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  • 201.
    Larsson, Ida
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Dalmo, Erika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology. Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala Univ, Dept Immunol Genet & Pathol, Uppsala, Sweden..
    Elgendy, Ramy
    Uppsala Univ, Dept Immunol Genet & Pathol, Uppsala, Sweden..
    Niklasson, Mia
    Uppsala Univ, Dept Immunol Genet & Pathol, Uppsala, Sweden..
    Doroszko, Milena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology. Uppsala Univ, Dept Immunol Genet & Pathol, Uppsala, Sweden..
    Segerman, Anna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology. Uppsala Univ, Dept Immunol Genet & Pathol, Uppsala, Sweden.;Uppsala Univ Hosp, Dept Med Sci Canc Pharmacol & Computat Med, Uppsala, Sweden..
    Jornsten, Rebecka
    Chalmers Univ Technol, Math Sci, Gothenburg, Sweden..
    Westermark, Bengt
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology.
    Nelander, Sven
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology.
    Modeling glioblastoma heterogeneity as a dynamic network of cell states2021In: Molecular Systems Biology, ISSN 1744-4292, E-ISSN 1744-4292, Vol. 17, no 9, article id e10105Article in journal (Refereed)
    Abstract [en]

    Tumor cell heterogeneity is a crucial characteristic of malignant brain tumors and underpins phenomena such as therapy resistance and tumor recurrence. Advances in single-cell analysis have enabled the delineation of distinct cellular states of brain tumor cells, but the time-dependent changes in such states remain poorly understood. Here, we construct quantitative models of the time-dependent transcriptional variation of patient-derived glioblastoma (GBM) cells. We build the models by sampling and profiling barcoded GBM cells and their progeny over the course of 3 weeks and by fitting a mathematical model to estimate changes in GBM cell states and their growth rates. Our model suggests a hierarchical yet plastic organization of GBM, where the rates and patterns of cell state switching are partly patient-specific. Therapeutic interventions produce complex dynamic effects, including inhibition of specific states and altered differentiation. Our method provides a general strategy to uncover time-dependent changes in cancer cells and offers a way to evaluate and predict how therapy affects cell state composition.

  • 202.
    Larsson, Ida
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Lundin, Erika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology. Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala Univ, Dept Immunol Genet & Pathol, Uppsala, Sweden..
    Elgendy, Ramy
    Uppsala Univ, Dept Immunol Genet & Pathol, Uppsala, Sweden..
    Niklasson, Mia
    Uppsala Univ, Dept Immunol Genet & Pathol, Uppsala, Sweden..
    Doroszko, Milena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology. Uppsala Univ, Dept Immunol Genet & Pathol, Uppsala, Sweden..
    Segerman, Anna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology. Uppsala Univ, Dept Immunol Genet & Pathol, Uppsala, Sweden.;Uppsala Univ Hosp, Dept Med Sci Canc Pharmacol & Computat Med, Uppsala, Sweden..
    Jornsten, Rebecka
    Chalmers Univ Technol, Math Sci, Gothenburg, Sweden..
    Westermark, Bengt
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology.
    Nelander, Sven
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology.
    Modeling glioblastoma heterogeneity as a dynamic network of cell states2021In: Molecular Systems Biology, ISSN 1744-4292, E-ISSN 1744-4292, Vol. 17, no 9, article id e10105Article in journal (Refereed)
    Abstract [en]

    Tumor cell heterogeneity is a crucial characteristic of malignant brain tumors and underpins phenomena such as therapy resistance and tumor recurrence. Advances in single-cell analysis have enabled the delineation of distinct cellular states of brain tumor cells, but the time-dependent changes in such states remain poorly understood. Here, we construct quantitative models of the time-dependent transcriptional variation of patient-derived glioblastoma (GBM) cells. We build the models by sampling and profiling barcoded GBM cells and their progeny over the course of 3 weeks and by fitting a mathematical model to estimate changes in GBM cell states and their growth rates. Our model suggests a hierarchical yet plastic organization of GBM, where the rates and patterns of cell state switching are partly patient-specific. Therapeutic interventions produce complex dynamic effects, including inhibition of specific states and altered differentiation. Our method provides a general strategy to uncover time-dependent changes in cancer cells and offers a way to evaluate and predict how therapy affects cell state composition.

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  • 203.
    Lau Börjesson, Joey
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical diabetology and metabolism.
    Vasylovska, Svitlana
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Kozlova, Elena N.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Carlsson, Per-Ola
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Transplantation and regenerative medicine.
    Surface Coating of Pancreatic Islets With Neural Crest Stem Cells Improves Engraftment and Function After Intraportal Transplantation2015In: Cell Transplantation, ISSN 0963-6897, E-ISSN 1555-3892, Vol. 24, no 11, p. 2263-2272Article in journal (Refereed)
    Abstract [en]

    The present study aimed to develop techniques for surface coating of islets with neural crest stem cells (NCSCs) in order to enable cotransplantation to the clinically used liver site and then investigate engraftment and function intraportally of such bioengineered islets. Mouse islets were coated during incubation with enhanced green fluorescent protein (EGFP)-expressing mouse NCSCs and transplanted into the portal vein to cure diabetic mice. An intravenous glucose tolerance test was performed at 1 month posttransplantation. Islet grafts were retrieved and evaluated for vascular density, nerves, and glial cells. NCSCs expressed a vast number of key angiogenic and neurotrophic factors. Mice transplanted with NCSC-bioengineered islets responded better to the glucose load than recipient mice with control islets. NCSCs remained present in the vicinity or had often migrated into the NCSC-coated islets, and an improved islet graft reinnervation and revascularization was observed. Transplanted NCSCs differentiated into both glial and neural cells in the islet grafts. We conclude that bioengineering of islets with NCSCs for intraportal transplantation provides a possibility to improve islet engraftment and function. Pending successful establishment of protocols for expansion of NCSCs from, for example, human skin or bone marrow, this strategy may be applied to clinical islet transplantation.

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  • 204.
    Laviña, Bàrbara
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology.
    Castro, Marco
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology.
    Niaudet, Colin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology.
    Bert, Cruys
    Peter, Carmeliet
    Bentley, Katie
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology. 4Computational Biology Laboratory, Center for Vascular Biology Research, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, USA.
    Cord, Brakebusch
    Betsholtz, Christer
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology.
    Gängel, Konstantin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology.
    Defective endothelial cell migration in the absence of Cdc42 leads to capillary-venous malformations: Cdc42 and vascular malformationsManuscript (preprint) (Other academic)
  • 205.
    Lees, John
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology. Uppsala Univ, Evolutionsbiol Centrum EBC, S-75236 Uppsala, Sweden..
    Pèrtille, Fábio
    Uppsala University, Disciplinary Domain of Science and Technology, Biology. Uppsala Univ, Evolutionsbiol Centrum EBC, S-75236 Uppsala, Sweden..
    Lotvedt, Pia
    Linköping Univ, Institutionen Fys, Kemi & Biol IFM, S-58330 Linköping, Sweden..
    Jensen, Per
    Linköping Univ, Institutionen Fys, Kemi & Biol IFM, S-58330 Linköping, Sweden..
    Guerrero-Bosagna, Carlos
    Uppsala University, Disciplinary Domain of Science and Technology, Biology.
    The mitoepigenome responds to stress, suggesting novel mito-nuclear interactions in vertebrates2023In: BMC Genomics, E-ISSN 1471-2164, Vol. 24, article id 561Article in journal (Refereed)
    Abstract [en]

    The mitochondria are central in the cellular response to changing environmental conditions resulting from disease states, environmental exposures or normal physiological processes. Although the influences of environmental stressors upon the nuclear epigenome are well characterized, the existence and role of the mitochondrial epigenome remains contentious. Here, by quantifying the mitochondrial epigenomic response of pineal gland cells to circadian stress, we confirm the presence of extensive cytosine methylation within the mitochondrial genome. Furthermore, we identify distinct epigenetically plastic regions (mtDMRs) which vary in cytosinic methylation, primarily in a non CpG context, in response to stress and in a sex-specific manner. Motifs enriched in mtDMRs contain recognition sites for nuclear-derived DNA-binding factors (ATF4, HNF4A) important in the cellular metabolic stress response, which we found to be conserved across diverse vertebrate taxa. Together, these findings suggest a new layer of mito-nuclear interaction in which the nuclear metabolic stress response could alter mitochondrial transcriptional dynamics through the binding of nuclear-derived transcription factors in a methylation-dependent context.

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  • 206.
    Lekholm, Emilia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Functional Pharmacology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Solute Carriers in Metabolism: Regulation of known and putative solute carriers in the central nervous system2017Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Solute carriers (SLCs) are membrane-bound transporter proteins, important for nutrient, ion, drug and metabolite transport across membranes. A quarter of the human genome codes for membrane-bound proteins, and SLCs make up the largest group of transporter proteins. Due to their ability to transport a large repertoire of substances across, not just the plasma membrane, but also the membrane of internal organelles, they hold a key position in maintaining homeostasis affecting metabolic pathways. Unfortunately, some of the more than 400 identified SLCs are still not fully characterized, even though a quarter of these are associated with human disease. In addition, there are about 30 membrane-bound proteins with strong resemblance to SLCs, of which very little is known. The aim of this thesis is to characterize some of these putative SLCs, focusing on their localization and function in the central nervous system. Since many of the known SLCs play a vital part in metabolism and related pathways, the response to different nutritional conditions has been used as a key method. MFSD14A and MFSD14B, characterized in Paper I, are putative SLCs belonging to the Major Facilitator Superfamily (MFS) and found to be neuronal, differentially expressed in the mouse central nervous system and transiently upregulated in mouse embryonic cortex cultures due to amino acid deprivation. They were also altered in areas of the mouse brain after starvation as well as after high fat diet. In Paper II, the effect on gene regulation due to complete amino acid starvation was monitored in a mouse hypothalamic cell line and 47 different genes belonging to SLCs, or putative SLCs, were found to be affected. Of these, 15 genes belonged to already known amino acid transporters, whereas 32 were putative SLCs with no known function or SLCs not known to react to amino acids. The three SV2 proteins, SV2A, SV2B and SV2C, were studied in Paper III using human neuroblastoma cell lines. The high metabolic state of cancers often result in an upregulation and alteration of transporter proteins, and alterations of the SV2 proteins were found following different treatments performed in this study. Paper IV focused on putative SLCs of MFS type and their role in glucose metabolism. Mouse embryonic cortex cultures were subjected to glucose starvation and the gene expression of 19 putative transporters were analyzed. All but four of the putative transporters were affected either at 3h or 12h of glucose deprivation. In conclusion, several SLCs and putative SLCs studied in this thesis are strongly affected by alteration in metabolism, either due to amino acids or glucose or both. This makes the putative SLCs dynamic membrane-bound proteins, possibly transporters, highly affected by nutritional status and most likely regulated to maintain homeostasis.

    List of papers
    1. Putative Membrane-Bound Transporters MFSD14A and MFSD14B Are Neuronal and Affected by Nutrient Availability
    Open this publication in new window or tab >>Putative Membrane-Bound Transporters MFSD14A and MFSD14B Are Neuronal and Affected by Nutrient Availability
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    2017 (English)In: Frontiers in Molecular Neuroscience, ISSN 1662-5099, Vol. 10, article id 11Article in journal (Refereed) Published
    Abstract [en]

    Characterization of orphan transporters is of importance due to their involvement in cellular homeostasis but also in pharmacokinetics and pharmacodynamics. The tissue and cellular localization, as well as function, is still unknown for many of the solute carriers belonging to the major facilitator superfamily (MFS) Pfam clan. Here, we have characterized two putative novel transporters MFSD14A (HIAT1) and MFSD14B (HIATL1) in the mouse central nervous system and found protein staining throughout the adult mouse brain. Both transporters localized to neurons and MFSD14A co-localized with the Golgi marker Giantin in primary embryonic cortex cultures, while MFSD14B staining co-localized with an endoplasmic retention marker, KDEL. Based on phylogenetic clustering analyses, we predict both to have organic substrate profiles, and possible involvement in energy homeostasis. Therefore, we monitored gene regulation changes in mouse embryonic primary cultures after amino acid starvations and found both transporters to be upregulated after 3 h of starvation. Interestingly, in mice subjected to 24 h of food starvation, both transporters were downregulated in the hypothalamus, while Mfsdl4a was also downregulated in the brainstem. In addition, in mice fed a high fat diet (HFD), upregulation of both transporters was seen in the striatum. Both MFSD14A and MFSD14B were intracellular neuronal membrane bound proteins, expressed in the Golgi and Endoplasmic reticulum, affected by both starvation and HFD to varying degree in the mouse brain.

    Keywords
    MFSD14A, HIAT1, MFSD14B, HIATL1, SLC, MFSD, transporter protein
    National Category
    Neurology
    Identifiers
    urn:nbn:se:uu:diva-316413 (URN)10.3389/fnmol.2017.00011 (DOI)000392751300001 ()28179877 (PubMedID)
    Funder
    Swedish Research CouncilThe Swedish Brain FoundationSwedish Society for Medical Research (SSMF)Novo NordiskMagnus Bergvall Foundation
    Available from: 2017-03-02 Created: 2017-03-02 Last updated: 2017-11-29Bibliographically approved
    2. The gene expression of numerous SLC transporters is altered in the immortalized hypothalamic cell line N25/2 following amino acid starvation
    Open this publication in new window or tab >>The gene expression of numerous SLC transporters is altered in the immortalized hypothalamic cell line N25/2 following amino acid starvation
    2017 (English)In: FEBS Open Bio, E-ISSN 2211-5463, Vol. 7, no 2, p. 249-264Article in journal (Refereed) Published
    Abstract [en]

    Amino acids are known to play a key role in gene expression regulation,and in mammalian cells, amino acid signaling is mainly mediated via twopathways, the mammalian target of rapamycin complex 1 (mTORC1) pathwayand the amino acid responsive (AAR) pathway. It is vital for cells tohave a system to sense amino acid levels, in order to control protein andamino acid synthesis and catabolism. Amino acid transporters are crucialin these pathways, due to both their sensing and transport functions. Inthis large-scale study, an immortalized mouse hypothalamic cell line (N25/2)was used to study the gene expression changes following 1, 2, 3, 5 or 16 hof amino acid starvation. We focused on genes encoding solute carriers(SLCs) and putative SLCs, more specifically on amino acid transporters.The microarray contained 28 270 genes and 86.2% of the genes wereexpressed in the cell line. At 5 h of starvation, 1001 genes were upregulatedand 848 genes were downregulated, and among these, 47 genes from theSLC superfamily or atypical SLCs were found. Of these, 15 were genesencoding amino acid transporters and 32 were genes encoding other SLCsor atypical SLCs. Increased expression was detected for genes encodingamino acid transporters from system A, ASC, L, N, T, xc-, and y+. UsingGO annotations, genes involved in amino acid transport and amino acidtransmembrane transporter activity were found to be most upregulated at3 h and 5 h of starvation.

    National Category
    Cell Biology
    Identifiers
    urn:nbn:se:uu:diva-331260 (URN)10.1002/2211-5463.12181 (DOI)000397220400011 ()28174690 (PubMedID)
    Funder
    Swedish Research CouncilNovo NordiskStiftelsen Olle Engkvist ByggmästareMagnus Bergvall Foundation
    Available from: 2017-10-12 Created: 2017-10-12 Last updated: 2018-09-07Bibliographically approved
    3. Treatment of two human neuroblastoma cell lines with growth factors, ATRA or VIP alters synaptic vesicle glycoprotein 2 (SV2) expression.
    Open this publication in new window or tab >>Treatment of two human neuroblastoma cell lines with growth factors, ATRA or VIP alters synaptic vesicle glycoprotein 2 (SV2) expression.
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    (English)Manuscript (preprint) (Other academic)
    National Category
    Biochemistry and Molecular Biology
    Identifiers
    urn:nbn:se:uu:diva-331276 (URN)
    Available from: 2017-10-12 Created: 2017-10-12 Last updated: 2017-10-12
    4. Glucose deprived mouse embryonal cortex cultures respond by altering MFS transporter expression and localization
    Open this publication in new window or tab >>Glucose deprived mouse embryonal cortex cultures respond by altering MFS transporter expression and localization
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    (English)Manuscript (preprint) (Other academic)
    National Category
    Biochemistry and Molecular Biology
    Identifiers
    urn:nbn:se:uu:diva-331327 (URN)
    Available from: 2017-10-12 Created: 2017-10-12 Last updated: 2017-10-12
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  • 207.
    Lendahl, Urban
    et al.
    Karolinska Inst, Dept Cell & Mol Biol, SE-17177 Stockholm, Sweden.;Karolinska Inst, Dept Neurobiol Care Sci & Soc, SE-14183 Huddinge, Sweden..
    Muhl, Lars
    Karolinska Inst, Dept Med, Blickagangen 16, SE-14157 Huddinge, Sweden..
    Betsholtz, Christer
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology. Uppsala University, Science for Life Laboratory, SciLifeLab. Karolinska Inst, Dept Med, Blickagangen 16, SE-14157 Huddinge, Sweden..
    Identification, discrimination and heterogeneity of fibroblasts2022In: Nature Communications, E-ISSN 2041-1723, Vol. 13, no 1, article id 3409Article, review/survey (Refereed)
    Abstract [en]

    Fibroblasts, the principal cell type of connective tissue, secrete extracellular matrix components during tissue development, homeostasis, repair and disease. Despite this crucial role, the identification and distinction of fibroblasts from other cell types are challenging and laden with caveats. Rapid progress in single-cell transcriptomics now yields detailed molecular portraits of fibroblasts and other cell types in our bodies, which complement and enrich classical histological and immunological descriptions, improve cell class definitions and guide further studies on the functional heterogeneity of cell subtypes and states, origins and fates in physiological and pathological processes. In this review, we summarize and discuss recent advances in the understanding of fibroblast identification and heterogeneity and how they discriminate from other cell types. In this review, the authors look at how recent progress in single-cell transcriptomics complement and enrich the classical, largely morphological, portraits of fibroblasts. The detailed molecular information now available provides new insights into fibroblast identity, heterogeneity and function.

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  • 208.
    Leuchowius, Karl Johan
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Weibrecht, Irene
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Söderberg, Ola
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular tools.
    In situ proximity ligation assay for microscopy and flow cytometry2011In: Current Protocols in Cytometry, ISSN 1934-9297, Vol. Suppl 56, p. Unit 9.36-Article in journal (Refereed)
    Abstract [en]

    The ability to study proteins and protein interactions inside cells and tissues is important for elucidating how cells function in health and disease. The in situ proximity ligation assay (in situ PLA) presented here can be used to visualize proteins, protein-protein interactions, and post-translational modifications in cells and tissues. The method is based upon the use of antibodies that target the proteins involved in an interaction; hence, the method has the advantage that it can be used in clinical specimens, providing localized, quantifiable single molecule detection in single cells. This unit describes how in situ PLA can be used with fluorescence microscopy and flow cytometry to study proteins (obtaining high sensitivity and specificity of detection) and protein interactions. It also includes information on expected results and information on how to troubleshoot the assay.

  • 209.
    Levine, Daniel C.
    et al.
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA..
    Kuo, Hsin-Yu
    Northwestern Univ, Dept Chem, Evanston, IL USA.;Northwestern Univ, Dept Biomed Engn, Evanston, IL USA.;Northwestern Univ, Dept Cell & Mol Biol, Evanston, IL USA..
    Hong, Hee-Kyung
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA..
    Cedernaes, Jonathan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Transplantation and regenerative medicine. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA.
    Hepler, Chelsea
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA..
    Wright, Alexandra G.
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA..
    Sommars, Meredith A.
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA..
    Kobayashi, Yumiko
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA..
    Marcheva, Biliana
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA..
    Gao, Peng
    Northwestern Univ, Feinberg Sch Med, Robert H Lurie Canc Ctr Metabol Core, Chicago, IL USA..
    Ilkayeva, Olga R.
    Duke Univ Sch Med, Dept Med, Div Endocrinol Metab & Nutr, Duke Mol Physiol Inst, Durham, NC USA..
    Omura, Chiaki
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA..
    Ramsey, Kathryn M.
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA..
    Newgard, Christopher B.
    Duke Univ Sch Med, Dept Med, Div Endocrinol Metab & Nutr, Duke Mol Physiol Inst, Durham, NC USA..
    Barish, Grant D.
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA..
    Peek, Clara Bien
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA.;Northwestern Univ, Feinberg Sch Med, Dept Biochem & Mol Genet, Chicago, IL USA..
    Chandel, Navdeep S.
    Northwestern Univ, Feinberg Sch Med, Dept Med, Chicago, IL USA..
    Mrksich, Milan
    Northwestern Univ, Dept Chem, Evanston, IL USA.;Northwestern Univ, Dept Biomed Engn, Evanston, IL USA.;Northwestern Univ, Dept Cell & Mol Biol, Evanston, IL USA..
    Bass, Joseph
    Northwestern Univ, Feinberg Sch Med, Div Endocrinol Metab & Mol Med, Dept Med, Chicago, IL 60208 USA..
    NADH inhibition of SIRT1 links energy state to transcription during time-restricted feeding2021In: Nature Metabolism, E-ISSN 2522-5812, Vol. 3, no 12, p. 1621-1632Article in journal (Refereed)
    Abstract [en]

    In mammals, circadian rhythms are entrained to the light cycle and drive daily oscillations in levels of NAD+, a cosubstrate of the class III histone deacetylase sirtuin 1 (SIRT1) that associates with clock transcription factors. Although NAD+ also participates in redox reactions, the extent to which NAD(H) couples nutrient state with circadian transcriptional cycles remains unknown. Here we show that nocturnal animals subjected to time-restricted feeding of a calorie-restricted diet (TRF-CR) only during night-time display reduced body temperature and elevated hepatic NADH during daytime. Genetic uncoupling of nutrient state from NADH redox state through transduction of the water-forming NADH oxidase from Lactobacillus brevis (LbNOX) increases daytime body temperature and blood and liver acyl-carnitines. LbNOX expression in TRF-CR mice induces oxidative gene networks controlled by brain and muscle Arnt-like protein 1 (BMAL1) and peroxisome proliferator-activated receptor alpha (PPARα) and suppresses amino acid catabolic pathways. Enzymatic analyses reveal that NADH inhibits SIRT1 in vitro, corresponding with reduced deacetylation of SIRT1 substrates during TRF-CR in vivo. Remarkably, Sirt1 liver nullizygous animals subjected to TRF-CR display persistent hypothermia even when NADH is oxidized by LbNOX. Our findings reveal that the hepatic NADH cycle links nutrient state to whole-body energetics through the rhythmic regulation of SIRT1.

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  • 210.
    Li, Hao
    et al.
    Uppsala Univ Hosp, Dept Surg Sci, Sect Otolaryngol, Head & Neck Surg, Uppsala, Sweden..
    Staxäng, Karin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Hodik, Monika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Melkersson, Karl-Gunnar
    Kolmardens Tropicarium AB, Kolmarden, Sweden..
    Rask-Andersen, Mathias
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medicinsk genetik och genomik. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Rask-Andersen, Helge
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Otolaryngology and Head and Neck Surgery.
    Regeneration in the Auditory Organ in Cuban and African Dwarf Crocodiles (Crocodylus rhombifer and Osteolaemus tetraspis) Can We Learn From the Crocodile How to Restore Our Hearing?2022In: Frontiers in Cell and Developmental Biology, E-ISSN 2296-634X, Vol. 10, article id 934571Article in journal (Refereed)
    Abstract [en]

    Background: In several non-mammalian species, auditory receptors undergo cell renewal after damage. This has raised hope of finding new options to treat human sensorineural deafness. Uncertainty remains as to the triggering mechanisms and whether hair cells are regenerated even under normal conditions. In the present investigation, we explored the auditory organ in the crocodile to validate possible ongoing natural hair cell regeneration. Materials and Methods: Two male Cuban crocodiles (Crocodylus rhombifer) and an adult male African Dwarf crocodile (Osteolaemus tetraspis) were analyzed using transmission electron microscopy and immunohistochemistry using confocal microscopy. The crocodile ears were fixed in formaldehyde and glutaraldehyde and underwent micro-computed tomography (micro-CT) and 3D reconstruction. The temporal bones were drilled out and decalcified. Results: The crocodile papilla basilaris contained tall (inner) and short (outer) hair cells surrounded by a mosaic of tightly connected supporting cells coupled with gap junctions. Afferent neurons with and without ribbon synapses innervated both hair cell types. Supporting cells occasionally showed signs of trans-differentiation into hair cells. They expressed the MAFA and SOX2 transcription factors. Supporting cells contained organelles that may transfer genetic information between cells, including the efferent nerve fibers during the regeneration process. The tectorial membrane showed signs of being replenished and its architecture being sculpted by extracellular exosome-like proteolysis. Discussion: Crocodilians seem to produce new hair cells during their life span from a range of supporting cells. Imposing efferent nerve fibers may play a role in regeneration and re-innervation of the auditory receptors, possibly triggered by apoptotic signals from wasted hair cells. Intercellular signaling may be accomplished by elaborate gap junction and organelle systems, including neural emperipolesis. Crocodilians seem to restore and sculpt their tectorial membranes throughout their lives.

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  • 211.
    Li, Jia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    ATP Dynamics in Pancreatic α- and β-cells2014Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Glucose metabolism in pancreatic α- and β-cells is believed to regulate secretion of glucagon and insulin, respectively. In β-cells, ATP links glucose metabolism to electrical activity and insulin secretion. In α-cells, ATP has been attributed various roles in glucose-regulated glucagon release, but the underlying mechanisms are poorly understood. Despite its importance in insulin and glucagon secretion little is known about ATP kinetics in α- and β-cells. In this thesis, the novel fluorescent ATP biosensor Perceval was used to monitor physiologically relevant ATP concentrations with little influence of ADP. Glucose stimulation of β-cells within mouse and human pancreatic islets induced pronounced rise of ATP with superimposed oscillations. Simultaneous measurements of the sub-plasma membrane ATP and Ca2+ concentrations revealed glucose-induced oscillations in opposite phase. ATP increased further and the oscillations ceased when voltage-dependent Ca2+ influx was prevented. In contrast, ATP promptly decreased in response to K+-depolarization-induced elevation of Ca2+. Also mobilization of Ca2+ from intracellular stores lowered ATP, but the negative effect was not due to increased ATP consumption by the sarco/endoplasmic reticulum Ca2+-ATPase. Store-operated Ca2+ entry alone had little effect but markedly elevated ATP when combined with muscarinic receptor activation. When comparing ATP and Ca2+ responses in α- and β-cells within the same islet, glucose-induced ATP generation was much less pronounced and the dose-response relationship left-shifted in the α-cells. At basal glucose, individual α-cells showed Ca2+ and concomitant ATP oscillations in opposite-phase with variable frequency. These oscillations largely cancelled out when averaging data from several α-cells. At high glucose, the Ca2+ and ATP oscillations in α-cells tended to synchronize with the corresponding β-cell oscillations. Since β-cell Ca2+ oscillations drive pulsatile insulin secretion, which is antiparallel to pulsatile glucagon secretion, there seems to be an inverse relationship between changes in α-cell Ca2+ and glucagon release. This paradox is attributed to paracrine inhibition overriding Ca2+ stimulation, since somatostatin receptor blockade potently stimulated glucagon release with little effect on α-cell Ca2+ signalling. The data indicate that complex ATP-Ca2+ interactions in α- and β-cells underlie cell-intrinsic regulation of glucagon and insulin secretion and that paracrine inhibition of glucagon release becomes important in hyperglycaemia.

    List of papers
    1. Oscillations of sub-membrane ATP in glucose-stimulated beta cells depend on negative feedback from Ca2+
    Open this publication in new window or tab >>Oscillations of sub-membrane ATP in glucose-stimulated beta cells depend on negative feedback from Ca2+
    2013 (English)In: Diabetologia, ISSN 0012-186X, E-ISSN 1432-0428, Vol. 56, no 7, p. 1577-1586Article in journal (Refereed) Published
    Abstract [en]

    ATP links changes in glucose metabolism to electrical activity, Ca2+ signalling and insulin secretion in pancreatic beta cells. There is evidence that beta cell metabolism oscillates, but little is known about ATP dynamics at the plasma membrane, where regulation of ion channels and exocytosis occur. The sub-plasma-membrane ATP concentration ([ATP](pm)) was recorded in beta cells in intact mouse and human islets using total internal reflection microscopy and the fluorescent reporter Perceval. Glucose dose-dependently increased [ATP](pm) with half-maximal and maximal effects at 5.2 and 9 mmol/l, respectively. Additional elevations of glucose to 11 to 20 mmol/l promoted pronounced [ATP](pm) oscillations that were synchronised between neighbouring beta cells. [ATP](pm) increased further and the oscillations disappeared when voltage-dependent Ca2+ influx was prevented. In contrast, K+-depolarisation induced prompt lowering of [ATP](pm). Simultaneous recordings of [ATP](pm) and the sub-plasma-membrane Ca2+ concentration ([Ca2+](pm)) during the early glucose-induced response revealed that the initial [ATP](pm) elevation preceded, and was temporarily interrupted by the rise of [Ca2+](pm). During subsequent glucose-induced oscillations, the increases of [Ca2+](pm) correlated with lowering of [ATP](pm). In beta cells, glucose promotes pronounced oscillations of [ATP](pm), which depend on negative feedback from Ca2+ (.) The bidirectional interplay between these messengers in the sub-membrane space generates the metabolic and ionic oscillations that underlie pulsatile insulin secretion.

    Keywords
    ATP, Ca2+, Human islets, Mouse islets, Oscillations, Pancreatic beta cell, Perceval, Plasma membrane
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-203521 (URN)10.1007/s00125-013-2894-0 (DOI)000319881300015 ()
    Available from: 2013-07-16 Created: 2013-07-15 Last updated: 2017-12-06Bibliographically approved
    2. Dual effects of muscarinic receptor activation on intracellular ATP in pancreatic ß-cells
    Open this publication in new window or tab >>Dual effects of muscarinic receptor activation on intracellular ATP in pancreatic ß-cells
    (English)Manuscript (preprint) (Other academic)
    National Category
    Cell and Molecular Biology
    Research subject
    Medical Cell Biology
    Identifiers
    urn:nbn:se:uu:diva-234963 (URN)
    Available from: 2014-10-27 Created: 2014-10-27 Last updated: 2018-01-11
    3. Sub-membrane ATP and Ca2+ kinetics in α-cells – unexpectedsignalling for glucagon secretion
    Open this publication in new window or tab >>Sub-membrane ATP and Ca2+ kinetics in α-cells – unexpectedsignalling for glucagon secretion
    Show others...
    (English)Manuscript (preprint) (Other (popular science, discussion, etc.))
    National Category
    Cell and Molecular Biology
    Research subject
    Medical Cell Biology
    Identifiers
    urn:nbn:se:uu:diva-234826 (URN)
    Available from: 2014-10-23 Created: 2014-10-23 Last updated: 2018-01-11
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  • 212.
    Li, Jiaxin
    et al.
    Lund Univ, Stem Cell Ctr, Lund, Sweden.;Lund Univ, Dept Clin Sci, Div Neurosurg, Lund, Sweden..
    Ek, Fredrik
    Lund Univ, Dept Expt Med Sci, Chem Biol & Therapeut, Lund, Sweden..
    Olsson, Roger
    Lund Univ, Dept Expt Med Sci, Chem Biol & Therapeut, Lund, Sweden..
    Belting, Mattias
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Neuro-Oncology. Uppsala University, Science for Life Laboratory, SciLifeLab. Lund Univ, Sect Oncol, Dept Clin Sci, Lund, Sweden.;Skane Univ Hosp, Dept Hematol Oncol & Radiophys, Lund, Sweden..
    Bengzon, Johan
    Lund Univ, Stem Cell Ctr, Lund, Sweden.;Lund Univ, Dept Clin Sci, Div Neurosurg, Lund, Sweden.;Skane Univ Hosp, Dept Neurosurg, Lund, Sweden..
    Glioblastoma CD105(+) cells define a SOX2(-) cancer stem cell-like subpopulation in the pre-invasive niche2022In: Acta neuropathologica communications, E-ISSN 2051-5960, Vol. 10, article id 126Article in journal (Refereed)
    Abstract [en]

    Glioblastoma (GBM) is the most common and most aggressive primary brain tumor in adults. Glioma stem like cells (GSC) represent the highest cellular hierarchy in GBM and have a determining role in tumor growth, recurrence and patient prognosis. However, a better definition of GSC subpopulations, especially at the surgical resection margin, is warranted for improved oncological treatment options. The present study interrogated cells expressing CD105 (CD105(+)) specifically within the tumor front and the pre-invasive niche as a potential GSC subpopulation. GBM primary cell lines were generated from patients (n = 18) and CD105(+) cells were isolated and assessed for stem-like characteristics. In vitro, CD105(+) cells proliferated and enriched in serum-containing medium but not in serum-free conditions. CD105(+) cells were characterized by Nestin(+), Vimentin(+) and SOX2(-), clearly distinguishing them from SOX2(+) GCS. GBM CD105(+) cells differentiated into osteocytes and adipocytes but not chondrocytes. Exome sequencing revealed that GBM CD105(+) cells matched 83% of somatic mutations in the Cancer cell line encyclopedia, indicating a malignant phenotype and in vivo xenotransplantation assays verified their tumorigenic potential. Cytokine assays showed that immunosuppressive and protumorigenic cytokines such as IL6, IL8, CCL2, CXCL-1 were produced by CD105(+) cells. Finally, screening for 88 clinical drugs revealed that GBM CD105(+) cells are resistant to most chemotherapeutics except Doxorubicin, Idarubicin, Fludarabine and ABT-751. Our study provides a rationale for targeting tumoral CD105(+) cells in order to reshape the tumor microenvironment and block GBM progression.

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  • 213.
    Li, Wen
    et al.
    Columbia Univ, Dept Biochem & Mol Biophys, 2-221 Blackwell,165 West 168th St, New York, NY 10032 USA..
    Liu, Zheng
    Columbia Univ, Dept Biochem & Mol Biophys, 2-221 Blackwell,165 West 168th St, New York, NY 10032 USA..
    Koripella, Ravi Kiran
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Langlois, Robert
    Columbia Univ, Dept Biochem & Mol Biophys, 2-221 Blackwell,165 West 168th St, New York, NY 10032 USA..
    Sanyal, Suparna
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Frank, Joachim
    Columbia Univ, Dept Biochem & Mol Biophys, 2-221 Blackwell,165 West 168th St, New York, NY 10032 USA.;Columbia Univ, Howard Hughes Med Inst, New York, NY 10032 USA.;Columbia Univ, Dept Biol Sci, New York, NY 10027 USA..
    Activation of GTP hydrolysis in mRNA-tRNA translocation by elongation factor G2015In: Science Advances, E-ISSN 2375-2548, Vol. 1, no 4, article id e1500169Article in journal (Refereed)
    Abstract [en]

    During protein synthesis, elongation of the polypeptide chain by each amino acid is followed by a translocation step in which mRNA and transfer RNA (tRNA) are advanced by one codon. This crucial step is catalyzed by elongation factor G (EF-G), a guanosine triphosphatase (GTPase), and accompanied by a rotation between the two ribosomal subunits. A mutant of EF-G, H91A, renders the factor impaired in guanosine triphosphate (GTP) hydrolysis and thereby stabilizes it on the ribosome. We use cryogenic electron microscopy (cryo-EM) at near-atomic resolution to investigate two complexes formed by EF-G H91A in its GTP state with the ribosome, distinguished by the presence or absence of the intersubunit rotation. Comparison of these two structures argues in favor of a direct role of the conserved histidine in the switch II loop of EF-G in GTPase activation, and explains why GTP hydrolysis cannot proceed with EF-G bound to the unrotated form of the ribosome.

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  • 214.
    Li, Zhifei
    et al.
    Peking Univ, Sch Life Sci, Peking Tsinghua Joint Ctr Life Sci, State Key Lab Membrane Biol, Beijing, Peoples R China; Tsinghua Univ, Beijing Adv Innovat Ctr Struct Biol, Sch Life Sci, Tsinghua Peking Joint Ctr Life Sci, Beijing, Peoples R China.
    Ge, Xueliang
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Zhang, Yixiao
    Peking Univ, Sch Life Sci, Peking Tsinghua Joint Ctr Life Sci, State Key Lab Membrane Biol, Beijing, Peoples R China.
    Zheng, Lvqin
    Peking Univ, Sch Life Sci, Peking Tsinghua Joint Ctr Life Sci, State Key Lab Membrane Biol, Beijing, Peoples R China.
    Sanyal, Suparna
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Gao, Ning
    Peking Univ, Sch Life Sci, Peking Tsinghua Joint Ctr Life Sci, State Key Lab Membrane Biol, Beijing, Peoples R China; Tsinghua Univ, Beijing Adv Innovat Ctr Struct Biol, Sch Life Sci, Tsinghua Peking Joint Ctr Life Sci, Beijing, Peoples R China.
    Cryo-EM structure of Mycobacterium smegmatis ribosome reveals two unidentified ribosomal proteins close to the functional centers2018In: Protein & Cell, ISSN 1674-800X, E-ISSN 1674-8018, Vol. 9, no 4, p. 384-388Article in journal (Other academic)
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  • 215. Liao, Zhen
    et al.
    Kjellin, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Microbiology.
    Fransson, Åsa
    Söderbom, Fredrik
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Microbiology.
    Functional analyses of RNA interference effectors in Dictyostelium discoideum during growth and developmentManuscript (preprint) (Other academic)
    Abstract [en]

    RNA interference (RNAi) is a widespread biological process, which regulates gene expression in eukaryotic cells. A complex of proteins and small RNAs, RISC, mediates this gene regulation. Central to the function of RISC are the Argonaute effector proteins, which bind the small RNAs, e.g. micro (mi)RNAs and small interfering (si)RNAs. It has previously been shown that RNAi is important to control transposon mobilization in the social amoeba Dictyostelium discoideum, a unicellular eukaryote that upon starvation enters a multicellular developmental program. Information concerning the five Argonautes in D. discoideum is scarce but several of them appear to inhibit transposon mobilization by RNAi related mechanisms. In a recent study, we showed that three of the Argonautes in D. discoideum are involved in controlling cell division. In this study, we perform mRNA- and small RNA-seq. from growing and developing cells, combined with phenotypic studies of D. discoideum strains depleted of AgnB, AgnC, and AgnE. The previously observed effect on cell division, i.e. faster growth for agnE-, and slower for agnB- and agnC- cells, is associated with increased and decreased expression, respectively, of genes involved in nucleotide metabolism. Furthermore, all three argonautes appear to be involved in downregulation of ribosomal protein genes during development while AgnE also contributes to reduced expression of protein coding genes during growth. These effects are likely mediated by small RNAs. We further report the subcellular localization of the three Argonautes, where AgnB is mainly localized in the cytoplasm, AgnC in the nucleus, and the previously reported cytoplasmic localization for AgnE was confirmed. Finally, we present data indicating that AgnB is interacting with miRNAs, suggesting that this Argonaute is involved in miRNA mediated gene regulation in D. discoideum.  Taken together, our data indicate that none of the Argonautes components are essential for cell division and development, but all participate in fine-tuning of gene expression for optimal growth and synchronous multicellular development.

  • 216.
    Lin, Yingbo
    et al.
    Karolinska Inst, Dept Pathol & Oncol, CCK R8 04, Stockholm, Sweden..
    Liu, Hongyu
    Karolinska Inst, Dept Pathol & Oncol, CCK R8 04, Stockholm, Sweden.;Guangdong Ocean Univ, Fisheries Coll, Lab Aquat Anim Nutr Feed, Zhanjiang, Peoples R China..
    Waraky, Ahmed
    Karolinska Inst, Dept Pathol & Oncol, CCK R8 04, Stockholm, Sweden..
    Haglund, Felix
    Karolinska Inst, Dept Pathol & Oncol, CCK R8 04, Stockholm, Sweden..
    Agarwal, Prasoon
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Experimental and Clinical Oncology. Karolinska Univ Hosp, Div Clin Immunol, Dept Lab Med LABMED H5, Stockholm, Sweden.
    Jernberg Wiklund, Helena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Experimental and Clinical Oncology.
    Warsito, Dudi
    Karolinska Inst, Dept Pathol & Oncol, CCK R8 04, Stockholm, Sweden..
    Larsson, Olle
    Karolinska Inst, Dept Pathol & Oncol, CCK R8 04, Stockholm, Sweden..
    SUMO-modified insulin-like growth factor 1 receptor (IGF-1R) increases cell cycle progression and cell proliferation2017In: Journal of Cellular Physiology, ISSN 0021-9541, E-ISSN 1097-4652, Vol. 232, no 10, p. 2722-2730Article in journal (Refereed)
    Abstract [en]

    Increasing number of studies have shown nuclear localization of the insulin-like growth factor 1 receptor (nIGF-1R) in tumor cells and its links to adverse clinical outcome in various cancers. Any obvious cell physiological roles of nIGF-1R have, however, still not been disclosed. Previously, we reported that IGF-1R translocates to cell nucleus and modulates gene expression by binding to enhancers, provided that the receptor is SUMOylated. In this study, we constructed stable transfectants of wild type IGF1R (WT) and triple-SUMO-site-mutated IGF1R (TSM) using igf1r knockout mouse fibroblasts (R-). Cell clones (R-WT and R-TSM) expressing equal amounts of IGF1R were selected for experiments. Phosphorylation of IGF-1R, Akt, and Erk upon IGF-1 stimulation was equal in R-WT and R-TSM. WT was confirmed to enter nuclei. TSM did also undergo nuclear translocation, although to a lesser extent. This may be explained by that TSM heterodimerizes with insulin receptor, which is known to translocate to cell nuclei. R-WT proliferated substantially faster than R-TSM, which did not differ significantly from the empty vector control. Upon IGF-1 stimulationG1-S-phase progression of R-WT increased from 12 to 38%, compared to 13 to 20% of R-TSM. The G1-S progression of R-WT correlated with increased expression of cyclin D1, A, and CDK2, as well as downregulation of p27. This suggests that SUMO-IGF-1R affects upstream mechanisms that control and coordinate expression of cell cycle regulators. Further studies to identify such SUMO-IGF-1R dependent mechanisms seem important.

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  • 217.
    Lindvall, Jenny
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Green and red fluorescent protein tagging of endogenous proteins in glioblastoma using the CRISPR (clustered regularly interspaced short palindromic repeats)-Cas9 system2016Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    Glioblastoma multiforme is the most malignant primary brain tumor that affects adults, recognized by the World Health Organization as an aggressive grade IV astrocytoma. Patients diagnosed with this type of tumor are left with a poor prognosis even with the most advanced treatment available. The cancer is quite heterogeneous and is typically categorized into four different subtypes depending on genetic aberrations and patient characteristics. Furthermore, researchers have discovered a subpopulation of glioblastoma cells, known as cancer stem cells, which are thought to be resistant to current therapies and responsible for tumor reoccurrence and relapse. Previous studies, in addition to this one, have found that the differentiation of glioblastoma cells downregulate nestin protein expression, the selected stem cell marker, and upregulate glial fibrillary acid protein expression, the selected differentiation marker, using immunofluorescence. Thus, one alternative treatment option is to understand the mechanism underlying the differentiation of cancer stem cells. Four cell cultures representative of each glioblastoma subtype will be endogenously tagged using the genome editing system, Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR/Cas9). The representative stem cell marker, nestin, will be tagged with a green fluorescent protein, while the chosen differentiation marker, glial fibrillary acid protein, will be tagged with a red fluorescent protein. Several drugs were screened to analyze whether the drugs had a differentiation effect on the glioblastoma cells. As a result, strong evidence indicated that bone morphogenetic protein four upregulated glial fibrillary acid protein expression levels to the same extent as the differentiation control media using 5% fetal bovine serum. The goal of this study is to establish a method to directly monitor the differentiation process of glioblastoma cells as a novel molecular screening method. In this case, all glioblastoma cells, even the ones resistant to treatment, can be eliminated through an initial “pre-treatment” by forcing differentiation of cancer stem cells, making the cells more susceptible to the chemotherapy drugs. In the long run, glioblastoma patients would have a chance at a more positive prognosis; a longer life that is free of glioblastoma.

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    Lindvall.Jenny_MasterThesis2016
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    Lindvall.Jenny_PopularScienceArticle2016
  • 218. Lisowska, Halina
    et al.
    Deperas-Kaminska, Marta
    Haghdoost, Siamak
    Stockholms universitet, Institutionen för genetik, mikrobiologi och toxikologi.
    Parmryd, Ingela
    Stockholms universitet, Avdelningen för cellbiologi.
    Wojcik, Andrzej
    Stockholms universitet, Institutionen för genetik, mikrobiologi och toxikologi.
    Radiation-induced DNA damage and repair in human gammadelta and alphabeta T-lymphocytes analysed by the alkaline comet assay2010In: Genome integrity, ISSN 2041-9414, Vol. 1, no 1, p. 8-Article in journal (Refereed)
    Abstract [en]

    It has been shown by a number of authors that the radiosensitivity of peripheral blood mononuclear cells (PBMC) is higher in cancer patients compared to healthy donors, which is interpreted as a sign of genomic instability. PBMC are composed of different cell subpopulations which are differently radiosensitive and the difference between cancer patients and healthy donors could also be due to different composition of their PBMC pools. Gamma-delta T-lymphocytes play an important role in immunosurveillance and are promising cells for immunotherapy. Their abundance is frequently reduced in cancer patients so should their sensitivity to radiation be lower than that of other T-lymphocytes, this could, at least partly explain the low radiosensitivity of PBMC from healthy individuals compared to cancer patients. The present investigation was carried out to test this. Using the alkaline comet assay we analysed the level of DNA damage and repair in isolated gammadelta T-lymphocytes, pan T-lymphocytes and in total PBMC exposed in vitro to gamma radiation. We found no difference in the level of DNA damage and the capacity of DNA repair between the T cell populations. This is the first study that addresses the question of sensitivity to radiation of gamma-delta T-cells.

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    Radiation-induced DNA damage and repair in human gammadelta and alphabeta T-lymphocytes analysed by the alkaline comet assay
  • 219.
    Liu, Chen
    et al.
    Univ Crete, Dept Biol, Iraklion, Greece.;Fdn Res & Technol Hellas, Inst Mol Biol & Biotechnol, Iraklion, Greece.;Swedish Univ Agr Sci, Dept Plant Biol, Uppsala BioCtr, Uppsala, Sweden..
    Mentzelopoulou, Andriani
    Univ Crete, Dept Biol, Iraklion, Greece.;Fdn Res & Technol Hellas, Inst Mol Biol & Biotechnol, Iraklion, Greece..
    Papagavriil, Fotini
    Univ Crete, Dept Biol, Iraklion, Greece.;Fdn Res & Technol Hellas, Inst Mol Biol & Biotechnol, Iraklion, Greece..
    Ramachandran, Prashanth
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Organismal Biology.
    Perraki, Artemis
    Fdn Res & Technol Hellas, Inst Mol Biol & Biotechnol, Iraklion, Greece..
    Claus, Lucas
    Univ Ghent, Dept Plant Biotechnol & Bioinformat, Ghent, Belgium.;Ctr Plant Syst Biol, Ghent, Belgium..
    Barg, Sebastian
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Dörmann, Peter
    Univ Bonn, Inst Mol Physiol & Biotechnol Plants, Bonn, Germany..
    Jaillais, Yvon
    Univ Claude Bernard Lyon 1, CNRS, Lab Reprod & Dev Plantes, INRAE,ENS Lyon, Lyon, France..
    Johnen, Philipp
    Univ Bonn, Inst Crop Sci & Resource Conservat, Dept Plant Nutr, Bonn, Germany..
    Russinova, Eugenia
    Univ Ghent, Dept Plant Biotechnol & Bioinformat, Ghent, Belgium.;Ctr Plant Syst Biol, Ghent, Belgium..
    Gizeli, Electra
    Univ Crete, Dept Biol, Iraklion, Greece.;Fdn Res & Technol Hellas, Inst Mol Biol & Biotechnol, Iraklion, Greece..
    Schaaf, Gabriel
    Univ Bonn, Inst Crop Sci & Resource Conservat, Dept Plant Nutr, Bonn, Germany..
    Moschou, Panagiotis Nikolaou
    Univ Crete, Dept Biol, Iraklion, Greece.;Fdn Res & Technol Hellas, Inst Mol Biol & Biotechnol, Iraklion, Greece.;Swedish Univ Agr Sci, Dept Plant Biol, Uppsala BioCtr, Uppsala, Sweden..
    SEC14-like condensate phase transitions at plasma membranes regulate root growth in Arabidopsis2023In: PLoS biology, ISSN 1544-9173, E-ISSN 1545-7885, Vol. 21, no 9, article id e3002305Article in journal (Refereed)
    Abstract [en]

    Protein function can be modulated by phase transitions in their material properties, which can range from liquid- to solid-like; yet, the mechanisms that drive these transitions and whether they are important for physiology are still unknown. In the model plant Arabidopsis, we show that developmental robustness is reinforced by phase transitions of the plasma membrane-bound lipid-binding protein SEC14-like. Using imaging, genetics, and in vitro reconstitution experiments, we show that SEC14-like undergoes liquid-like phase separation in the root stem cells. Outside the stem cell niche, SEC14-like associates with the caspase-like protease separase and conserved microtubule motors at unique polar plasma membrane interfaces. In these interfaces, SEC14-like undergoes processing by separase, which promotes its liquid-to-solid transition. This transition is important for root development, as lines expressing an uncleavable SEC14-like variant or mutants of separase and associated microtubule motors show similar developmental phenotypes. Furthermore, the processed and solidified but not the liquid form of SEC14-like interacts with and regulates the polarity of the auxin efflux carrier PINFORMED2. This work demonstrates that robust development can involve liquid-to-solid transitions mediated by proteolysis at unique plasma membrane interfaces.

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  • 220.
    Liu, Wei
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Otolaryngology and Head and Neck Surgery.
    Loewenheim, Hubert
    Eberhard Karls Univ Tubingen, Tubingen Hearing Res Ctr, Dept Otolaryngol Head & Neck Surg, D-72076 Tubingen, Germany.
    Santi, Peter A.
    Univ Minnesota, Dept Otolaryngol, 121 Lions Res Bldg,2001 Sixth St SE, Minneapolis, MN 55455 USA.
    Glueckert, Rudolf
    Med Univ Innsbruck, Dept Otolaryngol, Anichstr 35, A-6020 Innsbruck, Austria.
    Schrott-Fischer, Annelies
    Med Univ Innsbruck, Dept Otolaryngol, Anichstr 35, A-6020 Innsbruck, Austria.
    Rask-Andersen, Helge
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Otolaryngology and Head and Neck Surgery.
    Expression of trans-membrane serine protease 3 (TMPRSS3) in the human organ of Corti2018In: Cell and Tissue Research, ISSN 0302-766X, E-ISSN 1432-0878, Vol. 372, no 3, p. 445-456Article in journal (Refereed)
    Abstract [en]

    TMPRSS3 (Trans-membrane Serine Protease 3) is a type II trans-membrane serine protease that has proteolytic activity essential for hearing. Mutations in the gene cause non-syndromic autosomal recessive deafness (DFNB8/10) in humans. Knowledge about its cellular distribution in the human inner ear may increase our understanding of its physiological role and involvement in deafness, ultimately leading to therapeutic interventions. In this study, we used super-resolution structured illumination microscopy for the first time together with transmission electron microscopy to localize the TMPRSS3 protein in the human organ of Corti. Archival human cochleae were dissected out during petroclival meningioma surgery. Microscopy with Zeiss LSM710 microscope achieved a lateral resolution of approximately 80 nm. TMPRSS3 was found to be associated with actin in both inner and outer hair cells. TMPRSS3 was located in cell surface-associated cytoskeletal bodies (surfoskelosomes) in inner and outer pillar cells and Deiters cells and in subcuticular organelles in outer hair cells. Our results suggest that TMPRSS3 proteolysis is linked to hair cell sterociliary mechanics and to the actin/microtubule networks that support cell motility and integrity.

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  • 221.
    Looman, Camilla
    et al.
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology. Molekylär Immunologi.
    Åbrink, Magnus
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology. Molekylär Immunologi.
    Mark, Charlotta
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology. Molekylär Immunologi.
    Hellman, Lars
    Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology. Uppsala University, Teknisk-naturvetenskapliga vetenskapsområdet, Faculty of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Immunology. Molekylär Immunologi.
    MZF6D, a novel KRAB zinc finger gene expressed exclusively in meiotic male germ cells.2003In: DNA and Cell Biology, Vol. 22, p. 489-496Article in journal (Refereed)
  • 222.
    Lopes, Ricardo J.
    et al.
    Univ Porto, CIBIO InBIO, Ctr Invest Biodiversidade & Recursos Genet, Campus Agr Vairao, P-4485661 Vairao, Portugal..
    Johnson, James D.
    Auburn Univ, Dept Biol Sci, Auburn, AL 36849 USA..
    Toomey, Matthew B.
    Washington Univ, Sch Med, Dept Pathol & Immunol, St Louis, MO 63110 USA..
    Ferreira, Mafalda S.
    Univ Porto, CIBIO InBIO, Ctr Invest Biodiversidade & Recursos Genet, Campus Agr Vairao, P-4485661 Vairao, Portugal..
    Araujo, Pedro M.
    Univ Porto, CIBIO InBIO, Ctr Invest Biodiversidade & Recursos Genet, Campus Agr Vairao, P-4485661 Vairao, Portugal.;Univ Coimbra, Dept Life Sci, Marine & Environm Sci Ctr MARE, P-3004517 Coimbra, Portugal..
    Melo-Ferreira, Jose
    Univ Porto, CIBIO InBIO, Ctr Invest Biodiversidade & Recursos Genet, Campus Agr Vairao, P-4485661 Vairao, Portugal.;Univ Porto, Fac Ciencias, Dept Biol, Rua Campo Alegre S-N, P-4169007 Oporto, Portugal..
    Andersson, Leif
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Swedish Univ Agr Sci, Dept Anim Breeding & Genet, S-75007 Uppsala, Sweden.;Texas A&M Univ, Coll Vet Med & Biomed Sci, Dept Vet Integrat Biosci, College Stn, TX 77845 USA..
    Hill, Geoffrey E.
    Auburn Univ, Dept Biol Sci, Auburn, AL 36849 USA..
    Corbo, Joseph C.
    Washington Univ, Sch Med, Dept Pathol & Immunol, St Louis, MO 63110 USA..
    Carneiro, Miguel
    Univ Porto, CIBIO InBIO, Ctr Invest Biodiversidade & Recursos Genet, Campus Agr Vairao, P-4485661 Vairao, Portugal.;Univ Porto, Fac Ciencias, Dept Biol, Rua Campo Alegre S-N, P-4169007 Oporto, Portugal..
    Genetic Basis for Red Coloration in Birds2016In: Current Biology, ISSN 0960-9822, E-ISSN 1879-0445, Vol. 26, no 11, p. 1427-1434Article in journal (Refereed)
    Abstract [en]

    The yellow and red feather pigmentation of many bird species [1] plays pivotal roles in social signaling and mate choice [2, 3]. To produce red pigments, birds ingest yellow carotenoids and endogenously convert them into red ketocarotenoids via an oxidation reaction catalyzed by a previously unknown ketolase [4-6]. We investigated the genetic basis for red coloration in birds using whole-genome sequencing of red siskins (Spinus cucullata), common canaries (Serinus canaria), and "red factor" canaries, which are the hybrid product of crossing red siskins with common canaries [7]. We identified two genomic regions introgressed from red siskins into red factor canaries that are required for red coloration. One of these regions contains a gene encoding a cytochrome P450 enzyme, CYP2J19. Transcriptome analysis demonstrates that CYP2J19 is significantly upregulated in the skin and liver of red factor canaries, strongly implicating CYP2J19 as the ketolase that mediates red coloration in birds. Interestingly, a second introgressed region required for red feathers resides within the epidermal differentiation complex, a cluster of genes involved in development of the integument. Lastly, we present evidence that CYP2J19 is involved in ketocarotenoid formation in the retina. The discovery of the carotenoid ketolase has important implications for understanding sensory function and signaling mediated by carotenoid pigmentation.

  • 223. Lu, Xuedong
    et al.
    Nie, Shuping
    Xia, Chengjing
    Huang, Lie
    He, Ying
    Wu, Runxiang
    Zhang, Li
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structure and Molecular Biology.
    A rapid two-step algorithm detects and identifies clinical macrolide and beta-lactam antibiotic resistance in clinical bacterial isolates2014In: Journal of Microbiological Methods, ISSN 0167-7012, E-ISSN 1872-8359, Vol. 102, p. 26-31Article in journal (Refereed)
    Abstract [en]

    Purpose: Aiming to identify macrolide and beta-lactam resistance in clinical bacterial isolates rapidly and accurately, a two-step algorithm was developed based on detection of eight antibiotic resistance genes. Methods: Targeting at genes linked to bacterial macrolide (msrA, ermA, ermB, and ermC) and beta-lactam (bla(TEM), bla(SHV), bla(CTX-N-1), bin(CTX-M-9)) antibiotic resistances, this method includes a multiplex real-time PCR, a melting temperature profile analysis as well as a liquid bead microarray assay. Liquid bead microarray assay is applied only when indistinguishable T-m profile is observed. Results: The clinical validity of this method was assessed on clinical bacterial isolates. Among the total 580 isolates that were determined by our diagnostic method, 75% of them were identified by the multiplex real-time PCR with melting temperature analysis alone, while the remaining 25% required both multiplex real-time PCR with melting temperature analysis and liquid bead microarray assay for identification. Compared with the traditional phenotypic antibiotic susceptibility test, an overall agreement of 81.2% (kappa = 0.614, 95% Cl = 0550-0.679) was observed, with a sensitivity and specificity of 87.7% and 73% respectively. Besides, the average test turnaround time is 3.9 h, which is much shorter in comparison with more than 24 h for the traditional phenotypic tests. Conclusions: Having the advantages of the shorter operating time and comparable high sensitivity and specificity with the traditional phenotypic test, our two-step algorithm provides an efficient tool for rapid determination of macrolide and beta-lactam antibiotic resistances in clinical bacterial isolates.  

  • 224.
    Ludvigsen, Eva
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Cell Biology.
    Somatostatin Receptor Expression and Biological Functions in Endocrine Pancreatic Cells2006Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Type 1 diabetes is resulting from the selective destruction of insulin-producing beta-cells within the pancreatic islets. Somatostatin acts as an inhibitor of hormone secretion through specific receptors (sst1-5).

    All ssts were expressed in normal rat and mouse pancreatic islets, although the expression intensity and the co-expression pattern varied between ssts as well as between species. This may reflect a difference in response to somatostatin in islet cells of the two species.

    The Non-Obese Diabetic (NOD) mouse model is an experimental model of type 1 diabetes, with insulitis accompanied by spontaneous hyperglycaemia. Pancreatic specimens from NOD mice at different age and stage of disease were stained for ssts. The islet cells of diabetic NOD mice showed increased islet expression of sst2-5 compared to normoglycemic NOD mice. The increase in sst2-5 expression in the islets cells may suggest either a contributing factor in the process leading to diabetes, or a defense response against ongoing beta-cell destruction.

    Somatostatin analogues were tested on a human endocrine pancreatic tumour cell line and cultured pancreatic islets. Somatostatin analogues had an effect on cAMP accumulation, chromogranin A secretion and MAP kinase activity in the cell line. Treatment of rat pancreatic islets with somatostatin analogues with selective receptor affinity was not sufficient to induce an inhibition of insulin and glucagon secretion. However, a combination of selective analogues or non-selective analogues via co-stimulation of receptors can cause inhibition of hormone production. For insulin and glucagon, combinations of sst2 + sst5 and sst1 + sst2, respectively, showed a biological effect.

    In summary, knowledge of islet cell ssts expression and the effect of somatostatin analogues with high affinity to ssts may be valuable in the future attempts to influence beta-cell function in type 1 diabetes mellitus, since down-regulation of beta-cell function may promote survival of these cells during the autoimmune attack.

    List of papers
    1. Expression and distributon of somatostatin receptor subtypes in the pancreatic islets of mice and rats
    Open this publication in new window or tab >>Expression and distributon of somatostatin receptor subtypes in the pancreatic islets of mice and rats
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    2004 In: Journal of Histochemistry and Cytochemistry, ISSN 0022-1554, Vol. 52, no 3, p. 391-400Article in journal (Refereed) Published
    Identifiers
    urn:nbn:se:uu:diva-93887 (URN)
    Available from: 2006-01-13 Created: 2006-01-13Bibliographically approved
    2. Expression of somatostatin receptor subtypes 1-5 in pancreatic islets of nomoglycemic and diabetic NOD mice
    Open this publication in new window or tab >>Expression of somatostatin receptor subtypes 1-5 in pancreatic islets of nomoglycemic and diabetic NOD mice
    2005 In: European Journal of Endocrinology, ISSN 1479-693, Vol. 153, p. 445-454Article in journal (Refereed) Published
    Identifiers
    urn:nbn:se:uu:diva-93888 (URN)
    Available from: 2006-01-13 Created: 2006-01-13Bibliographically approved
    3. Subtype selective interactions of somatostatin and somatostatin analogs with sst1, sst2 and sst5 in BON-1 cells
    Open this publication in new window or tab >>Subtype selective interactions of somatostatin and somatostatin analogs with sst1, sst2 and sst5 in BON-1 cells
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    2004 In: Medical Oncology, ISSN 1357-0560, Vol. 21, no 3, p. 285-295Article in journal (Refereed) Published
    Identifiers
    urn:nbn:se:uu:diva-93889 (URN)
    Available from: 2006-01-13 Created: 2006-01-13Bibliographically approved
    4. Regulation of insulin and glucagon secretion from rat pancreatic islets in vitro by somatostatin analogues
    Open this publication in new window or tab >>Regulation of insulin and glucagon secretion from rat pancreatic islets in vitro by somatostatin analogues
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    2007 (English)In: Regulatory Peptides, ISSN 0167-0115, E-ISSN 1873-1686, Vol. 138, no 1, p. 1-9Article in journal (Refereed) Published
    Abstract [en]

    Somatostatin is an inhibitor of hormone secretion through specific receptors (sst1-5). The aim of this study was to investigate the putative regulatory role of somatostatin analogues on the secretion of insulin and glucagon by rat pancreatic islets. After 48 h exposure only the non-selective agonists (somatostatin, octreotide and SOM-230) inhibited insulin accumulation. The inhibition of insulin secretion was accompanied by increased islet insulin contents. None of the analogues showed a consistent effect on the glucagon accumulation in the medium after 48 h. Since we observed a difference in the regulatory effect between the non-selective and selective analogues, combinations of selective analogues were studied. Combination of sst2 + sst5 agonists inhibited the medium insulin accumulation, while combination of sst1 + sst2 analogues caused a decrease in glucagon accumulation. After removal of somatostatin a rebound effect with increased insulin secretion were observed. This effect was reversed after 6 h. For SOM-230 insulin secretion continued to be suppressed even after the analogue was removed and returned to control values after 3 h. As for glucagon secretion there was an initial decline after culture with octreotide, while the other substances failed to induce any changes. In summary, non-selective somatostatin analogues or combinations of receptor selective analogues may cause inhibition of hormone secretion from rat pancreatic islets. For insulin and glucagon, combinations of sst2 + sst5 and sst1 + sst2, respectively may exert this effects. Thus, our data suggest that more than one sst must be involved to down-regulate islet glucagon and insulin secretion.

    Keywords
    Glucagon secretion, Insulin secretion, Pancreatic islets, Somatostatin analogues, Somatostatin receptors
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-93890 (URN)10.1016/j.regpep.2006.07.006 (DOI)000243772600001 ()16935361 (PubMedID)
    Available from: 2006-01-13 Created: 2006-01-13 Last updated: 2017-12-14Bibliographically approved
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  • 225.
    Lugano, Roberta
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology.
    Ramachandran, Mohanraj
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology.
    Dimberg, Anna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Vascular Biology.
    Tumor angiogenesis: causes, consequences, challenges and opportunities2020In: Cellular and Molecular Life Sciences (CMLS), ISSN 1420-682X, E-ISSN 1420-9071, Vol. 77, no 9, p. 1745-1770Article, review/survey (Refereed)
    Abstract [en]

    Tumor vascularization occurs through several distinct biological processes, which not only vary between tumor type and anatomic location, but also occur simultaneously within the same cancer tissue. These processes are orchestrated by a range of secreted factors and signaling pathways and can involve participation of non-endothelial cells, such as progenitors or cancer stem cells. Anti-angiogenic therapies using either antibodies or tyrosine kinase inhibitors have been approved to treat several types of cancer. However, the benefit of treatment has so far been modest, some patients not responding at all and others acquiring resistance. It is becoming increasingly clear that blocking tumors from accessing the circulation is not an easy task to accomplish. Tumor vessel functionality and gene expression often differ vastly when comparing different cancer subtypes, and vessel phenotype can be markedly heterogeneous within a single tumor. Here, we summarize the current understanding of cellular and molecular mechanisms involved in tumor angiogenesis and discuss challenges and opportunities associated with vascular targeting.

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  • 226.
    Luna Marco, Clara
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Role of inositol requiring enzyme-1 alpha on lipid metabolism and response to doxorubicin in hepatocellular carcinoma2022Independent thesis Advanced level (degree of Master (Two Years)), 80 credits / 120 HE creditsStudent thesis
    Abstract [en]

    BACKGROUND:Hepatocellular carcinoma (HCC) is associated with endoplasmic reticulum (ER) stress and lipid metabolism disruption. Doxorubicin (DOX), a common treatment for HCC, and ER-stress can alter lipid composition of the tumour, contributing to tumorigenesis.AIM: to determine the role of inositol requiring enzyme-1 alpha (IRE1α) on lipid metabolism and response to DOX in HCC.METHODS:HepG2 cells and a diethylnitrosamine-induced HCC mouse model were treated with 4μ8C (IRE1α inhibitor), DOX or combination (4μ8C+DOX). Lipid accumulation in tumour was assessed by quantification of liver triglycerides (TGs) levels in mouse tissue. In HepG2 cells, Oil Red staining was used to detect lipid droplets and expression of lipid metabolic markers (adipose triglyceride lipase (ATGL), monoacylglycerol lipase (MGL) and lysosomal acid lipase (LAL)) was measured by RT-qPCR. Seahorse analysis was used to determine the oxygen consumption rate (OCR) in HepG2 cells under the effect of the treatments. ANOVA tests were applied for statistical analysis (p<0.05).RESULTS:Treatment with 4μ8C+DOX decreased TG enrichment in tumours from mice and reduced lipid droplet accumulation in HepG2 cells. ATGL, MGL and LAL mRNA expression was increased after treatment with DOX, but this was reverted to control levels after treating HepG2 cells with 4μ8C+DOX. Cell metabolism was increased (higher OCR levels) after treatment with DOX and 4μ8C+DOX in HepG2 cells.CONCLUSION: Inhibition of IRE1α ER-stress pathway combined with DOX treatment may reduce lipid accumulation in liver tumours from mice and HepG2 cells, potentially contributing to an improved cancer therapy. However, the underlying mechanisms of these observations need to be further investigated.

  • 227. Lyberg, Katarina
    et al.
    Ali, Hani Abdulkadir
    Grootens, Jennine
    Kjellander, Matilda
    Tirfing, Malin
    Arock, Michel
    Hägglund, Hans
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Haematology.
    Nilsson, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Haematology.
    Ungerstedt, Johanna
    Histone deacetylase inhibitor SAHA mediates mast cell death and epigenetic silencing of constitutively active D816V KIT in systemic mastocytosis.2017In: Oncotarget, E-ISSN 1949-2553, Vol. 8, no 6, p. 9647-9659Article in journal (Refereed)
    Abstract [en]

    Systemic mastocytosis (SM) is a clonal bone marrow disorder, where therapeutical options are limited. Over 90% of the patients carry the D816V point mutation in the KIT receptor that renders this receptor constitutively active. We assessed the sensitivity of primary mast cells (MC) and mast cell lines HMC1.2 (D816V mutated), ROSA (KIT WT) and ROSA (KIT D816V) cells to histone deacetylase inhibitor (HDACi) treatment. We found that of four HDACi, suberoyl anilide hydroxamic acid (SAHA) was the most effective in killing mutated MC. SAHA downregulated KIT, followed by major MC apoptosis. Primary SM patient MC cultured ex vivo were even more sensitive to SAHA than HMC1.2 cells, whereas primary MC from healthy subjects were less affected. There was a correlation between cell death and SM disease severity, where cell death was more pronounced in the case of aggressive SM, with almost 100% cell death among MC from the mast cell leukemia patient. Additionally, ROSA (KIT D816V) was more affected by HDACi than ROSA (KIT WT) cells. Using ChIP qPCR, we found that the level of active chromatin mark H3K18ac/H3 decreased significantly in the KIT region. This epigenetic silencing was seen only in the KIT region and not in control genes upstream and downstream of KIT, indicating that the downregulation of KIT is exerted by specific epigenetic silencing. In conclusion, KIT D816V mutation sensitized MC to HDACi mediated killing, and SAHA may be of value as specific treatment for SM, although the specific mechanism of action requires further investigation.

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  • 228.
    Lyngå, Olivia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Cancer precision medicine.
    Cellular repair of DNA double-strand breaks of different complexity2024Independent thesis Advanced level (professional degree), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    Cancer is a leading cause of death, with a worldwide mortality of approximately 10 million people every year. Cancer cells often have dysfunctional cell-cycle regulation and rapid cell division. Thus, most non-surgical cancer treatments stop the proliferation and kill the cancer cells. One option to further improve cancer therapy is to target proteins responsible for the repair of the most lethal form of DNA damage, double-strand breaks (DSBs), which are induced by ionising radiation and several anticancer drugs. The aim of the project was therefore to study the role of two proteins involved in the non-homologous end joining (NHEJ) pathway of DSB repair, DNA-dependent protein kinase catalytic subunit (DNA-PKcs) and X-ray repair cross-complementing protein 4 (XRCC4).

    The role of these repair proteins was studied by analysing the clonogenic survival of wild-type Chinese hamster ovary (CHO) cells and CHO cells where either DNA-PKcs or XRCC4 had been knocked-out (KO). The cells were exposed to two different DNA-damaging agents: ionising radiation (X-rays) or the cytotoxic drug Calicheamicin (CLM). The formation and repair of DSBs were analysed using immunohistochemical detection of the DSB markers 53BP1 and γH2AX, which could be visualized by fluorescence microscopy. To gather more knowledge about the kinetics of protein-protein interactions between these two repair proteins, proximity ligation was performed with normal human fibroblasts.

    The results showed that the repair proteins were equally important for cell survival with an increased sensitivity to DNA damage for the KO cell lines compared to the wild-type, and the survival remained equal for the two KO cell lines. However, XRCC4 was more important than DNA-PKcs in the repair of DSBs, suggesting that cell death in DNA-PKcs-defective cells is not only related to the repair capacity. During DSB repair the repair proteins were rapidly in close proximity following exposure to X-rays. The KO cell lines showed greater sensitivity to CLM than X-rays compared to wild-type, indicating that X-ray induced DSBs were less toxic than CLM induced DSBs.

    In conclusion, both DNA-PKcs and XRCC4 have key roles in DSB repair and may serve as potential targets in the development of drugs to sensitise cancer cells treated with DSB-inducing therapies such as X-rays.

    The full text will be freely available from 2025-06-03 18:43
  • 229.
    Löfås, Mathilda
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Förhoppningarna över CAR-T celler som behandlingsmetod mot hematologisk cancer2018Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesis
    Abstract [sv]

    Olika typer av blodcancer är några av de vanligast förekommande cancerformerna i världen. Ju tidigare sjukdomen upptäcks och beroende på vilken typ av blodcancer någon drabbats av ser prognosen i många fall god ut då dagens behandlingsmetoder under flera år effektiviserats. Dock finns det fall där patienter inte svarar på traditionella terapier som cytostatika och stålning eller gånger där sjukdomen kommer tillbaka i aggressivare former. I dessa fall krävs andra metoder för att försöka behandla och bota cancern.

     

    En metod som på senare tid fått mycket uppmärksamhet i forskarvärlden är användandet av kroppens egna immunceller som behandlingsform mot blodcancer. Genom att rena fram en patients egna T-celler från ett blodprov, kan dessa sedan genetiskt modifieras till att känna igen specifika tumörassocierade antigener (TAA) som bara vissa typer av cancerceller uttrycker. Metoden går ut på att T-cellerna får chimära antigen-receptorer (CAR), som uttrycks på cellytan, där CAR-T cellerna sedan injiceras tillbaka till patienten. CAR-T cellerna känner igen cancercellerna och attackerar sedan, med målet att patienten efter behandlingen inte ska ha några cancerceller kvar i kroppen.

     

    Kliniska försök gjorda på patienter med olika typer av blodcancer har visat lovande resultat, särskilt gällande patienter som fått återfall av blodcancertypen Akut Lymfatisk Leukemi (ALL). De som fått delta i studierna har haft mycket dåliga prognoser och har innan blivit behandlade med de konventionella behandlingsterapierna, men utan eller med mycket dåligt resultat. Förhoppningar som väckts från dessa forskningsresultat har lett till diskussion att CAR-T celler kan komma att förändra cancervården och i framtiden kanske vara en lika vanlig behandlingsmetod som strålning eller kemoterapi. Dock kvarstår många problem som forskarna måste lyckas lösa innan CAR-T celler kan räknas som konventionell. Bland annat finns stora risker att patienter vid behandling kan drabbas av cytokinfrisläppningssyndrom (CRS, eng. cytokine release syndrome), där immunförsvaret kan attackera kroppens egna organ som i värsta fall kan leda till döden. Cancercellerna kan även komma tillbaka efterbehandling, då de har utvecklat en resistens mot CAR-T cellerna. Metoder för att undvika dessa toxiska responser och göra T-cellerna mer effektiva är bara några av de problem som kvarstår.

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  • 230.
    Mahammad, Saleemulla
    Stockholms universitet, Wenner-Grens institut.
    Cholesterol in T cells: homeostasis, plasma membrane organization and signaling2010Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The plasma membrane of eukaryotic cells contains cholesterol and glycosphingolipids enriched nanodomains known as lipid rafts; which are believed to exist in a liquid ordered (lo) state. Methyl-beta-cyclodextrin (MBCD) is used to deplete cellular cholesterol and a widespread assumption is that MBCD preferentially targets cholesterol in lipid rafts. To analyze this in T cells a progressive cholesterol extraction protocols was established. At 37ºC, MBCD treatment does not lead to the preferential loss of cholesterol from TX-DRMs. At 0ºC only 35% of total cholesterol could be extracted demonstrating that less than 35% of the cell’s cholesterol is found in the plasma membrane. Moreover, incubation of cells at 0ºC causes loss of plasma membrane cholesterol and an increase in cholesteryl esters. The increase in cholesterol esters upon cold exposure is linked to the cholesterol concentration induced activation of ACAT enzyme which converts cholesterol to cholesteryl esters. Cholesterol concentration specific activation of ACAT and conversion of cholesterol to cholesteryl esters during the loading of cholesterol onto T cells by MBCD was also observed. By using MBCD for progressive cholesterol depletion from T cells at 37ºC, the effect of cholesterol depletion on T cell signaling was addressed. At 10-20% cholesterol depletion levels, tyrosine phosphorylation is increased and ERK is activated. Peripheral actin polymerization, cell spreading and membrane protrusions are also triggered by limited cholesterol depletion. Upon limited cholesterol depletion aggregation of lipid rafts in the plasma membrane was observed. The aggregation of lipid rafts upon cholesterol depletion does not dependent on the signaling proteins such as Src-kinases. Upon cholesterol depletion there is an increase in overall plasma membrane order, indicative of more ordered domains forming at the expense of disordered domains.

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  • 231.
    Mahammad, Saleemulla
    et al.
    Stockholms universitet, Wenner-Grens institut.
    Dinic, Jelena
    Stockholms universitet, Wenner-Grens institut.
    Adler, Jeremy
    Stockholms universitet, Wenner-Grens institut.
    Parmryd, Ingela
    Stockholms universitet, Wenner-Grens institut.
    Limited cholesterol depletion causes aggregation of plasma membrane lipid raftsinducing T cell activation2010In: Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids, ISSN 1388-1981, E-ISSN 1879-2618, Vol. 1801, no 6, p. 625-634Article in journal (Refereed)
    Abstract [en]

    Acute cholesterol depletion is generally associated with decreased or abolished T cell signalling but it can also cause T cell activation. This anomaly has been addressed in Jurkat T cells using progressive cholesterol depletion with methyl-beta-cyclodextrin (MBCD). At depletion levels higher than 50% there is substantial cell death, which explains reports of signalling inhibition. At 10–20% depletion levels, tyrosine phosphorylation is increased, ERK is activated and there is a small increase in cytoplasmic Ca2+. Peripheral actin polymerisation is also triggered by limited cholesterol depletion. Strikingly, the lipid raft marker GM1 aggregates upon cholesterol depletion and these aggregated domains concentrate the signalling proteins Lck and LAT, whereas the opposite is true for the non lipid raft marker the transferrin receptor. Using PP2, an inhibitor of Src family kinase activation, it is demonstrated that the lipid raft aggregation occurs independently of and thus upstream of the signalling response. Upon cholesterol depletion there is an increase in overall plasma membrane order, indicative of more ordered domains forming at the expense of disordered domains. That cholesterol depletion and not unspecific effects of MBCD was behind the reported results was confirmed by performing all experiments with MBCD–cholesterol, when no net cholesterol extraction took place. We conclude that non-lethal cholesterol depletion causes the aggregation of lipid rafts which then induces T cell signalling.

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  • 232.
    Mahammad, Saleemulla
    et al.
    Stockholms universitet, Wenner-Grens institut.
    Parmryd, Ingela
    Stockholms universitet, Wenner-Grens institut.
    Cholesterol homeostasis in T cells. Methyl-beta-cyclodextrin treatment results in equal loss of cholesterol from Triton X-100 soluble and insoluble fractions.2008In: Biochimica et Biophysica Acta, ISSN 0006-3002, E-ISSN 1878-2434, Vol. 1778, no 5, p. 1251-8Article in journal (Refereed)
    Abstract [en]

    Methyl-beta-cycloclextrin (MBCD) is frequently used to acutely deplete cells of cholesterol. A widespread assumption is that MBCD preferentially targets cholesterol in lipid rafts and that sensitivity to MBCD is proof of lipid raft involvement in a cellular process. To analyse any MBCD preference systematically, progressive cholesterol depletion of Jurkat T cells was performed using MBCD and [H-3]-cholesterol. It was found that at 37 degrees C, MBCD extracts similar proportions of cholesterol from the Triton X-100 resistant (lipid raft enriched) as it does from other cellular fractions and that the cells rapidly reestablish the relative differences in cholesterol concentration between different compartments. Moreover, cells restore the cholesterol level in the plasma membrane by mobilising cholesterol from intracellular cholesterol stores. Interestingly, mere incubation at 0 degrees C caused a loss of plasma membrane cholesterol with a concomitant increase in cholesteryl esters and adiposomes. Moreover, only 35% of total cholesterol could be extracted by MBCD at 0 degrees C and was accompanied by a complete loss of plasma membrane and endocytotic recycling centre filipin staining. This study clearly shows that MBCD does not specifically extract cholesterol from any cellular fraction, that cholesterol redistributes upon temperature changes and that intracellular cholesterol stores can be used to replenish plasma membrane cholesterol.

  • 233.
    Majda, Mateusz
    et al.
    Swedish Univ Agr Sci, Dept Forest Genet & Plant Physiol, UPSC, S-90183 Umea, Sweden..
    Grones, Peter
    Swedish Univ Agr Sci, Dept Forest Genet & Plant Physiol, UPSC, S-90183 Umea, Sweden..
    Sintorn, Ida-Maria
    Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Computerized Image Analysis and Human-Computer Interaction. Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Division of Visual Information and Interaction. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Vain, Thomas
    Swedish Univ Agr Sci, Dept Forest Genet & Plant Physiol, UPSC, S-90183 Umea, Sweden..
    Milani, Pascale
    Univ Lyon, ENS Lyon, UCB Lyon 1, Lab Reprod & Dev Plantes,CNRS,INRA, F-69342 Lyon, France..
    Krupinski, Pawel
    Lund Univ, Dept Astron & Theoret Phys, Computat Biol & Biol Phys, Solvegatan 14A, S-22362 Lund, Sweden..
    Zagorska-Marek, Beata
    Univ Wroclaw, Inst Expt Biol, Dept Plant Dev Biol, Kanonia 6-8, PL-50328 Wroclaw, Poland..
    Viotti, Corrado
    Umea Univ, Dept Plant Physiol, UPSC, S-90187 Umea, Sweden.;Univ Potsdam, Inst Biochem & Biol, Plant Physiol, D-14476 Potsdam, Germany..
    Jonsson, Henrik
    Lund Univ, Dept Astron & Theoret Phys, Computat Biol & Biol Phys, Solvegatan 14A, S-22362 Lund, Sweden.;Univ Cambridge, Sainsbury Lab, Bateman St, Cambridge CB2 1LR, England.;Univ Cambridge, Dept Math & Theoret Phys, Cambridge CB3 0WA, England..
    Mellerowicz, Ewa J.
    Swedish Univ Agr Sci, Dept Forest Genet & Plant Physiol, UPSC, S-90183 Umea, Sweden..
    Hamant, Olivier
    Univ Lyon, ENS Lyon, UCB Lyon 1, Lab Reprod & Dev Plantes,CNRS,INRA, F-69342 Lyon, France..
    Robert, Stephanie
    Swedish Univ Agr Sci, Dept Forest Genet & Plant Physiol, UPSC, S-90183 Umea, Sweden..
    Mechanochemical Polarization of Contiguous Cell Walls Shapes Plant Pavement Cells2017In: Developmental Cell, ISSN 1534-5807, E-ISSN 1878-1551, Vol. 43, no 3, p. 290-304Article in journal (Refereed)
    Abstract [en]

    The epidermis of aerial plant organs is thought to be limiting for growth, because it acts as a continuous load-bearing layer, resisting tension. Leaf epidermis contains jigsaw puzzle piece-shaped pavement cells whose shape has been proposed to be a result of subcellular variations in expansion rate that induce local buckling events. Paradoxically, such local compressive buckling should not occur given the tensile stresses across the epidermis. Using computational modeling, we show that the simplest scenario to explain pavement cell shapes within an epidermis under tension must involve mechanical wall heterogeneities across and along the anticlinal pavement cell walls between adjacent cells. Combining genetics, atomic force microscopy, and immunolabeling, we demonstrate that contiguous cell walls indeed exhibit hybrid mechanochemical properties. Such biochemical wall heterogeneities precede wall bending. Altogether, this provides a possible mechanism for the generation of complex plant cell shapes.

  • 234.
    Maklakov, Alexei A.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Ecology and Genetics, Animal ecology.
    Friberg, Urban
    Linköping Univ, IFM Biol AVIAN Behav Genom & Physiol Grp, S-58183 Linköping, Sweden..
    Ageing: Why Males Curtail the Longevity of Their Mates2016In: Current Biology, ISSN 0960-9822, E-ISSN 1879-0445, Vol. 26, no 20, p. R929-R932Article in journal (Other academic)
    Abstract [en]

    Male nematodes secrete pheromones that accelerate the somatic senescence of potential mates. A new study shows that this harm most likely is an unintended by-product of the males' aim to speed up sexual maturation and delay reproductive senescence of future partners.

  • 235.
    Maklakov, Alexei A.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Ecology and Genetics, Animal ecology.
    Immler, Simone
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Ecology and Genetics, Evolutionary Biology.
    The Expensive Germline and the Evolution of Ageing2016In: Current Biology, ISSN 0960-9822, E-ISSN 1879-0445, Vol. 26, no 13, p. R577-R586Article, review/survey (Refereed)
    Abstract [en]

    The trade-off between survival and reproduction is the bedrock of the evolutionary theory of ageing. The reproductive system regulates ageing of the soma, and removal of germ cells extends somatic lifespan and increases resistance to a broad variety of abiotic and biotic stresses. The general explanation for this somatic response is that reduced reproduction frees up resources for survival. Remarkably, however, the disruption of molecular signaling pathways that regulate ageing increases lifespan without the obligatory reduction in fecundity, thus challenging the key role of the survival-reproduction trade-off. Here, we review the diverse literature on the costs of lifespan extension and suggest that the current paradigm is overly centered on the trade-off between lifespan and fecundity, often neglecting key aspects of fitness, such as development time, defense against parasites and, in particular, the high costs of germline maintenance. Compromised germline maintenance increases germline mutation rate, which reduces offspring fitness and ultimately can terminate germline proliferation across generations. We propose that future work should incorporate the costs of germline maintenance in the study of ageing evolution, as well as in applied biomedical research, by assessing offspring fitness.

  • 236. Maksimov, Vladimir
    et al.
    Nakamura, Miyuki
    Wildhaber, Thomas
    Nanni, Paolo
    Ramström, Margareta
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Bergquist, Jonas
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Hennig, Lars
    The H3 chaperone function of NASP is conserved in Arabidopsis.2016In: The Plant Journal, ISSN 0960-7412, E-ISSN 1365-313X, Vol. 88, no 3, p. 425-436Article in journal (Refereed)
    Abstract [en]

    Histones are abundant cellular proteins but, if not incorporated into chromatin, they are usually bound by histone chaperones. Here, we identify Arabidopsis NASP as a chaperone for histones H3.1 and H3.3. NASP interacts in vitro with monomeric H3.1 and H3.3 as well as with histone H3.1-H4 and H3.3-H4 dimers. However, NASP does not bind to monomeric H4. NASP shifts the equilibrium between histone dimers and tetramers towards tetramers but does not interact with tetramers in vitro. Arabidopsis NASP promotes [H3-H4]2 tetrasome formation, possibly by providing preassembled histone tetramers. However, NASP does not promote disassembly of in vitro preassembled tetrasomes. In contrast to its mammalian homolog, Arabidopsis NASP is a predominantly nuclear protein. In vivo, NASP binds mainly monomeric H3.1 and H3.3. Pulldown experiments indicated that NASP may also interact with the histone chaperone MSI1 and a HSC70 heat shock protein.

  • 237.
    Mangano, Alexander
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre.
    Hormonal Influence on the Proliferation Potential of C17.2 Neuronal Progenitor Cells2024Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesis
    Abstract [en]

    The brain is arguably the most complex organ of the body: Controlling muscles, maintaining homeostasis, processing information. It has the longest developmental period of any organ, and hormones are essential during its development. Endocrine disruptive chemicals can disturb hormones in any part of their signaling pathway and can act as agonists or antagonists on their receptors. Should the balance of stem cell or progenitor cell proliferation be skewed in any direction during early brain development, it may lead to cognitive disfunctions or diseases. In this study, v-myc induced mouse-derived neural progenitor cells (NPCs) were exposed to retinoid and glucocorticoid receptor agonists and antagonists to investigate the involvement of these pathways on NPC proliferation. NPCs were exposed for 24 hours, after which they were read in a Tecan. Statistical analyses revealed that no treatment yielded statistically significant responses, however in the case for mifepristone (glucocorticoid receptor antagonist), dexamethasone (glucocorticoid receptor agonist) and Agn (retinoid receptor antagonist), a small negative response trend could be observed. Cell viability analyses also revealed that the concentrations of chemicals did not induce cytotoxicity. The unexpectedly high and variable data suggest errors were made during the seeding of the 96-well microplates. In conclusion, these findings suggest that with further development, this in vitro method could be used in conjunction with other in vitro models for the assessment of neurodevelopmental toxicity and for identification of EDCs in the future.

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    Bachelor's thesis
  • 238.
    Markó, Dorottya
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Biology Education Centre. AstraZeneca.
    Derivation and characterization of pacemaker cells from human induced pluripotent stem cells for investigation of cardiotoxicity2021Independent thesis Advanced level (degree of Master (Two Years)), 40 credits / 60 HE creditsStudent thesis
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  • 239.
    Marsit, Souhir
    et al.
    Univ Laval, IBIS, Dept Biol, PROTEO, Quebec City, PQ, Canada.
    Dion-Côté, Anne-Marie
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Ecology and Genetics, Evolutionary Biology. Cornell Univ, Dept Mol Biol & Genet, Ithaca, NY USA.
    Barbash, Daniel A.
    Cornell Univ, Dept Mol Biol & Genet, Ithaca, NY USA.
    Did Mitochondria Kill the Frog?2018In: Developmental Cell, ISSN 1534-5807, E-ISSN 1878-1551, Vol. 44, no 5, p. 539-541Article in journal (Other academic)
    Abstract [en]

    Genomic divergence can cause reproductive isolation between species. The molecular mechanisms underlying reproductive isolation can thus reveal which genomic features evolve rapidly and become unstable or incompatible in hybrids. In a recent paper in Nature, Gibeaux et al. (2018) report paternal genome instability and metabolic imbalance in hybrids between frog species.

  • 240. Masson, Patrick
    et al.
    Lundin, Daniel
    Söderbom, Fredrik
    Department of Molecular Biology, Swedish University of Agricultural Sciences (SLU), Uppsala Biomedical Center (BMC), Box 590, S-75124 Uppsala, Sweden .
    Young, Patrick
    Characterization of a REG/PA28 proteasome activator homolog in Dictyostelium discoideum indicates that the ubiquitin- and ATP-independent REGgamma proteasome is an ancient nuclear protease2009In: Eukaryotic Cell, ISSN 1535-9778, E-ISSN 1535-9786, Vol. 8, no 6, p. 844-851Article in journal (Refereed)
    Abstract [en]

    The nuclear proteasome activator REGgamma/PA28gamma is an ATP- and ubiquitin-independent activator of the 20S proteasome and has been proposed to degrade and thereby regulate both a key human oncogene, encoding the coactivator SRC-3/AIB1, and the cyclin-dependent kinase inhibitor p21 (Waf/Cip1). We report the identification and characterization of a PA28/REG homolog in Dictyostelium. Association of a recombinant Dictyostelium REG with the purified Dictyostelium 20S proteasome led to the preferential stimulation of the trypsin-like proteasome peptidase activity. Immunolocalization studies demonstrated that the proteasome activator is localized to the nucleus and is present in growing as well as starving Dictyostelium cells. Our results indicate that the Dictyostelium PA28/REG activator can stimulate both the trypsin-like and chymotrypsin-like activities of the 20S proteasome and supports the idea that the REGgamma-20S proteasome represents an early unique nuclear degradation pathway for eukaryotic cells.

  • 241.
    Maturi, Varun
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Gene regulation by different proteins of TGFβ superfamily2018Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The present thesis discusses how gene regulation by transforming growth factor β (TGFβ) family cytokines is affected by post-translational modifications of different transcription factors. The thesis also focuses on gene regulation by transcription factors involved in TGFβ signaling.

    The importance of the poly ADP-ribose polymerase (PARP) family in controlling gene expression in response to TGFβ and bone morphogenetic protein (BMP) is analyzed first. PARP2, along with PARP1, ADP-ribosylates Smad2 and Smad3, the signaling mediators of TGFβ. On the other hand, poly ADP-ribose glycohydrolase (PARG) removes the ADP-ribose from Smad2/3 and antagonizes PARP1 and PARP2. ADP-ribosylation of Smads in turn affects their DNA binding capacity. We then illustrate how PARP1 and PARG can regulate gene expression in response to BMP that signals via Smad1, 5. Over-expression of PARP1 suppressed the transcriptional activity of Smad1/5. Knockdown of PARP1 or over-expression of PARG enhanced the transcriptional activity of BMP-Smads on target genes. Hence our data suggest that ADP-ribosylation of Smad proteins controls both TGFβ and BMP signaling. 

    I then focus on elucidating novel genes that are regulated by ZEB1 and Snail1, two key transcriptional factors in TGFβ signaling, known for their ability to induce EMT and cancer metastasis. Chromatin immunoprecipitation-sequencing (ChIP-seq) and targeted whole genome transcriptomics in triple negative breast cancer cells were used, to find binding regions and the functional impact of ZEB1 and Snail1 throughout the genome. ZEB1 binds to the regulatory sequences of a wide range of genes, not only related to cell invasion, pointing to new functions of ZEB1. On the other hand, Snail1 regulated only a few genes, especially related to signal transduction and cellular movement. Further functional analysis revealed that ZEB1 could regulate the anchorage-independent growth of the triple negative breast cancer cells, whereas Snail1 could regulate the expression of BMP6 in these cells. We have therefore elucidated novel functional roles of the two transcription factors, Snail1 and ZEB1 in triple negative breast cancer cells.

    List of papers
    1. Fine-Tuning of Smad Protein Function by Poly(ADP-Ribose) Polymerases and Poly(ADP-Ribose) Glycohydrolase during Transforming Growth Factor β Signaling
    Open this publication in new window or tab >>Fine-Tuning of Smad Protein Function by Poly(ADP-Ribose) Polymerases and Poly(ADP-Ribose) Glycohydrolase during Transforming Growth Factor β Signaling
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    2014 (English)In: PLOS ONE, E-ISSN 1932-6203, Vol. 9, no 8, p. e103651-Article in journal (Refereed) Published
    Abstract [en]

    BACKGROUND:

    Initiation, amplitude, duration and termination of transforming growth factor β (TGFβ) signaling via Smad proteins is regulated by post-translational modifications, including phosphorylation, ubiquitination and acetylation. We previously reported that ADP-ribosylation of Smads by poly(ADP-ribose) polymerase 1 (PARP-1) negatively influences Smad-mediated transcription. PARP-1 is known to functionally interact with PARP-2 in the nucleus and the enzyme poly(ADP-ribose) glycohydrolase (PARG) can remove poly(ADP-ribose) chains from target proteins. Here we aimed at analyzing possible cooperation between PARP-1, PARP-2 and PARG in regulation of TGFβ signaling.

    METHODS:

    A robust cell model of TGFβ signaling, i.e. human HaCaT keratinocytes, was used. Endogenous Smad3 ADP-ribosylation and protein complexes between Smads and PARPs were studied using proximity ligation assays and co-immunoprecipitation assays, which were complemented by in vitro ADP-ribosylation assays using recombinant proteins. Real-time RT-PCR analysis of mRNA levels and promoter-reporter assays provided quantitative analysis of gene expression in response to TGFβ stimulation and after genetic perturbations of PARP-1/-2 and PARG based on RNA interference.

    RESULTS:

    TGFβ signaling rapidly induces nuclear ADP-ribosylation of Smad3 that coincides with a relative enhancement of nuclear complexes of Smads with PARP-1 and PARP-2. Inversely, PARG interacts with Smads and can de-ADP-ribosylate Smad3 in vitro. PARP-1 and PARP-2 also form complexes with each other, and Smads interact and activate auto-ADP-ribosylation of both PARP-1 and PARP-2. PARP-2, similar to PARP-1, negatively regulates specific TGFβ target genes (fibronectin, Smad7) and Smad transcriptional responses, and PARG positively regulates these genes. Accordingly, inhibition of TGFβ-mediated transcription caused by silencing endogenous PARG expression could be relieved after simultaneous depletion of PARP-1.

    CONCLUSION:

    Nuclear Smad function is negatively regulated by PARP-1 that is assisted by PARP-2 and positively regulated by PARG during the course of TGFβ signaling.

    National Category
    Clinical Medicine
    Identifiers
    urn:nbn:se:uu:diva-231920 (URN)10.1371/journal.pone.0103651 (DOI)000341302700014 ()25133494 (PubMedID)
    Available from: 2014-09-11 Created: 2014-09-11 Last updated: 2022-01-28Bibliographically approved
    2. Regulation of Bone Morphogenetic Protein Signaling by ADP-ribosylation
    Open this publication in new window or tab >>Regulation of Bone Morphogenetic Protein Signaling by ADP-ribosylation
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    2016 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 291, no 24, p. 12706-12723Article in journal (Refereed) Published
    Abstract [en]

    We previously established a mechanism of negative regulation of transforming growth factor beta signaling mediated by the nuclear ADP-ribosylating enzyme poly-(ADP-ribose) polymerase 1 (PARP1) and the deribosylating enzyme poly-(ADP-ribose) glycohydrolase (PARG), which dynamically regulate ADP-ribosylation of Smad3 and Smad4, two central signaling proteins of the pathway. Here we demonstrate that the bone morphogenetic protein (BMP) pathway can also be regulated by the opposing actions of PARP1 and PARG. PARG positively contributes to BMP signaling and forms physical complexes with Smad5 and Smad4. The positive role PARG plays during BMP signaling can be neutralized by PARP1, as demonstrated by experiments where PARG and PARP1 are simultaneously silenced. In contrast to PARG, ectopic expression of PARP1 suppresses BMP signaling, whereas silencing of endogenous PARP1 enhances signaling and BMP-induced differentiation. The two major Smad proteins of the BMP pathway, Smad1 and Smad5, interact with PARP1 and can be ADP-ribosylated in vitro, whereas PARG causes deribosylation. The overall outcome of this mode of regulation of BMP signal transduction provides a fine-tuning mechanism based on the two major enzymes that control cellular ADP-ribosylation.

    National Category
    Cell and Molecular Biology
    Identifiers
    urn:nbn:se:uu:diva-299724 (URN)10.1074/jbc.M116.729699 (DOI)000378119900024 ()27129221 (PubMedID)
    Funder
    Swedish Research Council, K2010-67X-14936-07-3 K2013-66X-14936-10-5
    Available from: 2016-07-26 Created: 2016-07-26 Last updated: 2018-10-23Bibliographically approved
    3. Genome-wide binding of transcription factor ZEB1 in triple-negative breast cancer cells
    Open this publication in new window or tab >>Genome-wide binding of transcription factor ZEB1 in triple-negative breast cancer cells
    2018 (English)In: Journal of Cellular Physiology, ISSN 0021-9541, E-ISSN 1097-4652, Vol. 233, no 10, p. 7113-7127Article in journal (Refereed) Published
    Abstract [en]

    Zinc finger E-box binding homeobox 1 (ZEB1) is a transcriptional regulator involved in embryonic development and cancer progression. ZEB1 induces epithelial-mesenchymal transition (EMT). Triple-negative human breast cancers express high ZEB1 mRNA levels and exhibit features of EMT. In the human triple-negative breast cancer cell model Hs578T, ZEB1 associates with almost 2,000 genes, representing many cellular functions, including cell polarity regulation (DLG2 and FAT3). By introducing a CRISPR-Cas9-mediated 30bp deletion into the ZEB1 second exon, we observed reduced migratory and anchorage-independent growth capacity of these tumor cells. Transcriptomic analysis of control and ZEB1 knockout cells, revealed 1,372 differentially expressed genes. The TIMP metallopeptidase inhibitor 3 and the teneurin transmembrane protein 2 genes showed increased expression upon loss of ZEB1, possibly mediating pro-tumorigenic actions of ZEB1. This work provides a resource for regulators of cancer progression that function under the transcriptional control of ZEB1. The data confirm that removing a single EMT transcription factor, such as ZEB1, is not sufficient for reverting the triple-negative mesenchymal breast cancer cells into more differentiated, epithelial-like clones, but can reduce tumorigenic potential, suggesting that not all pro-tumorigenic actions of ZEB1 are linked to the EMT.

    Keywords
    ZEB1, EMT, ChIP-seq, CRISPR-Cas9
    National Category
    Cell Biology
    Research subject
    Biochemistry; Biology with specialization in Molecular Cell Biology
    Identifiers
    urn:nbn:se:uu:diva-334438 (URN)10.1002/jcp.26634 (DOI)000438352300071 ()29744893 (PubMedID)
    Funder
    Swedish Research Council, 2015-02757Swedish Research Council, K2013-66X-14936-10-5Swedish Cancer Society, CAN 2012/438Swedish Cancer Society, CAN 2015/438Swedish Cancer Society, CAN 2016/445
    Available from: 2017-11-23 Created: 2017-11-23 Last updated: 2018-09-24Bibliographically approved
    4. Genome-wide binding of transcription factor Snail1 in triple-negative breast cancer cells
    Open this publication in new window or tab >>Genome-wide binding of transcription factor Snail1 in triple-negative breast cancer cells
    (English)Manuscript (preprint) (Other academic)
    Keywords
    ChIP-Seq, Snail1, Ampliseq, EMtT, BMP6
    National Category
    Cell Biology
    Research subject
    Biochemistry; Biology with specialization in Molecular Cell Biology
    Identifiers
    urn:nbn:se:uu:diva-334433 (URN)
    Available from: 2017-11-23 Created: 2017-11-23 Last updated: 2017-11-30
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  • 242. Maturi, Varun
    et al.
    Enroth, Stefan
    Heldin, Carl-Henrik
    Moustakas, Aristidis
    Genome-wide binding of transcription factor Snail1 in triple-negative breast cancer cellsManuscript (preprint) (Other academic)
  • 243.
    Maturi, Varun
    et al.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Enroth, Stefan
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Heldin, Carl-Henrik
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Moustakas, Aristidis
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Genome-wide binding of transcription factor ZEB1 in triple-negative breast cancer cells2018In: Journal of Cellular Physiology, ISSN 0021-9541, E-ISSN 1097-4652, Vol. 233, no 10, p. 7113-7127Article in journal (Refereed)
    Abstract [en]

    Zinc finger E-box binding homeobox 1 (ZEB1) is a transcriptional regulator involved in embryonic development and cancer progression. ZEB1 induces epithelial-mesenchymal transition (EMT). Triple-negative human breast cancers express high ZEB1 mRNA levels and exhibit features of EMT. In the human triple-negative breast cancer cell model Hs578T, ZEB1 associates with almost 2,000 genes, representing many cellular functions, including cell polarity regulation (DLG2 and FAT3). By introducing a CRISPR-Cas9-mediated 30bp deletion into the ZEB1 second exon, we observed reduced migratory and anchorage-independent growth capacity of these tumor cells. Transcriptomic analysis of control and ZEB1 knockout cells, revealed 1,372 differentially expressed genes. The TIMP metallopeptidase inhibitor 3 and the teneurin transmembrane protein 2 genes showed increased expression upon loss of ZEB1, possibly mediating pro-tumorigenic actions of ZEB1. This work provides a resource for regulators of cancer progression that function under the transcriptional control of ZEB1. The data confirm that removing a single EMT transcription factor, such as ZEB1, is not sufficient for reverting the triple-negative mesenchymal breast cancer cells into more differentiated, epithelial-like clones, but can reduce tumorigenic potential, suggesting that not all pro-tumorigenic actions of ZEB1 are linked to the EMT.

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    fulltext
  • 244. Mehdi, Saher
    et al.
    Derkacheva, Maria
    Ramström, Margareta
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Kralemann, Lejon
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Hennig, Lars
    The WD40 Domain Protein MSI1 Functions in a Histone Deacetylase Complex to Fine-Tune Abscisic Acid Signaling2016In: The Plant Cell, ISSN 1040-4651, E-ISSN 1532-298X, Vol. 28, no 1, p. 42-54Article in journal (Refereed)
    Abstract [en]

    MSI1 belongs to a family of histone binding WD40-repeat proteins. Arabidopsis thaliana contains five genes encoding MSI1-like proteins, but their functions in diverse chromatin-associated complexes are poorly understood. Here, we show that MSI1 is part of a histone deacetylase complex. We copurified HISTONE DEACETYLASE19 (HDA19) with MSI1 and transcriptional regulatory SIN3-like proteins and provide evidence that MSI1 and HDA19 associate into the same complex in vivo. These data suggest that MSI1, HDA19, and HISTONE DEACETYLATION COMPLEX1 protein form a core complex that can integrate various SIN3-like proteins. We found that reduction of MSI1 or HDA19 causes upregulation of abscisic acid (ABA) receptor genes and hypersensitivity of ABA-responsive genes. The MSI1-HDA19 complex fine-tunes ABA signaling by binding to the chromatin of ABA receptor genes and by maintaining low levels of acetylation of histone H3 at lysine 9, thereby affecting the expression levels of ABA receptor genes. Reduced MSI1 or HDA19 levels led to increased tolerance to salt stress corresponding to the increased ABA sensitivity of gene expression. Together, our results reveal the presence of an MSI1-HDA19 complex that fine-tunes ABA signaling in Arabidopsis.

  • 245.
    Migliano, Simona M.
    et al.
    Univ Oslo, Fac Med, Ctr Canc Cell Reprogramming, Oslo, Norway.;Oslo Univ Hosp, Inst Canc Res, Dept Mol Cell Biol, Oslo, Norway..
    Schultz, Sebastian W.
    Univ Oslo, Fac Med, Ctr Canc Cell Reprogramming, Oslo, Norway.;Oslo Univ Hosp, Inst Canc Res, Dept Mol Cell Biol, Oslo, Norway..
    Wenzel, Eva M.
    Univ Oslo, Fac Med, Ctr Canc Cell Reprogramming, Oslo, Norway.;Oslo Univ Hosp, Inst Canc Res, Dept Mol Cell Biol, Oslo, Norway..
    Takáts, Szabolcs
    Univ Oslo, Fac Med, Ctr Canc Cell Reprogramming, Oslo, Norway.;Oslo Univ Hosp, Inst Canc Res, Dept Mol Cell Biol, Oslo, Norway.;Eotvos Lorand Univ, Dept Anat Cell & Dev Biol, Budapest, Hungary..
    Liu, Dan
    Univ Oslo, Fac Med, Ctr Canc Cell Reprogramming, Oslo, Norway.;Oslo Univ Hosp, Inst Canc Res, Dept Mol Cell Biol, Oslo, Norway..
    Mørk, Silje
    Univ Oslo, Fac Med, Ctr Canc Cell Reprogramming, Oslo, Norway.;Oslo Univ Hosp, Inst Canc Res, Dept Mol Cell Biol, Oslo, Norway..
    Tan, Kia Wee
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Univ Oslo, Fac Med, Ctr Canc Cell Reprogramming, Oslo, Norway; Oslo Univ Hosp, Inst Canc Res, Dept Mol Cell Biol, Oslo, Norway.
    Rusten, Tor Erik
    Univ Oslo, Fac Med, Ctr Canc Cell Reprogramming, Oslo, Norway.;Oslo Univ Hosp, Inst Canc Res, Dept Mol Cell Biol, Oslo, Norway..
    Raiborg, Camilla
    Univ Oslo, Fac Med, Ctr Canc Cell Reprogramming, Oslo, Norway.;Oslo Univ Hosp, Inst Canc Res, Dept Mol Cell Biol, Oslo, Norway..
    Stenmark, Harald
    Univ Oslo, Fac Med, Ctr Canc Cell Reprogramming, Oslo, Norway.;Oslo Univ Hosp, Inst Canc Res, Dept Mol Cell Biol, Oslo, Norway..
    Removal of hypersignaling endosomes by simaphagy2024In: Autophagy, ISSN 1554-8627, E-ISSN 1554-8635, Vol. 20, no 4, p. 769-791Article in journal (Refereed)
    Abstract [en]

    Activated transmembrane receptors continue to signal following endocytosis and are only silenced upon ESCRT-mediated internalization of the receptors into intralumenal vesicles (ILVs) of the endosomes. Accordingly, endosomes with dysfunctional receptor internalization into ILVs can cause sustained receptor signaling which has been implicated in cancer progression. Here, we describe a surveillance mechanism that allows cells to detect and clear physically intact endosomes with aberrant receptor accumulation and elevated signaling. Proximity biotinylation and proteomics analyses of ESCRT-0 defective endosomes revealed a strong enrichment of the ubiquitin-binding macroautophagy/autophagy receptors SQSTM1 and NBR1, a phenotype that was confirmed in cell culture and fly tissue. Live cell microscopy demonstrated that loss of the ESCRT-0 subunit HGS/HRS or the ESCRT-I subunit VPS37 led to high levels of ubiquitinated and phosphorylated receptors on endosomes. This was accompanied by dynamic recruitment of NBR1 and SQSTM1 as well as proteins involved in autophagy initiation and autophagosome biogenesis. Light microscopy and electron tomography revealed that endosomes with intact limiting membrane, but aberrant receptor downregulation were engulfed by phagophores. Inhibition of autophagy caused increased intra- and intercellular signaling and directed cell migration. We conclude that dysfunctional endosomes are surveyed and cleared by an autophagic process, simaphagy, which serves as a failsafe mechanism in signal termination.

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  • 246. Minczuk, Michal
    et al.
    He, Jiuya
    Duch, Anna
    Ettema, Thijs
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Evolution.
    Chlebowski, Aleksander
    Dzionek, Karol
    Nijtmans, Leo
    Huynen, Martijn
    Holt, Ian
    TEFM (c17orf42) is necessary for transcription of human mtDNA2011In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 39, no 10, p. 4284-4299Article in journal (Refereed)
    Abstract [en]

    Here we show that c17orf42, hereafter TEFM (transcription elongation factor of mitochondria), makes a critical contribution to mitochondrial transcription. Inactivation of TEFM in cells by RNA interference results in respiratory incompetence owing to decreased levels of H- and L-strand promoter-distal mitochondrial transcripts. Affinity purification of TEFM from human mitochondria yielded a complex comprising mitochondrial transcripts, mitochondrial RNA polymerase (POLRMT), pentatricopeptide repeat domain 3 protein (PTCD3), and a putative DEAD-box RNA helicase, DHX30. After RNase treatment only POLRMT remained associated with TEFM, and in human cultured cells TEFM formed foci coincident with newly synthesized mitochondrial RNA. Based on deletion mutants, TEFM interacts with the catalytic region of POLRMT, and in vitro TEFM enhanced POLRMT processivity on ss- and dsDNA templates. TEFM contains two HhH motifs and a Ribonuclease H fold, similar to the nuclear transcription elongation regulator Spt6. These findings lead us to propose that TEFM is a mitochondrial transcription elongation factor.

  • 247.
    Mohammed, Mubasher
    et al.
    Stockholm Univ, Wenner Gren Inst, Dept Mol Biosci, Stockholm, Sweden..
    Dziedziech, Alexis
    Stockholm Univ, Wenner Gren Inst, Dept Mol Biosci, Stockholm, Sweden..
    Sekar, Vaishnovi
    Stockholm Univ, Wenner Gren Inst, Dept Mol Biosci, Sci Life Lab, Stockholm, Sweden..
    Ernest, Medard
    NIH, Natl Inst Allergy & Infect Dis, Lab Malaria & Vector Res, Rockville, MD 20892 USA..
    Silva, Thiago Luiz
    NIH, Natl Inst Allergy & Infect Dis, Lab Malaria & Vector Res, Rockville, MD 20892 USA..
    Balan, Balu
    Walter & Eliza Hall Inst Med Res, Populat Hlth & Immun Div, Melbourne, Vic, Australia.;Univ Melbourne, Fac Vet & Agr Sci, Parkville, Vic, Australia..
    Emami, S. Noushin
    Stockholm Univ, Wenner Gren Inst, Dept Mol Biosci, Stockholm, Sweden..
    Biryukova, Inna
    Stockholm Univ, Wenner Gren Inst, Dept Mol Biosci, Sci Life Lab, Stockholm, Sweden..
    Friedländer, Marc R.
    Stockholm Univ, Wenner Gren Inst, Dept Mol Biosci, Sci Life Lab, Stockholm, Sweden..
    Jex, Aaron
    Walter & Eliza Hall Inst Med Res, Populat Hlth & Immun Div, Melbourne, Vic, Australia.;Univ Melbourne, Fac Vet & Agr Sci, Parkville, Vic, Australia..
    Jacobs-Lorena, Marcelo
    Johns Hopkins Univ, Bloomberg Sch Publ Hlth, Dept Mol Microbiol & Immunol, Baltimore, MD USA..
    Henriksson, Johan
    Umeå Univ, Dept Mol Biol, Lab Mol Infect Med Sweden MIMS, Umeå, Sweden..
    Vega-Rodriguez, Joel
    NIH, Natl Inst Allergy & Infect Dis, Lab Malaria & Vector Res, Rockville, MD 20892 USA..
    Ankarklev, Johan
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular Evolution. Stockholm Univ, Wenner Gren Inst, Dept Mol Biosci, Stockholm, Sweden.
    Single-Cell Transcriptomics To Define Plasmodium falciparum Stage Transition in the Mosquito Midgut2023In: Microbiology Spectrum, E-ISSN 2165-0497, Vol. 11, no 2Article in journal (Refereed)
    Abstract [en]

    Malaria inflicts the highest rate of morbidity and mortality among the vector-borne diseases. The dramatic bottleneck of parasite numbers that occurs in the gut of the obligatory mosquito vector provides a promising target for novel control strategies. Using single-cell transcriptomics, we analyzed Plasmodium falciparum development in the mosquito gut, from unfertilized female gametes through the first 20 h after blood feeding, including the zygote and ookinete stages. This study revealed the temporal gene expression of the ApiAP2 family of transcription factors and of parasite stress genes in response to the harsh environment of the mosquito midgut. Further, employing structural protein prediction analyses, we found several upregulated genes predicted to encode intrinsically disordered proteins (IDPs), a category of proteins known for their importance in regulation of transcription, translation, and protein-protein interactions. IDPs are known for their antigenic properties and may serve as suitable targets for antibody- or peptide-based transmission suppression strategies. In total, this study uncovers the P. falciparum transcriptome from early to late parasite development in the mosquito midgut, inside its natural vector, which provides an important resource for future malaria transmission-blocking initiatives.

    IMPORTANCE The malaria parasite Plasmodium falciparum causes more than half a million deaths per year. The current treatment regimen targets the symptom-causing blood stage inside the human host. However, recent incentives in the field call for novel interventions to block parasite transmission from humans to the mosquito vector. Therefore, we need to better understand the parasite biology during its development inside the mosquito, including a deeper understanding of the expression of genes controlling parasite progression during these stages. Here, we have generated single-cell transcriptome data, covering P. falciparum’s development, from gamete to ookinete inside the mosquito midgut, uncovering previously untapped parasite biology, including a repertoire of novel biomarkers to be explored in future transmission-blocking efforts. We anticipate that our study provides an important resource, which can be further explored to improve our understanding of the parasite biology as well as aid in guiding future malaria intervention strategies.

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    FULLTEXT01
  • 248. Mohell, N.
    et al.
    Alfredsson, J.
    Fransson, A.
    Uustalu, M.
    Bystrom, S.
    Gullbo, Joachim
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Experimental and Clinical Oncology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Cancer Pharmacology and Computational Medicine.
    Hallberg, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Organic Pharmaceutical Chemistry.
    Bykov, V. J. N.
    Bjorklund, U.
    Wiman, K. G.
    APR-246 overcomes resistance to cisplatin and doxorubicin in ovarian cancer cells2015In: Cell Death and Disease, ISSN 2041-4889, E-ISSN 2041-4889, Vol. 6, article id e1794Article in journal (Refereed)
    Abstract [en]

    Two main causes of platinum resistance are mutation in the tumor suppressor gene TP53 and drug-induced increase in intracellular glutathione concentration. Mutations in TP53 occur in about 50% of human tumors. APR-246 (PRIMA-1(MET)) is the first clinical-stage compound that reactivates mutant p53 and induces apoptosis. APR-246 is a prodrug that is converted to the active compound methylene quinuclidinone (MQ), a Michael acceptor that binds to cysteine residues in mutant p53 and restores its wildtype conformation. Here, we show that MQ also binds to cysteine in glutathione, thus decreasing intracellular free glutathione concentration. We also show that treatment with APR-246 completely restores the cisplatin and doxorubicin sensitivity to p53-mutant drug-resistant ovarian cancer cells. We propose that this unique ability of APR-246/MQ to bind to cysteines in both mutant p53 and glutathione has a key role in the resensitization as well as in the outstanding synergistic effects observed with APR-246 in combination with platinum compounds in ovarian cancer cell lines and primary cancer cells. However, MQ binding to cysteines in other targets, for example, thioredoxin reductase, may contribute as well. Strong synergy was also observed with the DNA-damaging drugs doxorubicin and gemcitabine, while additive effects were found with the taxane docetaxel. Our results provide a strong rationale for the ongoing clinical study with APR-246 in combination with platinum-based therapy in patients with p53-mutant recurrent high-grade serous (HGS) ovarian cancer. More than 96% of these patients carry TP53 mutations. Combined treatment with APR-246 and platinum or other DNA-damaging drugs could allow dramatically improved therapy of a wide range of therapy refractory p53 mutant tumors.

    Download full text (pdf)
    fulltext
  • 249. Molnar, Matyas
    et al.
    Chen, Yun
    Friberg, Peter
    L. M., Gan
    Fu, Ying
    Molecular imaging of inflammation-activated endothelium using bio-conjugated quantum dotsManuscript (preprint) (Other academic)
  • 250. Molnar, Matyas
    et al.
    Chen, Yun
    Friberg, Peter
    L. M., Gan
    Fu, Ying
    Significant blue-shift of luminescence peak from VCAM-1-binding peptide-functionalized quantum dots upon binding with VCAM-1Manuscript (preprint) (Other academic)
2345678 201 - 250 of 426
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