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Publications (10 of 98) Show all publications
Wåhlén, E., Olsson, F., Raykova, D., Söderberg, O., Heldin, J. & Lennartsson, J. (2023). Activated EGFR and PDGFR internalize in separate vesicles and downstream AKT and ERK1/2 signaling are differentially impacted by cholesterol depletion. Biochemical and Biophysical Research Communications - BBRC, 665, 195-201
Open this publication in new window or tab >>Activated EGFR and PDGFR internalize in separate vesicles and downstream AKT and ERK1/2 signaling are differentially impacted by cholesterol depletion
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2023 (English)In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 665, p. 195-201Article in journal (Refereed) Published
Abstract [en]

The interplay between membrane subregions and receptor tyrosine kinases (RTK) will influence signaling in both normal and pathological RTK conditions. In this study, epidermal growth factor receptor (EGFR) and platelet-derived growth factor receptor β (PDGFR-β) internalizations were investigated by immunofluorescent microscopy following simultaneous treatment with EGF and PDGF-BB. We found that the two receptors utilize separate routes of internalization, which merges in a common perinuclear endosomal compartment after 45 min of stimulation. This is further strengthened when contrasting the recruitment of either EGFR or PDGFR-β to either clathrin or caveolin-1: PDGFR-β dissociates from caveolin-1 upon stimulation, and engages clathrin, whilst an increased recruitment of EGFR, to both clathrin and caveolin-1, was observed upon EGF stimulation. The association between EGFR and caveolin-1 is supported by the observation that EGFR was localized in lipid raft associated fractions, whereas PDGFR-β was not. We also found that disruption of lipid rafts using MβCD led to an increased EGFR dimerization and phosphorylation in response to ligand, as well as a dramatic decrease in AKT- and a smaller but robust decrease in ERK1/2 phosphorylation. This suggest that lipid rafts may be important to effectively connect the EGFR with downstream proteins to facilitate signaling. Our data implies that cholesterol depletion of the plasma membrane affect the signaling of EGFR and PDGFRβ differently.

Place, publisher, year, edition, pages
Elsevier, 2023
Keywords
EGFR, EGF, PDGFR, PDGF, Membrane raft, Lipid rafts, Receptor tyrosine kinase, Internalization
National Category
Cell Biology Pharmacology and Toxicology
Identifiers
urn:nbn:se:uu:diva-506912 (URN)10.1016/j.bbrc.2023.04.099 (DOI)001004994200001 ()37163940 (PubMedID)
Funder
Swedish Cancer Society, 21 1427 Pj 01HSwedish Cancer Society, 22 2306 Pj
Available from: 2023-07-03 Created: 2023-07-03 Last updated: 2023-07-03Bibliographically approved
Mihalič, F., Simonetti, L., Giudice, G., Rubin Sander, M., Lindqvist, R., Peters, M. B., . . . Ivarsson, Y. (2023). Large-scale phage-based screening reveals extensive pan-viral mimicry of host short linear motifs. Nature Communications, 14(1), Article ID 2409.
Open this publication in new window or tab >>Large-scale phage-based screening reveals extensive pan-viral mimicry of host short linear motifs
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2023 (English)In: Nature Communications, E-ISSN 2041-1723, Vol. 14, no 1, article id 2409Article in journal (Refereed) Published
Abstract [en]

Viruses mimic host short linear motifs (SLiMs) to hijack and deregulate cellular functions. Studies of motif-mediated interactions therefore provide insight into virus-host dependencies, and reveal targets for therapeutic intervention. Here, we describe the pan-viral discovery of 1712 SLiM-based virus-host interactions using a phage peptidome tiling the intrinsically disordered protein regions of 229 RNA viruses. We find mimicry of host SLiMs to be a ubiquitous viral strategy, reveal novel host proteins hijacked by viruses, and identify cellular pathways frequently deregulated by viral motif mimicry. Using structural and biophysical analyses, we show that viral mimicry-based interactions have similar binding strength and bound conformations as endogenous interactions. Finally, we establish polyadenylate-binding protein 1 as a potential target for broad-spectrum antiviral agent development. Our platform enables rapid discovery of mechanisms of viral interference and the identification of potential therapeutic targets which can aid in combating future epidemics and pandemics.

Place, publisher, year, edition, pages
Springer Nature, 2023
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-503184 (URN)10.1038/s41467-023-38015-5 (DOI)000979744000013 ()37100772 (PubMedID)
Funder
Swedish Foundation for Strategic Research, SB16-0039Swedish Research Council, 2018-05851Swedish Research Council, 2020.0182Swedish Research Council, VR-RFI 2016-00968Knut and Alice Wallenberg Foundation, NNF14CC0001Knut and Alice Wallenberg Foundation
Available from: 2023-06-30 Created: 2023-06-30 Last updated: 2023-06-30Bibliographically approved
Leino, M. & Söderberg, O. (2023). Purification of DNA oligonucleotides to improve hybridization chain reaction performance. New Biotechnology, 76, 33-40
Open this publication in new window or tab >>Purification of DNA oligonucleotides to improve hybridization chain reaction performance
2023 (English)In: New Biotechnology, ISSN 1871-6784, E-ISSN 1876-4347, Vol. 76, p. 33-40Article in journal (Refereed) Published
Abstract [en]

Hybridization chain-reaction (HCR) is technique to generate a linear polymerization of oligonucleotide hairpins, used in multiple molecular biology methods. The HCR reaction is dependent on that every hairpin is metastable in the absence of a triggering oligonucleotide and that every hairpin can continue the polymerization, which places a strong demand on oligonucleotide quality. In this paper we show how further purification can greatly increase polymerization potential. We found that a single extra PAGE-purification could greatly enhance hairpin polymerization both in solution and in situ. Purification using a ligation-based method further improved polymerization, yielding in situ immunoHCR stains at least 3.4-times stronger than non-purified control. This demonstrates the importance of not only good sequence design of the oligonucleotide hairpins, but also the demand for high quality oligonucleotides to accomplish a potent and specific HCR.

Place, publisher, year, edition, pages
Elsevier, 2023
Keywords
DNA hairpin, hybridization chain reaction, in situ hybridization, purification
National Category
Biochemistry and Molecular Biology
Research subject
Biology with specialization in Molecular Biotechnology
Identifiers
urn:nbn:se:uu:diva-496885 (URN)10.1016/j.nbt.2023.04.004 (DOI)000983409800001 ()37059331 (PubMedID)
Funder
Swedish Cancer Society, 22 2306 PjSwedish Research Council, 2017-01775
Available from: 2023-02-22 Created: 2023-02-22 Last updated: 2023-05-26Bibliographically approved
Raykova, D., Kermpatsou, D., Malmqvist, T., Harrison, P. J., Rubin Sander, M., Stiller, C., . . . Söderberg, O. (2022). A method for Boolean analysis of protein interactions at a molecular level. Nature Communications, 13(1), Article ID 4755.
Open this publication in new window or tab >>A method for Boolean analysis of protein interactions at a molecular level
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2022 (English)In: Nature Communications, E-ISSN 2041-1723, Vol. 13, no 1, article id 4755Article in journal (Refereed) Published
Abstract [en]

Determination of interactions between native proteins in cells is important for understanding function. Here the authors report MolBoolean as a method to detect interactions between endogenous proteins in subcellular compartments, using antibody-DNA conjugates for identification and signal amplification. Determining the levels of protein-protein interactions is essential for the analysis of signaling within the cell, characterization of mutation effects, protein function and activation in health and disease, among others. Herein, we describe MolBoolean - a method to detect interactions between endogenous proteins in various subcellular compartments, utilizing antibody-DNA conjugates for identification and signal amplification. In contrast to proximity ligation assays, MolBoolean simultaneously indicates the relative abundances of protein A and B not interacting with each other, as well as the pool of A and B proteins that are proximal enough to be considered an AB complex. MolBoolean is applicable both in fixed cells and tissue sections. The specific and quantifiable data that the method generates provide opportunities for both diagnostic use and medical research.

Place, publisher, year, edition, pages
Springer Nature, 2022
National Category
Biochemistry and Molecular Biology Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:uu:diva-482674 (URN)10.1038/s41467-022-32395-w (DOI)000840338100011 ()35963857 (PubMedID)
Funder
Swedish Foundation for Strategic ResearchSwedish Cancer SocietySwedish Research Council
Note

Correction in: Nature Communications volume 14, Article number: 5450 (2023)

DOI: 10.1038/s41467-023-41325-3

Available from: 2022-09-20 Created: 2022-09-20 Last updated: 2023-10-24Bibliographically approved
Wåhlén, E., Olsson, F., Söderberg, O., Lennartsson, J. & Heldin, J. (2022). Differential impact of lipid raft depletion on platelet-derived growth factor (PDGF)-induced ERK1/2 MAP-kinase, SRC and AKT signaling. Cellular Signalling, 96, Article ID 110356.
Open this publication in new window or tab >>Differential impact of lipid raft depletion on platelet-derived growth factor (PDGF)-induced ERK1/2 MAP-kinase, SRC and AKT signaling
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2022 (English)In: Cellular Signalling, ISSN 0898-6568, E-ISSN 1873-3913, Vol. 96, article id 110356Article in journal (Refereed) Published
Abstract [en]

It has become clear that lipid rafts functions as signaling hotspots connecting cell surface receptors to intracellular signaling pathways. However, the exact involvement of lipid rafts in receptor tyrosine kinase signaling is still poorly understood. In this study, we have analyzed platelet-derived growth factor (PDGF) receptor β (PDGFR-β) signaling in two different cell lines depleted of cholesterol, and as a consequence, disruption of lipid rafts. Cholesterol depletion of BJ-hTERT fibroblasts using methyl-β-cyclodextrin (MβCD) did not affect PDGFR-β activation as measured by its tyrosine phosphorylation. However, we did observe a small reduction in AKT phosphorylation and a more robust decrease of ERK1/2 activation. In contrast, in the osteosarcoma cell line U2OS, we noticed a deficient receptor activation. Interestingly, in U2OS cells, the ERK1/2 pathway was unaffected, but instead AKT and SRC signaling was reduced. These results suggest that cell type specific wiring of signaling pathways can lead to differential sensitivity to cholesterol depletion. Furthermore, MβCD treatment had a much more pronounced morphological effect on U2OS compared to BJ-hTERT cells. This is consistent with a previous report claiming that cancer cells are more sensitive to cholesterol depletion than normal cells. Our data supports the possibility that cholesterol lowering drugs may impede tumor growth.

Place, publisher, year, edition, pages
Elsevier, 2022
Keywords
PDGFR, Lipid rafts, Membrane rafts, M?CD, BJ-hTERT and U2OS
National Category
Cell Biology Cell and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-478575 (URN)10.1016/j.cellsig.2022.110356 (DOI)000808581500004 ()35605761 (PubMedID)
Funder
Swedish Research CouncilSwedish Cancer Society, CAN2018/425
Note

De två sista författarna delar sistaförfattarskapet

Correction in: Cellular Signalling, vol. 98, article-id 110411

doi: 10.1016/j.cellsig.2022.110411

Available from: 2022-06-28 Created: 2022-06-28 Last updated: 2023-04-14Bibliographically approved
Frances-Soriano, L., Leino, M., Dos Santos, M. C., Kovacs, D., Borbas, K. E., Söderberg, O. & Hildebrandt, N. (2021). In Situ Rolling Circle Amplification Förster Resonance Energy Transfer (RCA-FRET) for Washing-Free Real-Time Single-Protein Imaging. Analytical Chemistry, 93(3), 1842-1850
Open this publication in new window or tab >>In Situ Rolling Circle Amplification Förster Resonance Energy Transfer (RCA-FRET) for Washing-Free Real-Time Single-Protein Imaging
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2021 (English)In: Analytical Chemistry, ISSN 0003-2700, E-ISSN 1520-6882, Vol. 93, no 3, p. 1842-1850Article in journal (Refereed) Published
Abstract [en]

Fluorescence signal enhancement via isothermal nucleic acid amplification is an important approach for sensitive imaging of intra- or extracellular nucleic acid or protein biomarkers. Rolling circle amplification (RCA) is frequently applied for fluorescence in situ imaging but faces limitations concerning multiplexing, dynamic range, and the required multiple washing steps before imaging. Here, we show that Forster resonance energy transfer (FRET) between fluorescent dyes and between lanthanide (Ln) complexes and dyes that hybridize to beta-actin-specific RCA products in HaCaT cells can afford washing-free imaging of single beta-actin proteins. Proximity-dependent FRET could be monitored directly after or during (real-time monitoring) dye or Ln DNA probe incubation and could efficiently distinguish between photoluminescence from beta-actin-specific RCA and DNA probes freely diffusing in solution or nonspecifically attached to cells. Moreover, time-gated FRET imaging with the Ln-dye FRET pairs efficiently suppressed sample autofluorescence and improved the signal-to-background ratio. Our results present an important proof of concept of RCA-FRET imaging with a strong potential to advance in situ RCA toward easier sample preparation, higher-order multiplexing, autofluorescence-free detection, and increased dynamic range by real-time monitoring of in situ RCA.

Place, publisher, year, edition, pages
American Chemical Society (ACS)American Chemical Society (ACS), 2021
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-437243 (URN)10.1021/acs.analchem.0c04828 (DOI)000613922400082 ()33356162 (PubMedID)
Funder
Swedish Research CouncilSwedish Cancer Society
Available from: 2021-03-12 Created: 2021-03-12 Last updated: 2024-01-15Bibliographically approved
Bivehed, E., Söderberg, O. & Hellman, B. (2020). Flash-comet: Significantly improved speed and sensitivity of the comet assay through the introduction of lithium-based solutions and a more gentle lysis. Mutation research. Genetic toxicology and environmental mutagenesis, 858, Article ID 503240.
Open this publication in new window or tab >>Flash-comet: Significantly improved speed and sensitivity of the comet assay through the introduction of lithium-based solutions and a more gentle lysis
2020 (English)In: Mutation research. Genetic toxicology and environmental mutagenesis, ISSN 1383-5718, E-ISSN 1879-3592, Vol. 858, article id 503240Article in journal (Refereed) Published
Abstract [en]

Evaluation of primary DNA-damage is one way to identify potential genotoxic agents and for this purpose the Comet assay has, for the last decades, been used to monitor DNA single strand and double strand breaks in individual cells. Various attempts have been made to modify the different steps in the in vitro protocol for the Comet assay in order to improve its sensitivity. However, to the best of our knowledge, nobody has tried to replace the traditionally used NaOH-based electrophoresis solution (pH > 13), with another type of solution. In the present paper, using TK-6 cells exposed to different concentrations of H2O2 or ionizing radiation, we present evidence clearly showing that a low-conductive LiOH-based electrophoresis solution at pH 12.5, and a more gentle lysis procedure, significantly improved both the speed and sensitivity of the assay. The new approach, which we call the Flash-comet, is based on a lysis buffer at pH 8.5, an unwinding time of 2.5 min in a LiOH solution without EDTA at pH 12.5, and an electrophoresis time of 1 min at 150 V (5 V/cm) using the same solution.

Place, publisher, year, edition, pages
ELSEVIER, 2020
Keywords
DNA-damage, Flash-comet assay, Low conductivity electrophoresis solution, Method development, Lithium hydroxide, Genotoxicity testing
National Category
Pharmacology and Toxicology
Identifiers
urn:nbn:se:uu:diva-428302 (URN)10.1016/j.mrgentox.2020.503240 (DOI)000591710900003 ()33198930 (PubMedID)
Funder
Swedish Research Council
Note

Correction in: MUTATION RESEARCH-GENETIC TOXICOLOGY AND ENVIRONMENTAL MUTAGENESIS, Volume:863, Article Number:503323, DOI:10.1016/j.mrgentox.2021.503323

Available from: 2020-12-15 Created: 2020-12-15 Last updated: 2023-09-03Bibliographically approved
Heldin, J., Rubin Sander, M., Leino, M., Thomsson, S., Lennartsson, J. & Söderberg, O. (2019). Dynamin inhibitors impair platelet-derived growth factor beta-receptor dimerization and signaling. Experimental Cell Research, 380(1), 69-79
Open this publication in new window or tab >>Dynamin inhibitors impair platelet-derived growth factor beta-receptor dimerization and signaling
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2019 (English)In: Experimental Cell Research, ISSN 0014-4827, E-ISSN 1090-2422, Vol. 380, no 1, p. 69-79Article in journal (Refereed) Published
Abstract [en]

The role of plasma membrane composition and dynamics in the activation process of receptor tyrosine kinases (RTKs) is still poorly understood. In this study we have investigated how signaling via the RTK, platelet-derived growth factor beta-receptor (PDGFR-beta) is affected by Dynasore or Dyngo-4a, which are commonly used dynamin inhibitors. PDGFR-beta preferentially internalizes via clathrin-coated pits and in this pathway, Dynamin II has a major role in the formation and release of vesicles from the plasma membrane by performing the membrane scission. We have found that dynamin inhibitors impedes the activation of PDGFR-beta by impairing ligand-induced dimerization of the receptor monomers, which leads to a subsequent lack of phosphorylation and activation both of receptors and downstream effectors, such as ERK1/2 and AKT. In contrast, dynamin inhibitors did not affect epidermal growth factor receptor (EGFR) dimerization and phosphorylation. Our findings suggest that there is a link between plasma membrane dynamics and PDGFR-beta activation, and that this link is not shared with the epidermal growth factor receptor.

Place, publisher, year, edition, pages
ELSEVIER INC, 2019
Keywords
RTK signaling, EGFR, PDGFR-beta, Dynasore, Dyngo, Dimerization
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-384986 (URN)10.1016/j.yexcr.2019.04.004 (DOI)000468124700008 ()30970237 (PubMedID)
Funder
Swedish Foundation for Strategic Research Swedish Research CouncilSwedish Cancer Society, CAN 2018/425
Available from: 2019-06-14 Created: 2019-06-14 Last updated: 2023-03-10Bibliographically approved
Wu, C.-C., Klaesson, A., Buskas, J., Ranefall, P., Mirzazadeh, R., Söderberg, O. & Wolf, J. B. W. (2019). In situ quantification of individual mRNA transcripts in melanocytes discloses gene regulation of relevance to speciation. Journal of Experimental Biology, 222(5)
Open this publication in new window or tab >>In situ quantification of individual mRNA transcripts in melanocytes discloses gene regulation of relevance to speciation
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2019 (English)In: Journal of Experimental Biology, ISSN 0022-0949, E-ISSN 1477-9145, Vol. 222, no 5Article in journal (Refereed) Published
National Category
Genetics
Identifiers
urn:nbn:se:uu:diva-381095 (URN)10.1242/jeb.194431 (DOI)000461414600021 ()30718374 (PubMedID)
Funder
EU, European Research Council, ERCStG-336536Swedish Research Council, 621-2013-4510Knut and Alice Wallenberg Foundation
Available from: 2019-03-08 Created: 2019-04-15 Last updated: 2022-01-29Bibliographically approved
Leino, M., Heldin, J., Rubin Sander, M., Kermpatsou, D., Raykova, D., Koos, B. & Söderberg, O. (2019). Optimization of proximity-dependent initiation of hybridization chain reaction for improved performance. Molecular Systems Design & Engineering , 4(5), 1058-1065
Open this publication in new window or tab >>Optimization of proximity-dependent initiation of hybridization chain reaction for improved performance
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2019 (English)In: Molecular Systems Design & Engineering , E-ISSN 2058-9689, Vol. 4, no 5, p. 1058-1065Article in journal (Refereed) Published
Abstract [en]

Proximity based detection methods are invaluable tools in the field of molecular biology, increasing selectivity and allowing for analysis of protein interactions. ProxHCR utilizes pairs of antibodies labelled with oligonucleotides to probe for proximal binding and to initiate a hybridization chain reaction (HCR) to generate an amplified detection signal. As HCR is based upon hybridization of DNA hairpins, the performance is dependent on salt concentrations and temperature. Herein we have redesigned the proxHCR system to increase the performance and to reduce dependency on temperature and salt concentrations. The new oligonucleotides provide an increased signal when performed at physiological salt concentrations and in room temperature.

National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-396655 (URN)10.1039/c9me00079h (DOI)000489041600007 ()
Funder
Swedish Foundation for Strategic Research Swedish Research Council
Available from: 2019-11-14 Created: 2019-11-14 Last updated: 2023-03-01Bibliographically approved
Projects
Next generation tissue profiling [2014-02968_VR]; Uppsala Universitysingle cell biochemistry [2017-00653_VR]; Uppsala UniversityMolecular evaluation of cell signaling in tumors [2017-01775_VR]; Uppsala University
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0003-2883-1925

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