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  • 101.
    Davis, Hayley
    et al.
    Univ Oxford, Wellcome Trust Ctr Human Genet, Gastrointestinal Stem Cell Biol Lab, Oxford, England..
    Raja, Erna
    Univ Tokyo, Grad Sch Med, Dept Mol Pathol, Tokyo, Japan..
    Miyazono, Kohei
    Univ Tokyo, Grad Sch Med, Dept Mol Pathol, Tokyo, Japan..
    Tsubakihara, Yutaro
    Ehime Univ, Grad Sch Med, Dept Mol Med Pathogenesis, Matsuyama, Ehime 790, Japan.;Uppsala Univ, Biomed Ctr, Sci Life Lab, Ludwig Inst Canc Res, Box 595, S-75124 Uppsala, Sweden..
    Moustakas, Aristidis
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Mechanisms of action of bone morphogenetic proteins in cancer2016In: Cytokine & growth factor reviews, ISSN 1359-6101, E-ISSN 1879-0305, Vol. 27, p. 81-92Article, review/survey (Refereed)
    Abstract [en]

    The bone morphogenetic proteins (BMPs) play fundamental roles in embryonic development and control differentiation of a diverse set of cell types. It is therefore of no surprise that the BMPs also contribute to the process of tumourigenesis and regulate cancer progression through various stages. We summarise here key roles of BMP ligands, receptors, their signalling mediators, mainly focusing on proteins of the Smad family, and extracellular antagonists, that contribute to the onset of tumourigenesis and to cancer progression in diverse tissues. Overall, the BMP pathways seem to act as tumour suppressors that maintain physiological tissue homeostasis and which are perturbed in cancer either via genetic mutation or via epigenetic misregulation of key gene components. BMPs also control the self-renewal and fate choices made by stem cells in several tissues. By promoting cell differentiation, including inhibition of the process of epithelial-mesenchymal transition, BMPs contribute to the malignant progression of cancer at advanced stages. It is therefore reasonable that pharmaceutical industries continuously develop biological agents and chemical modulators of BMP signalling with the aim to improve therapeutic regimes against several types of cancer.

  • 102.
    Davoodpour, Padideh
    Uppsala University, Units outside the University, Ludwig Institute for Cancer Research.
    2-ME-Induced Apoptotic Signalling in Prostate Cancer PC3 Cells2005Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Prostate cancer is common in the Western society and current treatments are often associated with side effects, therefore improved therapeutic strategies are desired. 2-methoxyestradiol (2-ME), an endogenous metabolite of estradiol-17β inhibits tumor growth in vivo as it prevents angiogenesis. 2-ME has also direct cytotoxic effects on tumor cells. In this study, we have investigated the potential use of PET to record effects 2-ME on prostate cancer cell (PC3) aggregates. The anti-proliferative and pro-apoptotic effects of 2-ME on PC3 cell aggregates in vitro were correlated with the uptake of deoxy-D-glucose, FMAU and choline labeled with 18F, 11C or 3H. 2-ME clearly reduced growth of PC3 aggregates and induced apoptosis in a dose-dependent manner. However, the PET tracers failed to record the cytotoxicity of 2-ME on PC3 aggregates.

    Further, the signaling events responsible for 2-ME induced prostate cancer cell death were investigated. We found that Smad7, previously implicated in TGF-β-induced responses, is required for 2-ME-induced p38 MAPK activation and subsequent apoptosis in PC-3U cells, as shown by the use of antisense or siRNA techniques and a specific inhibitor of p38 MAPK (SB203580). Interestingly, Smad7 also regulated the expression of the pro-apoptotic Bim protein.

    Shb is a Src Homology 2 domain adapter protein with pro-apoptotic effects. PC3 clones overexpressing Shb exhibited increased rates of apoptosis, both in the presence or absence of 2-ME, as they failed to activate survival mechanisms through ERK and Akt in response to 2-ME. Notably, Shb cells displayed increased activity of the pro-apoptotic kinase c-Abl. Pre-treatment with SB203580 or c-Abl (STI-571) inhibitors completely blocked the apoptotic response to 2-ME.

    In conclusion, Smad7 and Shb appear to be crucial for 2-ME-induced PC3 cell apoptosis via their activation of p38 MAPK and c-Abl. Future therapies exploring these pathways can be envisaged as treatment of prostate cancer.

    List of papers
    1. Effects of 2-ME on proliferation, apoptosis and PET-tracer uptake in human prostate cancer cell aggregates
    Open this publication in new window or tab >>Effects of 2-ME on proliferation, apoptosis and PET-tracer uptake in human prostate cancer cell aggregates
    2004 In: Nuclear Medicine and Biology, ISSN 0969-8051, Vol. 7, no Oct 31, p. 867-874Article in journal (Refereed) Published
    Identifiers
    urn:nbn:se:uu:diva-93755 (URN)
    Available from: 2005-11-17 Created: 2005-11-17Bibliographically approved
    2. 2-Methoxyestradiol-induced apoptosis in prostate cancer cells requires Smad7
    Open this publication in new window or tab >>2-Methoxyestradiol-induced apoptosis in prostate cancer cells requires Smad7
    2005 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 280, no 15, p. 14773-14779Article in journal (Refereed) Published
    Abstract [en]

    Prostate cancer is the second most common cause of death related to cancer in Western society. 2-Methoxyestradiol (2-ME), an endogenous metabolite of estradiol-17beta, inhibits tumor angiogenesis while also exerting potent cytotoxic effects on various cancer cells. 2-ME has been shown to activate the p38 MAPK and JNK pathways and to induce apoptosis in cells, although the underlying molecular mechanisms for this are unknown. Here we report that the expression of Smad7, an adaptor molecule required to activate p38 MAPK in the transforming growth factor beta signaling pathway, is also required for 2-ME-induced p38 activation and apoptosis in human prostate cancer cells (PC-3U). PC-3U/AS-S7 cells stably transfected with an antisense Smad7 construct, or PC-3U cells transiently transfected with short interfering RNA for Smad7, were protected against 2-ME-induced apoptosis. 2-ME-induced apoptosis was found to involve p38 MAPK and JNK, because simultaneous treatments with 2-ME and a specific p38 inhibitor (SB203580) or an inhibitor of JNK (L-JNK1) prevented 2-ME-induced apoptosis. Most interestingly, Smad7 was shown by both antisense and short interfering RNA techniques to affect levels of beta-catenin, which has been implicated previously in the regulation of apoptosis. Moreover, Smad7 was found to be important for the basal expression of Bim, a pro-apoptotic Bcl-2 family member, and for 2-ME-induced expression of Bim. These results suggest that expression of Smad7 is crucial for 2-ME-induced apoptosis in human prostate cancer cells.

    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-93756 (URN)10.1074/jbc.M414470200 (DOI)15708859 (PubMedID)
    Available from: 2005-11-17 Created: 2005-11-17 Last updated: 2017-12-14Bibliographically approved
    3. Increased apoptosis and C-Abl activity in PC3 prostate cancer overexpressing the Shb adapter protein
    Open this publication in new window or tab >>Increased apoptosis and C-Abl activity in PC3 prostate cancer overexpressing the Shb adapter protein
    Manuscript (Other academic)
    Identifiers
    urn:nbn:se:uu:diva-93757 (URN)
    Available from: 2005-11-17 Created: 2005-11-17 Last updated: 2010-01-13Bibliographically approved
  • 103.
    Davoodpour, Padideh
    et al.
    Uppsala University, Units outside the University, Ludwig Institute for Cancer Research.
    Bergström, Mats
    Landström, Marene
    Effects of 2-ME on proliferation, apoptosis and PET-tracer uptake in human prostate cancer cell aggregates2004In: Nuclear Medicine and Biology, ISSN 0969-8051, Vol. 7, no Oct 31, p. 867-874Article in journal (Refereed)
  • 104.
    Davoodpour, Padideh
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Bergström, Mats
    Landström, Maréne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Effects of 2-methoxyestradiol on proliferation, apoptosis and PET-tracer uptake in human prostate cancer cell aggregates2004In: Nuclear Medicine and Biology, ISSN 0969-8051, E-ISSN 1872-9614, Vol. 31, no 7, p. 867-874Article in journal (Refereed)
    Abstract [en]

    The purpose of this study was to investigate the potential use of PET in vivo to record cytotoxic effects of 2-methoxyestradiol (2-ME), an endogenous metabolite of 17beta-estradiol. The anti-proliferative and pro-apoptotic effects of 2-ME on human prostate cancer cell (PC3) aggregates in vitro, were correlated with the uptake of fluoro-deoxy-D-glucose, FMAU and choline labelled with 18F, 11C, or 3H. 2-ME clearly reduced growth of PC3 aggregates and induced apoptosis in a dose-dependent manner. However, the uptake of the putative proliferation markers 11C-FMAU or 3H-choline failed to record the growth inhibitory effects of 2-ME on PC3 cell aggregates. The uptake of 18F-FDG was used as a marker for effects on cellular metabolism and also failed to show any dose-dependent effects in PC3 aggregates. The use of these PET-tracers in vivo is therefore not recommended in order to evaluate the cytotoxic effects of 2-ME on human prostate cancer cells.

  • 105.
    Davoodpour, Padideh
    et al.
    Uppsala University, Units outside the University, Ludwig Institute for Cancer Research.
    Landström, Marene
    Welsh, Michael
    Increased apoptosis and C-Abl activity in PC3 prostate cancer overexpressing the Shb adapter proteinManuscript (Other academic)
  • 106.
    Davoodpour, Padideh
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Landström, Maréne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    2-Methoxyestradiol-induced apoptosis in prostate cancer cells requires Smad72005In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 280, no 15, p. 14773-14779Article in journal (Refereed)
    Abstract [en]

    Prostate cancer is the second most common cause of death related to cancer in Western society. 2-Methoxyestradiol (2-ME), an endogenous metabolite of estradiol-17beta, inhibits tumor angiogenesis while also exerting potent cytotoxic effects on various cancer cells. 2-ME has been shown to activate the p38 MAPK and JNK pathways and to induce apoptosis in cells, although the underlying molecular mechanisms for this are unknown. Here we report that the expression of Smad7, an adaptor molecule required to activate p38 MAPK in the transforming growth factor beta signaling pathway, is also required for 2-ME-induced p38 activation and apoptosis in human prostate cancer cells (PC-3U). PC-3U/AS-S7 cells stably transfected with an antisense Smad7 construct, or PC-3U cells transiently transfected with short interfering RNA for Smad7, were protected against 2-ME-induced apoptosis. 2-ME-induced apoptosis was found to involve p38 MAPK and JNK, because simultaneous treatments with 2-ME and a specific p38 inhibitor (SB203580) or an inhibitor of JNK (L-JNK1) prevented 2-ME-induced apoptosis. Most interestingly, Smad7 was shown by both antisense and short interfering RNA techniques to affect levels of beta-catenin, which has been implicated previously in the regulation of apoptosis. Moreover, Smad7 was found to be important for the basal expression of Bim, a pro-apoptotic Bcl-2 family member, and for 2-ME-induced expression of Bim. These results suggest that expression of Smad7 is crucial for 2-ME-induced apoptosis in human prostate cancer cells.

  • 107.
    Davoodpour, Padideh
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Landström, Maréne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Reduced tumor growth in vivo and increased c-Abl activity in PC3 prostate cancer cells overexpressing the Shb adapter protein2007In: BMC Cancer, ISSN 1471-2407, E-ISSN 1471-2407, Vol. 7, p. 161-Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Induction of apoptosis is one strategy for treatment of prostate cancer. The Shb adapter protein has been found to regulate apoptosis in various cell types and consequently human prostate cancer 3 (PC3) cells were transfected to obtain cells overexpressing Shb in order to increase our understanding of the mechanisms regulating PC3 cell apoptosis. METHODS: Human prostate cancer cells (PC3) were transfected with control vector or a vector containing the Shb cDNA. Clones overexpressing Shb were studied with respect to apoptosis (Dapi, M30) and c-Abl activation (Western blot for pY-245-Abl). The cells were exposed to the anti-tumor agent 2-methoxyestradiol (2-ME) and the p38 MAPK and c-Abl inhibitors SB203580 and STI-571, respectively, after which cell death was determined. In vivo tumor growth and tumor cell proliferation (Ki-67 staining) or apoptosis (active caspase 3 staining) were also determined in nude mice. RESULTS: PC3 cells overexpressing Shb exhibited increased rates of apoptosis in the presence of the anti-tumor agent 2-ME. The Shb cells displayed increased activity of the pro-apoptotic kinase c-Abl. Pre-treatment with p38 MAPK (SB203580) or c-Abl (STI-571) inhibitors completely blocked 2-ME-induced apoptosis, implicating these two pathways in the response. The PC3-Shb cells displayed reduced tumor growth in vivo, an effect occurring as a consequence of increased apoptosis and reduced DNA synthesis. CONCLUSION: It is concluded that Shb promotes 2-ME-induced PC3 cell apoptosis by increased pro-apoptotic signaling via the c-Abl pathway and that this causes reduced tumor growth in vivo.

  • 108.
    De Boeck, Miriam
    et al.
    Department of Molecular Cell Biology and Centre for Biomedical Genetics, Leiden University Medical Center, Postbus 9600, 2300 RC, Leiden, The Netherlands.
    ten Dijke, Peter
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Key role for ubiquitin protein modification in TGFβ signal transduction2012In: Upsala Journal of Medical Sciences, ISSN 0300-9734, E-ISSN 2000-1967, Vol. 117, no 2, p. 153-165Article, review/survey (Refereed)
    Abstract [en]

    The transforming growth factor β (TGFβ) superfamily of signal transduction molecules plays crucial roles in the regulation of cell behavior. TGFβ regulates gene transcription through Smad proteins and signals via non-Smad pathways. The TGFβ pathway is strictly regulated, and perturbations lead to tumorigenesis. Several pathway components are known to be targeted for proteasomal degradation via ubiquitination by E3 ligases. Smurfs are well known negative regulators of TGFβ, which function as E3 ligases recruited by adaptors such as I-Smads. TGFβ signaling can also be enhanced by E3 ligases, such as Arkadia, that target repressors for degradation. It is becoming clear that E3 ligases often target multiple pathways, thereby acting as mediators of signaling cross-talk. Regulation via ubiquitination involves a complex network of E3 ligases, adaptor proteins, and deubiquitinating enzymes (DUBs), the last-mentioned acting by removing ubiquitin from its targets. Interestingly, also non-degradative ubiquitin modifications are known to play important roles in TGFβ signaling. Ubiquitin modifications thus play a key role in TGFβ signal transduction, and in this review we provide an overview of known players, focusing on recent advances.

  • 109.
    de Kruijf, E M
    et al.
    Departments of Surgery, Leiden University Medical Center, Leiden, The Netherlands.
    Dekker, T J A
    Departments of Surgery, Leiden University Medical Center, Leiden, The Netherlands.
    Hawinkels, L J A C
    Departments of Molecular Cell Biology and Centre for Biomedical Genetics, Leiden University Medical Center, Leiden, The Netherlands.
    Putter, H
    Departments of Medical Statistics, Leiden University Medical Center, Leiden, The Netherlands.
    Smit, V T H B M
    Departments of Pathology, Leiden University Medical Center, Leiden, The Netherlands.
    Kroep, J R
    Departments of Medical Oncology, Leiden University Medical Center, Leiden, The Netherlands.
    Kuppen, P J K
    Departments of Surgery, Leiden University Medical Center, Leiden, The Netherlands.
    van de Velde, C J H
    Departments of Surgery, Leiden University Medical Center, Leiden, The Netherlands.
    ten Dijke, Peter
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Tollenaar, R A E M
    Departments of Surgery, Leiden University Medical Center, Leiden, The Netherlands.
    Mesker, W E
    Departments of Surgery, Leiden University Medical Center, Leiden, The Netherlands.
    The prognostic role of TGF-β signaling pathway in breast cancer patients2013In: Annals of Oncology, ISSN 0923-7534, E-ISSN 1569-8041, Vol. 24, no 2, p. 384-390Article in journal (Refereed)
    Abstract [en]

    Background

    The transforming growth factor-β (TGF-β) pathway has dual effects on tumor growth. Seemingly, discordant results have been published on the relation between TGF-β signaling markers and prognosis in breast cancer. Improved prognostic information for breast cancer patients might be obtained by assessing interactions among TGF-β signaling biomarkers.

    Patients and methods

    The expression of nuclear Smad4, nuclear phosphorylated-Smad2 (p-Smad2), and the membranous expression of TGF-β receptors I and II (TβRI and TβRII) was determined on a tissue microarray of 574 breast carcinomas. Tumors were stratified according to the Smad4 expression in combination with p-Smad2 expression or Smad4 in combination with the expression of both TGF-β receptors.

    Results

    Tumors with high expression of TβRII, TβRI and TβRII, and p-Smad2 (P = 0.018, 0.005, and 0.022, respectively), and low expression of Smad4 (P = 0.005) had an unfavorable prognosis concerning progression-free survival. Low Smad4 expression combined with high p-Smad2 expression or low expression of Smad4 combined with high expression of both TGF-β receptors displayed an increased hazard ratio of 3.04 [95% confidence interval (CI) 1.390-6.658] and 2.20 (95% CI 1.464-3.307), respectively, for disease relapse.

    Conclusions

    Combining TGF-β biomarkers provides prognostic information for patients with stage I-III breast cancer. This can identify patients at increased risk for disease recurrence that might therefore be candidates for additional treatment.

  • 110. de Miranda, Noel F. C. C.
    et al.
    van Dinther, Maarten
    van den Akker, Brendy E. W. M.
    van Wezel, Tom
    ten Dijke, Peter
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Morreau, Hans
    Transforming Growth Factor beta Signaling in Colorectal Cancer Cells With Microsatellite Instability Despite Biallelic Mutations in TGFBR22015In: Gastroenterology, ISSN 0016-5085, E-ISSN 1528-0012, Vol. 148, no 7, p. 1427-1437.e8Article in journal (Refereed)
    Abstract [en]

    BACKGROUND & AIMS: Most colorectal cancer (CRC) cells with high levels of microsatellite instability (MSI-H) accumulate mutations at a microsatellite sequence in the gene encoding transforming growth factor beta receptor II (TGFBR2). TGF beta signaling therefore is believed to be defective in these tumors, although CRC cells with TGFBR2 mutations have been reported to remain sensitive to TGF beta. We investigated how TGF beta signaling might continue in MSI-H CRC cells. METHODS: We sequenced the 10-adenines microsatellite sequence in the TGFBR2 gene of 32 MSI-H colon cancer tissues and 6 cell lines (HCT116, LS180, LS411N, RKO, SW48, and SW837). Activation of TGF beta signaling was detected by SMAD2 phosphorylation and through use of a TGF beta-responsive reporter construct in all CRC cell lines. Transcripts of TGFBR2 were knocked-down in CRC cells using short hairpin RNA. Full-length and mutant forms of TGFBR2 were expressed in LS411N cells, which do not respond to TGF beta, and their activities were measured. RESULTS: SMAD2 was phosphorylated in most MSI-H CRC tissues (strong detection in 44% and weak detection in 34% of MSI-H tumors). Phosphorylation of SMAD2 in MSI-H cells required TGFBR2-even the form encoding a frameshift mutation. Transcription and translation of TGFBR2 with a 1-nucleotide deletion at its microsatellite sequence still produced a full-length TGFBR2 protein. However, protein expression required preservation of the TGFBR2 microsatellite sequence; cells in which this sequence was replaced with a synonymous nonmicrosatellite sequence did not produce functional TGFBR2 protein. CONCLUSION: TGF beta signaling remains active in some MSI-H CRC cells despite the presence of frameshift mutations in the TGFBR2 gene because the mutated gene still expresses a functional protein. Strategies to reactivate TGF beta signaling in colorectal tumors might not be warranted, and the functional effects of mutations at other regions of microsatellite instability should be evaluated.

  • 111. Dekker, T. J. A.
    et al.
    Charehbili, A.
    Smit, V. T. H. B. M.
    ten Dijke, Peter
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Kranenbarg, E. Meershoek-Klein
    van de Velde, C. J. H.
    Nortier, J. W. R.
    Tollenaar, R. A. E. M.
    Mesker, W. E.
    Kroep, J. R. t f
    Disorganised stroma determined on pre-treatment breast cancer biopsies is associated with poor response to neoadjuvant chemotherapy: Results from the NEOZOTAC trial2015In: Molecular Oncology, ISSN 1574-7891, E-ISSN 1878-0261, Vol. 9, no 6, p. 1120-1128Article in journal (Refereed)
    Abstract [en]

    Introduction: The tumor-associated stoma is of importance for tumor progression and is generally accepted to have a significant influence on patient prognosis. However, little is known regarding specific features of tumor-associated stromal tissues and response to (neoadjuvant) chemotherapy. This study investigated the predictive value of extracellular matrix organization on response to chemotherapy in patients treated in the NEOZOTAC trial. Methods: Stromal organisation was analyzed via a simple method using image analysis software on hematoxylin and eosin (H&E)-stained slides from primary tumor biopsies collected as part of the NEOZOTAC trial. Heidenhain's AZAN trichrome-stained slides were also analyzed for comparison of collagen evaluation. Sections were stained for phospho-Smad2 (pS2) in order to determine the relationship of TGF-beta signaling with stromal organization. Results: A statistically significant relationship was observed between stroma consisting of organised collagen and pathological response to neoadjuvant chemotherapy (Odds Ratio 0.276, 95%CI 0.124-0.614, P = 0.002). This parameter was also related to ER-status (P = 0.003), clinical tumor -status (P = 0.041), nodal status (P = 0.029) and pS2 status (P = 0.025). Correlation between stromal organisation determined on H&E-stained and AZAN-stained tissue sections was high (Pearson's correlation coefficient = 0.806). Conclusion: Intratumoral stromal organisation determined using pre-treatment breast cancer biopsies was related to pathological response to chemotherapy. This parameter might play a role in the management of breast cancer for identifying those patients that are likely to benefit from neoadjuvant chemotherapy.

  • 112.
    Delis, Costas
    et al.
    Univ Thessaly, Dept Biochem & Biotechnol, Larisa 41221, Greece.;TEI Peloponnese, Kalamata, Greece..
    Krokida, Afrodite
    Univ Thessaly, Dept Biochem & Biotechnol, Larisa 41221, Greece..
    Tomatsidou, Anastasia
    Univ Thessaly, Dept Biochem & Biotechnol, Larisa 41221, Greece.;Univ Crete, Dept Biol, Iraklion, Greece..
    Tsikou, Daniela
    Univ Thessaly, Dept Biochem & Biotechnol, Larisa 41221, Greece..
    Beta, Rafailia A. A.
    Univ Thessaly, Dept Biochem & Biotechnol, Larisa 41221, Greece..
    Tsioumpekou, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Univ Thessaly, Dept Biochem & Biotechnol, Larisa 41221, Greece..
    Moustaka, Julietta
    Univ Thessaly, Dept Biochem & Biotechnol, Larisa 41221, Greece.;Aristotle Univ Thessaloniki, Dept Bot, Thessaloniki, Greece..
    Stravodimos, Georgios
    Univ Thessaly, Dept Biochem & Biotechnol, Larisa 41221, Greece..
    Leonidas, Demetres D.
    Univ Thessaly, Dept Biochem & Biotechnol, Larisa 41221, Greece..
    Balatsos, Nikolaos A. A.
    Univ Thessaly, Dept Biochem & Biotechnol, Larisa 41221, Greece..
    Papadopoulou, Kalliope K.
    Univ Thessaly, Dept Biochem & Biotechnol, Larisa 41221, Greece..
    AtHESPERIN: a novel regulator of circadian rhythms with poly(A)-degrading activity in plants2016In: RNA Biology, ISSN 1547-6286, E-ISSN 1555-8584, Vol. 13, no 1, p. 68-82Article in journal (Refereed)
    Abstract [en]

    We report the identification and characterization of a novel gene, AtHesperin (AtHESP) that codes for a deadenylase in Arabidopsis thaliana. The gene is under circadian clock-gene regulation and has similarity to the mammalian Nocturnin. AtHESP can efficiently degrade poly(A) substrates exhibiting allosteric kinetics. Size exclusion chromatography and native electrophoresis coupled with kinetic analysis support that the native enzyme is oligomeric with at least 3 binding sites. Knockdown and overexpression of AtHESP in plant lines affects the expression and rhythmicity of the clock core oscillator genes TOC1 and CCA1. This study demonstrates an evolutionary conserved poly(A)-degrading activity in plants and suggests deadenylation as a mechanism involved in the regulation of the circadian clock. A role of AtHESP in stress response in plants is also depicted.

  • 113.
    Demoulin, Jean-Baptiste
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Enarsson, Mia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Larsson, Jimmy
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Essaghir, Ahmed
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Forsberg-Nilsson, Karin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    The gene expression profile of PDGF-treated neural stem cells corresponds to partially differentiated neurons and glia2006In: Growth Factors, ISSN 0897-7194, E-ISSN 1029-2292, Vol. 24, no 3, p. 184-196Article in journal (Refereed)
    Abstract [en]

    We have previously shown that platelet-derived growth factor AA (PDGF-AA) stimulates the expansion of neuronal progenitors from neural stem cells, but is unable to replace fibroblast-growth factor 2 (FGF-2) as a stem cell mitogen. In the present study, we compared gene expression in neural stem cells that were grown in the presence of FGF-2 and in cells cultured with PDGF-AA or in the absence of growth factor, which induces differentiation. The genetic program elicited by PDGF-AA (156 significantly regulated genes) was not unique, but an intermediate between the ones of FGF-2-cultured stem cells and differentiated cells. These observations are compatible with the hypothesis that PDGF-AA induces a partial differentiation of neural stem cells, which retain the ability to proliferate, rather than acting solely as an instructing agent for neuronal differentiation. Finally, the transcriptional signature of stem cells grown with FGF-2 included a large number of genes over-expressed in gliomas and a core set of conserved genes periodically expressed during the eukaryote cell cycle.

  • 114.
    Demoulin, Jean-Baptiste
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Ericsson, Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Kallin, Anders
    Rorsman, Charlotte
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Rönnstrand, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Platelet-derived growth factor stimulates membrane lipid synthesis through activation of phosphatidylinositol 3-kinase and sterol regulatory element-binding proteins2004In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 279, no 34, p. 35392-35402Article in journal (Refereed)
    Abstract [en]

    We analyzed the transcriptional program elicited by stimulation of normal human fibroblasts with platelet-derived growth factor (PDGF) using cDNA microarrays. 103 significantly regulated transcripts that had not been previously linked to PDGF signaling were identified. Among them, a cluster of genes involved in fatty acid and cholesterol biosynthesis, including stearoyl-CoA desaturase (SCD), fatty acid synthase, and hydroxymethylglutaryl-CoA synthase (HMGCS), was up-regulated by PDGF after 24 h of treatment, and their expression correlated with increased membrane lipid production. These genes are known to be controlled by sterol regulatory element-binding proteins (SREBP). PDGF increased the amount of mature SREBP-1 and regulated the promoters of SCD and HMGCS in an SREBP-dependent manner. In line with these results, blocking SREBP processing by addition of 25-hydroxycholesterol blunted the effects of PDGF on lipogenic enzymes. SREBP activation was dependent on the phosphatidylinositol 3-kinase (PI3K) pathway, as judged from the effects of the inhibitor LY294002 and mutation of the PDGFbeta receptor tyrosines that bind the PI3K adaptor subunit p85. Fibroblast growth factors (FGF-2 and FGF-4) and other growth factors mimicked the effects of PDGF on NIH3T3 and human fibroblasts. In conclusion, our results suggest that growth factors induce membrane lipid synthesis via the activation SREBP and PI3K.

  • 115. Dib, Karim
    et al.
    Melander, Fredrik
    Axelsson, Lena
    Dagher, Marie-Claire
    Aspenström, Pontus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Andersson, Tommy
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Down-regulation of Rac activity during beta 2 integrin-mediated adhesion of human neutrophils2003In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 278, no 26, p. 24181-24188Article in journal (Refereed)
    Abstract [en]

    In human neutrophils, beta2 integrin engagement mediated a decrease in GTP-bound Rac1 and Rac2. Pretreatment of neutrophils with LY294002 or PP1 (inhibiting phosphatidylinositol 3-kinase (PI 3-kinase) and Src kinases, respectively) partly reversed the beta2 integrin-induced down-regulation of Rac activities. In contrast, beta2 integrins induced stimulation of Cdc42 that was independent of Src family members. The PI 3-kinase dependence of the beta2 integrin-mediated decrease in GTP-bound Rac could be explained by an enhanced Rac-GAP activity, since this activity was blocked by LY204002, whereas PP1 only had a minor effect. The fact that only Rac1 but not Rac2 (the dominating Rac) redistributed to the detergent-insoluble fraction and that it was independent of GTP loading excludes the possibility that down-regulation of Rac activities was due to depletion of GTP-bound Rac from the detergent-soluble fraction. The beta2 integrin-triggered relocalization of Rac1 to the cytoskeleton was enabled by a PI 3-kinase-induced dissociation of Rac1 from LyGDI. The dissociations of Rac1 and Rac2 from LyGDI also explained the PI 3-kinase-dependent translocations of Rac GTPases to the plasma membrane. However, these accumulations of Rac in the membrane, as well as that of p47phox and p67phox, were also regulated by Src tyrosine kinases. Inasmuch as Rac GTPases are part of the NADPH oxidase and the respiratory burst is elicited in neutrophils adherent by beta2 integrins, our results indicate that activation of the NADPH oxidase does not depend on the levels of Rac-GTP but instead requires a beta2 integrin-induced targeting of the Rac GTPases as well as p47phox and p67phox to the plasma membrane.

  • 116.
    Dikic, Inga
    Uppsala University, Units outside the University, Ludwig Institute for Cancer Research.
    Signal Transduction by Proline-Rich Tyrosine Kinase Pyk22002Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The proline-rich tyrosine kinase (Pyk2) together with focal adhesion kinase (FAK) define a family of non-receptor protein tyrosine kinases that are regulated by diverse stimuli. Activation of Pyk2 has been implicated in multiple signaling events, including modulation of ion channels, activation of MAP kinase cascades and apoptotic cell death. This thesis investigates the role of Pyk2 in the regulation of mitogenic signals and cell cytoskeleton.

    We identified a hematopoietic isoform of Pyk2 (designated Pyk2-H)that is generated by alternative RNA splicing and is mainly expressed in thymocytes, B cells and natural killer cells. In addition, we demonstrated that engagement of antigen receptors in lymphocytes leads to rapid tyrosine phosphorylation of Pyk2-H suggesting a potential role in host immune responses. These findings were corroborated by defects in B cell-mediated immune responses of Pyk2-/- mice.

    Several reports have previously indicated that Pyk2 acts as an upstream regulator of ERK and JNK MAP kinase cascades in response to numerous extracellular signals. Which MAP kinase pathway is activated by Pyk2 depends on arrays of effector proteins associated with Pyk2. We proposed a model where the formation of Pyk2-Src complexes results in phosphorylation of Shc, p130Cas and Pyk2. This creates binding sites for the SH2 domains of adaptor proteins Grb2 and Crk, which in turn recruit exchange factors for Ras and Rho GTPases that specifically activate ERK or JNK.

    Integration of signaling pathways initiated by receptor tyrosine kinases and integrins is essential for growth factor-mediated biological responses. We described neuronal cellular models where activation of both growth factor receptors and integrins is required for neurite outgrowth. In these cells, Pyk2 and FAK associate with integrin-linked complexes containing EGF receptors via their C- and N-terminal domains. Inhibition of Pyk2/FAK functions was sufficient to block neurite outgrowth and effectors of the C-terminal domain of Pyk2/FAK, including paxillin, were shown to regulate neurite outgrowth independently of ERK/MAP kinase in these cells. We thus proposed that Pyk2 and FAK play important roles in signal integration proximal to the integrin-growth factor receptor complexes.

  • 117.
    Drabsch, Yvette
    et al.
    Department of Molecular Cell Biology, Cancer Genomics Centre Netherlands, Leiden University Medical Center, Postbus 9600 2300, RC, Leiden, The Nethe.
    He, Shuning
    Institute of Biology, Leiden University, Einsteinweg 55, 2333, CC Leiden, The Netherlands .
    Zhang, Long
    Department of Molecular Cell Biology, Cancer Genomics Centre Netherlands, Leiden University Medical Center, Postbus 9600 2300, RC, Leiden, The Nethe.
    Snaar-Jagalska, B Ewa
    Institute of Biology, Leiden University, Einsteinweg 55, 2333, CC Leiden, The Netherlands .
    ten Dijke, Peter
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research. Department of Molecular Cell Biology, Cancer Genomics Centre Netherlands, Leiden University Medical Center, Postbus 9600 2300, RC, Leiden, The Netherlands .
    Transforming growth factor-β signalling controls human breast cancer metastasis in a zebrafish xenograft model2013In: Breast Cancer Research, ISSN 1465-5411, E-ISSN 1465-542X, Vol. 15, no 6, p. R106-Article in journal (Refereed)
    Abstract [en]

    INTRODUCTION: The transforming growth factor beta (TGF-β) signalling pathway is known to control human breast cancer invasion and metastasis. We demonstrate that the zebrafish xenograft assay is a robust and dependable animal model for examining the role of pharmacological modulators and genetic perturbation of TGF-β signalling in human breast tumour cells.

    METHODS: We injected cancer cells into the embryonic circulation (duct of cuvier) and examined their invasion and metastasis into the avascular collagenous tail. Various aspects of the TGF-β signalling pathway were blocked by chemical inhibition, small interfering RNA (siRNA), or small hairpin RNA (shRNA). Analysis was conducted using fluorescent microscopy.

    RESULTS: Breast cancer cells with different levels of malignancy, according to in vitro and in vivo mouse studies, demonstrated invasive and metastatic properties within the embryonic zebrafish model that nicely correlated with their differential tumourigenicity in mouse models. Interestingly, MCF10A M2 and M4 cells invaded into the caudal hematopoietic tissue and were visible as a cluster of cells, whereas MDA MB 231 cells invaded into the tail fin and were visible as individual cells. Pharmacological inhibition with TGF-β receptor kinase inhibitors or tumour specific Smad4 knockdown disturbed invasion and metastasis in the zebrafish xenograft model and closely mimicked the results we obtained with these cells in a mouse metastasis model. Inhibition of matrix metallo proteinases, which are induced by TGF-β in breast cancer cells, blocked invasion and metastasis of breast cancer cells.

    CONCLUSIONS: The zebrafish-embryonic breast cancer xenograft model is applicable for the mechanistic understanding, screening and development of anti-TGF-β drugs for the treatment of metastatic breast cancer in a timely and cost-effective manner.

  • 118.
    Drabsch, Yvette
    et al.
    Department of Molecular Cell Biology and Centre for Biomedical Genetics, Leiden University Medical Center, Postbus 9600, 2300 RC Leiden, The Netherlands.
    ten Dijke, Peter
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    TGF-beta Signaling in Breast Cancer Cell Invasion and Bone Metastasis2011In: Journal of mammary gland biology and neoplasia, ISSN 1083-3021, E-ISSN 1573-7039, Vol. 16, no 2, p. 97-108Article in journal (Refereed)
    Abstract [en]

    The contribution of transforming growth factor beta (TGF-beta) signaling to breast cancer has been studied for more than two decades. In an early phase TGF-beta may act as a tumour suppressor, while later, when cells have become resistant to its anti-mitogenic effects, the role of TGF-beta switches towards malignant conversion and progression. TGF-beta stimulates cell invasion and modifies the microenvironment to the advantage of cancer cells. Studies have shown that TGF-beta promotes bone and lung metastasis via different mechanisms. The therapeutic strategies to target the TGF-beta pathway in breast cancer are becoming increasingly clear. This review will focus on the role TGF-beta in breast cancer invasion and metastasis.

  • 119.
    Drabsch, Yvette
    et al.
    Department of Molecular Cell Biology and Centre for Biomedical Genetics, Leiden University Medical Center, Postbus 9600, 2300 RC Leiden, The Netherlands.
    ten Dijke, Peter
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    TGF-β signalling and its role in cancer progression and metastasis2012In: Cancer Metastasis Review, ISSN 0167-7659, E-ISSN 1573-7233, Vol. 31, no 3-4, p. 553-568Article, review/survey (Refereed)
    Abstract [en]

    The transforming growth factor-β (TGF-β) system signals via protein kinase receptors and SMAD mediators to regulate a large number of biological processes. Alterations of the TGF-β signalling pathway are implicated in human cancer. Prior to tumour initiation and early during progression, TGF-β acts as a tumour suppressor; however, at later stages, it is often a tumour promoter. Knowledge about the mechanisms involved in TGF-β signal transduction has allowed a better understanding of cancer progression, invasion, metastasis and epithelial-to-mesenchymal transition. Furthermore, several molecular targets with great potential in therapeutic interventions have been identified. This review discusses the TGF-β signalling pathway, its involvement in cancer and current therapeutic approaches.

  • 120.
    Dubrovska, Anna
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Kanamoto, Takashi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Lomnytska, Marta
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Volodko, Natalya
    Souchelnytskyi, Serhiy
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    TGFbeta1/Smad3 counteracts BRCA1-dependent repair of DNA damage2005In: Oncogene, ISSN 0950-9232, E-ISSN 1476-5594, Vol. 24, no 14, p. 2289-2297Article in journal (Refereed)
    Abstract [en]

    Inactivation of the BRCA1 gene has been found to confer susceptibility to early-onset familial breast and ovarian cancers. BRCA1 regulates DNA repair, chromatin remodeling and affects gene transcription. Transforming growth factor-beta (TGFbeta) is a potent regulator of growth, apoptosis and invasiveness of tumor cells, including breast cancer cells. Here we show that Smad3 which is a component of the TGFbeta signaling pathway, forms a complex with BRCA1 in vitro and in vivo. The interaction is mediated by the MH1 domain of Smad3 and the C-terminal part of BRCA1. We observed a co-localization of Smad3 and BRCA1 in nuclear complexes. We also found that TGFbeta1/Smad3 counteracted BRCA1-dependent repair of DNA double-strand breaks in human breast epithelial cells, as evaluated by BRCA1 nuclear foci formation, single-cell gel electrophoresis and cell survival assays. Thus, TGFbeta1/Smad3 suppresses BRCA1-dependent DNA repair in response to a DNA damaging agent.

  • 121.
    Dubrovska, Anna
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Souchelnytskyi, Serhiy
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Efficient enrichment of intact phosphorylated proteins by modified immobilized metal-affinity chromatography2005In: Proteomics, ISSN 1615-9853, E-ISSN 1615-9861, Vol. 5, no 18, p. 4678-4683Article in journal (Refereed)
    Abstract [en]

    Phosphoproteome studies are hampered by the lack of methods which allow a comprehensive and fast analysis of intact phosphoproteins. Here we describe an immobilized metal-affinity chromatography (IMAC)-based technique for the enrichment of phosphorylated proteins, which allows recovery of up to 90% of phosphoproteins. This technique is compatible with 2-DE and can be applied to cultured cells and tissues.

  • 122.
    Edlund, Sofia
    Uppsala University, Units outside the University, Ludwig Institute for Cancer Research.
    Mechanisms for TGF-β-Mediated Regulation of the Actin Filament System and Apoptosis2003Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Transforming growth factor-β (TGF-β) is a member of a large superfamily of cytokines which participate in many different types of cellular processes, such as growth inhibition, cell migration, differentiation, cell adhesion, wound healing and immunosuppression. Alterations of TGF-β superfamily signalling results in several different disorders, including bone disease, vascular disease and cancer. The TGF-β signalling pathways involve several different proteins, such as the Smad proteins, which upon receptor activation are translocated to the nucleus, where they affect transcriptional responses.

    The actin cytoskeleton is an organised network of filaments with a highly dynamic structure, which is under a continuous reconstruction to control the morphology, survival, growth and motility of eukaryotic cells. The members of the family of small GTP-binding proteins have been shown to be important regulators of the actin cytoskeleton.

    TGF-β was found to induce short term as well as long term actin reorganisation in prostate cancer cells. The short term response included membrane ruffling, and required signalling by the small GTPases Cdc42 and Rho as well as, the involvement of the mitogen-activated protein kinases p38 (p38 MAPK). The long term response included formation of stress fibers and required a cooperation between Smad and Rho GTPase signalling pathways involving the Rho-associated coiled-coil-containing protein kinase 1 (ROCK1).

    The TGF-β-induced activation of Cdc42 was, furthermore, shown to require the inhibitory Smad7 and p38 MAP kinase, via a PI3K-dependent pathway. Mixed lineage kinase 3 (MLK3), a mediator downstream of Cdc42, was necessary for the Cdc42-dependent actin filament reorganisation.

    Apoptosis is an important and carefully regulated process in human development and disease, which allows the multicellular organisms to remove cells that are in excess or potentially dangerous. TGF-β family members can induce apoptosis in many different cell types, in the presence or absence of other growth factors. Smad7 had previously been shown to be necessary for TGF-β-induced apoptosis of epithelial cells. We could show that Smad7 is required for TGF-β-induced activation of the TGF-β activated kinase 1 (TAK1)-mitogen-activated protein kinase kinase 3 (MKK3)-p38 MAPK pathway, which subsequently leads to apoptosis in prostate cancer cells.

    Members of the lymphoid enhancer factor-1/T-cell factor (LEF1/TCF) family of transcription factors have, together with β-catenin, been shown to be nuclear effectors in the Wnt-signalling pathway. We investigated a possible cross-talk between the TGF-β and Wnt signalling pathways. We found that TGF-β, in a Smad7-dependent manner induced a nuclear accumulation of β-catenin and enhanced the transcriptional activity of β-catenin and the induction of the downstream target gene c-myc. Since β-catenin and c-Myc has been shown to promote apoptosis, our results suggests the possibility that β-catenin contributes to TGF-β-induced apoptosis

    List of papers
    1. Transforming growth factor-beta-induced mobilization of actin cytoskeleton required signaling by small GTPases Cdc42 and RhoA
    Open this publication in new window or tab >>Transforming growth factor-beta-induced mobilization of actin cytoskeleton required signaling by small GTPases Cdc42 and RhoA
    2002 (English)In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 13, no 3, p. 902-914Article in journal (Refereed) Published
    Abstract [en]

    Transforming growth factor-beta (TGF-beta) is a potent regulator of cell growth and differentiation in many cell types. The Smad signaling pathway constitutes a main signal transduction route downstream of TGF-beta receptors. We studied TGF-beta-induced rearrangements of the actin filament system and found that TGF-beta 1 treatment of PC-3U human prostate carcinoma cells resulted in a rapid formation of lamellipodia. Interestingly, this response was shown to be independent of the Smad signaling pathway; instead, it required the activity of the Rho GTPases Cdc42 and RhoA, because ectopic expression of dominant negative mutant Cdc42 and RhoA abrogated the response. Long-term stimulation with TGF-beta 1 resulted in an assembly of stress fibers; this response required both signaling via Cdc42 and RhoA, and Smad proteins. A known downstream effector of Cdc42 is p38(MAPK); treatment of the cells with the p38(MAPK) inhibitor 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(pyridyl)1H-imidazole (SB203580), as well as ectopic expression of a kinase-inactive p38(MAPK), abrogated the TGF-beta-induced actin reorganization. Moreover, treatment of cells with the inhibitors of the RhoA target-protein Rho-associated coiled-coil kinase (+)-R-trans-4-(aminoethyl)-N-(4-pyridyl) cyclohexanecarboxamide (Y-27632) and 1-5(-isoquinolinesulfonyl)homopiperazine (HA-1077), as well as ectopic expression of kinase-inactive Rho coiled-coil kinase-1, abrogated the TGF-beta 1-induced formation of stress fibers. Collectively, these data indicate that TGF-beta-induced membrane ruffles occur via Rho GTPase-dependent pathways, whereas long-term effects require cooperation between Smad and Rho GTPase signaling pathways.

    Place, publisher, year, edition, pages
    The American Society for Cell Biology, 2002
    Keywords
    Actins/*metabolism, Amides/pharmacology, Animals, Cell Surface Extensions/metabolism, Cytoskeleton/drug effects/*metabolism, DNA-Binding Proteins/genetics/metabolism, Enzyme Inhibitors/pharmacology, Humans, Imidazoles/pharmacology, Intracellular Signaling Peptides and Proteins, Mitogen-Activated Protein Kinases/antagonists & inhibitors/genetics/metabolism, Protein-Serine-Threonine Kinases/antagonists & inhibitors/genetics/metabolism, Pyridines/pharmacology, Rats, Recombinant Fusion Proteins/metabolism, Signal Transduction/*physiology, Smad4 Protein, Stress Fibers/metabolism, Trans-Activators/genetics/metabolism, Transforming Growth Factor beta/*metabolism, Tumor Cells; Cultured, cdc42 GTP-Binding Protein/*metabolism, p38 Mitogen-Activated Protein Kinases, rac1 GTP-Binding Protein/metabolism, rhoA GTP-Binding Protein/*metabolism
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-90259 (URN)10.1091/mbc.01–08–0398 (DOI)11907271 (PubMedID)
    Available from: 2003-04-03 Created: 2003-04-03 Last updated: 2017-12-14Bibliographically approved
    2.
    The record could not be found. The reason may be that the record is no longer available or you may have typed in a wrong id in the address field.
    3. Transforming growth factor-beta1-induced apoptosis of prostate cancer cells involves Smad7-dependent activation of p38 by TGF-beta-activated kinase 1 and mitogen-activated protein kinase kinase 3
    Open this publication in new window or tab >>Transforming growth factor-beta1-induced apoptosis of prostate cancer cells involves Smad7-dependent activation of p38 by TGF-beta-activated kinase 1 and mitogen-activated protein kinase kinase 3
    Show others...
    2003 (English)In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 14, no 2, p. 529-544Article in journal (Refereed) Published
    Abstract [en]

    The inhibitory Smad7, a direct target gene for transforming growth factor-beta (TGF-beta), mediates TGF-beta1-induced apoptosis in several cell types. Herein, we report that apoptosis of human prostate cancer PC-3U cells induced by TGF-beta1 or Smad7 overexpression is caused by a specific activation of the p38 mitogen-activated protein kinase pathway in a TGF-beta-activated kinase 1 (TAK1)- and mitogen-activated protein kinase kinase 3 (MKK3)-dependent manner. Expression of dominant negative p38, dominant negative MKK3, or incubation with the p38 selective inhibitor [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole], prevented TGF-beta1-induced apoptosis. The expression of Smad7 was required for TGF-beta-induced activation of MKK3 and p38 kinases, and endogenous Smad7 was found to interact with phosphorylated p38 in a ligand-dependent manner. Ectopic expression of wild-type TAK1 promoted TGF-beta1-induced phosphorylation of p38 and apoptosis, whereas dominant negative TAK1 reduced TGF-beta1-induced phosphorylation of p38 and apoptosis. Endogenous Smad7 was found to interact with TAK1, and TAK1, MKK3, and p38 were coimmunoprecipitated with Smad7 in transiently transfected COS1 cells. Moreover, ectopically expressed Smad7 enhanced the coimmunoprecipitation of HA-MKK3 and Flag-p38, supporting the notion that Smad7 may act as a scaffolding protein and facilitate TAK1- and MKK3-mediated activation of p38.

    Place, publisher, year, edition, pages
    The American Society for Cell Biology, 2003
    Keywords
    Animals, Apoptosis, Blotting; Western, COS Cells, DNA Fragmentation, DNA-Binding Proteins/*metabolism, Enzyme Activation, Enzyme Inhibitors/pharmacology, Genes; Dominant, Humans, In Situ Nick-End Labeling, MAP Kinase Kinase 3, MAP Kinase Kinase Kinases/*metabolism, Male, Microscopy; Fluorescence, Mitogen-Activated Protein Kinase Kinases/*metabolism, Mitogen-Activated Protein Kinases/*metabolism, Phosphorylation, Precipitin Tests, Prostatic Neoplasms/*pathology, Protein Binding, Protein-Tyrosine Kinase/*metabolism, Research Support; Non-U.S. Gov't, Time Factors, Trans-Activators/*metabolism, Transfection, Transforming Growth Factor beta/metabolism/*physiology, Tumor Cells; Cultured, p38 Mitogen-Activated Protein Kinases
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-90261 (URN)10.1091/mbc.02–03–0037 (DOI)12589052 (PubMedID)
    Available from: 2003-04-03 Created: 2003-04-03 Last updated: 2017-12-14Bibliographically approved
    4.
    The record could not be found. The reason may be that the record is no longer available or you may have typed in a wrong id in the address field.
  • 123.
    Edlund, Sofia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Bu, Shizhong
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Schuster, Norbert
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Aspenström, Pontus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Heuchel, Rainer
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Heldin, Nils-Erik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
    ten Dijke, Peter
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Landström, Maréne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Transforming growth factor-beta1-induced apoptosis of prostate cancer cells involves Smad7-dependent activation of p38 by TGF-beta-activated kinase 1 and mitogen-activated protein kinase kinase 32003In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 14, no 2, p. 529-544Article in journal (Refereed)
    Abstract [en]

    The inhibitory Smad7, a direct target gene for transforming growth factor-beta (TGF-beta), mediates TGF-beta1-induced apoptosis in several cell types. Herein, we report that apoptosis of human prostate cancer PC-3U cells induced by TGF-beta1 or Smad7 overexpression is caused by a specific activation of the p38 mitogen-activated protein kinase pathway in a TGF-beta-activated kinase 1 (TAK1)- and mitogen-activated protein kinase kinase 3 (MKK3)-dependent manner. Expression of dominant negative p38, dominant negative MKK3, or incubation with the p38 selective inhibitor [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole], prevented TGF-beta1-induced apoptosis. The expression of Smad7 was required for TGF-beta-induced activation of MKK3 and p38 kinases, and endogenous Smad7 was found to interact with phosphorylated p38 in a ligand-dependent manner. Ectopic expression of wild-type TAK1 promoted TGF-beta1-induced phosphorylation of p38 and apoptosis, whereas dominant negative TAK1 reduced TGF-beta1-induced phosphorylation of p38 and apoptosis. Endogenous Smad7 was found to interact with TAK1, and TAK1, MKK3, and p38 were coimmunoprecipitated with Smad7 in transiently transfected COS1 cells. Moreover, ectopically expressed Smad7 enhanced the coimmunoprecipitation of HA-MKK3 and Flag-p38, supporting the notion that Smad7 may act as a scaffolding protein and facilitate TAK1- and MKK3-mediated activation of p38.

  • 124.
    Edlund, Sofia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Landström, Maréne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Aspenström, Pontus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Smad7 is required for TGF-ß-induced activation of the small GTPase Cdc422004In: Journal of Cell Science, ISSN 0021-9533, E-ISSN 1477-9137, Vol. 117, no Pt 9, p. 1835-1847Article in journal (Refereed)
    Abstract [en]

    Transforming growth factor beta (TGF-beta) is a potent regulator of cell growth and differentiation in many cell types. The Smad signaling pathway constitutes a main signal transduction route downstream of TGF-beta receptors. The inhibitory Smads, Smad6 and Smad7, are considered to function as negative regulators of the TGF-beta/Smad signaling cascade. In a previous study, we found that TGF-beta induces rearrangements of the actin filament system in human prostate carcinoma cells and that this response requires the small GTPases Cdc42 and RhoA. On the basis of the current view on the function of Smad7 in TGF-beta signaling, we hypothesized that Smad7 would function as a negative regulator of the TGF-beta-induced activation of Cdc42 and RhoA, but instead we found that the reverse is the case; Smad7 is required for the TGF-beta-induced activation of Cdc42 and the concomitant reorganization of the actin filament system. These observations propose a novel role for Smad7 in TGF-beta-dependent activation of Rho GTPases.

  • 125.
    Edlund, Sofia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Landström, Maréne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Aspenström, Pontus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Transforming growth factor-beta-induced mobilization of actin cytoskeleton required signaling by small GTPases Cdc42 and RhoA2002In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 13, no 3, p. 902-914Article in journal (Refereed)
    Abstract [en]

    Transforming growth factor-beta (TGF-beta) is a potent regulator of cell growth and differentiation in many cell types. The Smad signaling pathway constitutes a main signal transduction route downstream of TGF-beta receptors. We studied TGF-beta-induced rearrangements of the actin filament system and found that TGF-beta 1 treatment of PC-3U human prostate carcinoma cells resulted in a rapid formation of lamellipodia. Interestingly, this response was shown to be independent of the Smad signaling pathway; instead, it required the activity of the Rho GTPases Cdc42 and RhoA, because ectopic expression of dominant negative mutant Cdc42 and RhoA abrogated the response. Long-term stimulation with TGF-beta 1 resulted in an assembly of stress fibers; this response required both signaling via Cdc42 and RhoA, and Smad proteins. A known downstream effector of Cdc42 is p38(MAPK); treatment of the cells with the p38(MAPK) inhibitor 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(pyridyl)1H-imidazole (SB203580), as well as ectopic expression of a kinase-inactive p38(MAPK), abrogated the TGF-beta-induced actin reorganization. Moreover, treatment of cells with the inhibitors of the RhoA target-protein Rho-associated coiled-coil kinase (+)-R-trans-4-(aminoethyl)-N-(4-pyridyl) cyclohexanecarboxamide (Y-27632) and 1-5(-isoquinolinesulfonyl)homopiperazine (HA-1077), as well as ectopic expression of kinase-inactive Rho coiled-coil kinase-1, abrogated the TGF-beta 1-induced formation of stress fibers. Collectively, these data indicate that TGF-beta-induced membrane ruffles occur via Rho GTPase-dependent pathways, whereas long-term effects require cooperation between Smad and Rho GTPase signaling pathways.

  • 126.
    Edlund, Sofia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Lee, So Young
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Grimsby, Susanne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Zhang, Shouthing
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Aspenström, Pontus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Landström, Maréne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Interaction between Smad7 and beta-catenin: importance for transforming growth factor beta-induced apoptosis2005In: Molecular and Cellular Biology, ISSN 0270-7306, E-ISSN 1098-5549, Vol. 25, no 4, p. 1475-1488Article in journal (Refereed)
    Abstract [en]

    Members of the transforming growth factor beta (TGF-beta) and Wnt/wingless superfamilies regulate cell fate during development and tissue maintenance. Here we report that Smad7 interacts with beta-catenin and lymphoid enhancer binding factor 1/T-cell-specific factor (LEF1/TCF), transcriptional regulators in Wnt signaling, in a TGF-beta-dependent manner. Smad7 was found to be required for TGF-beta1-induced accumulation of beta-catenin and LEF1 in human prostate cancer (PC-3U) cells as well as in human keratinocytes (HaCaT cells). Moreover, when the endogenous Smad7 was repressed by specific small interfering RNA, TGF-beta-induced increase of activated p38, Akt phosphorylated on Ser473, glycogen synthase kinase 3beta phosphorylated on Ser9 was prevented, as well as the TGF-beta-induced association between beta-catenin and LEF1. Notably, the observed physical association of Smad7 and beta-catenin was found to be important for TGF-beta-induced apoptosis, since suppression of beta-catenin expression by small interfering RNA decreased the apoptotic response to TGF-beta.

  • 127.
    Eger, Glenda
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Regulation and Function of MAP Kinases in PDGF Signaling2016Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Platelet-derived growth factor (PDGF) is a family of signaling molecules that stimulates cell growth, survival and migration. PDGF is recognized by specific transmembrane proteins, the PDGF receptors, which relay the signals to the cell activating the Mitogen-activated protein (MAP) kinases and other signaling pathways. Aberrant activation of these pathways is frequently detected in cancer. Hence, the study of these processes is essential for identifying potential drug targets or diagnostic markers.

    In paper I, we identified Receptor Subfamily 4 Group A Member 1 NR4A1 to be regulated by PDGF via MAP kinases, clarifying the role of Extracellular signal–regulated kinases (Erk) 1/2, Erk5 and Nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) in its regulation. NR4A1 was found to be important for the tumorigenic potential, measured as anchorage-independent growth, of glioblastoma cells.

    Since the cellular responses elicited by PDGF result from the balance between phosphorylation and dephosphorylation events, we investigated the role of the dual specificity phosphatases DUSP4/MKP-2 and DUSP6/MKP-3. In paper II, we describe the crucial role of Erk1/2 and p53 in the expression of DUSP4/MKP2. Moreover, we observed that DUSP4/MKP-2 downregulation decreases Erk5 activation and accelerates PDGFRβ internalization and downregulation resulting in a specific inhibition of Signal transducers and activators of transcription (Stat) 3, Src and protein kinase C (PKC), and partially of p38, Stat1/5 and Phoshoplipase Cγ (PLCγ).

    In paper III, we report that DUSP6/MKP-3 creates a negative cross-talk between Erk1/2 and Erk5 and an auto-inhibitory feedback loop on the PI3-kinase/Akt pathway. In paper IV, we identify a new regulative mechanism of the PDGF pathway. PDGF induces Erk5 expression and activation that modulates the PDGFRβ activity. After Erk5 downregulation, the receptor undergoes to a faster and stronger activation that results in a faster internalization and degradation.

    In conclusion, we present a mechanism through which the PDGF/MAP kinases support tumor growth, and elucidate different regulatory pathways involved in PDGF signaling.

    List of papers
    1. NR4A1 Promotes PDGF-BB-Induced Cell Colony Formation in Soft Agar
    Open this publication in new window or tab >>NR4A1 Promotes PDGF-BB-Induced Cell Colony Formation in Soft Agar
    2014 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 9, no 9, p. e109047-Article in journal (Refereed) Published
    Abstract [en]

    The fibroblast mitogen platelet-derived growth factor -BB (PDGF-BB) induces a transient expression of the orphan nuclear receptor NR4A1 (also named Nur77, TR3 or NGFIB). The aim of the present study was to investigate the pathways through which NR4A1 is induced by PDGF-BB and its functional role. We demonstrate that in PDGF-BB stimulated NIH3T3 cells, the MEK1/2 inhibitor CI-1040 strongly represses NR4A1 expression, whereas Erk5 downregulation delays the expression, but does not block it. Moreover, we report that treatment with the NF-κB inhibitor BAY11-7082 suppresses NR4A1 mRNA and protein expression. The majority of NR4A1 in NIH3T3 was found to be localized in the cytoplasm and only a fraction was translocated to the nucleus after continued PDGF-BB treatment. Silencing NR4A1 slightly increased the proliferation rate of NIH3T3 cells; however, it did not affect the chemotactic or survival abilities conferred by PDGF-BB. Moreover, overexpression of NR4A1 promoted anchorage-independent growth of NIH3T3 cells and the glioblastoma cell lines U-105MG and U-251MG. Thus, whereas NR4A1, induced by PDGF-BB, suppresses cell growth on a solid surface, it increases anchorage-independent growth.

    National Category
    Cancer and Oncology
    Identifiers
    urn:nbn:se:uu:diva-233387 (URN)10.1371/journal.pone.0109047 (DOI)000343671700217 ()25269081 (PubMedID)
    Funder
    Swedish Research Council, K2011-67X-21859-01-6Swedish Cancer Society, 130519
    Available from: 2014-10-02 Created: 2014-10-02 Last updated: 2017-12-05Bibliographically approved
    2. Depletion of DUSP4 results in enhanced PDGFRβ cell surface clearance and suppression of PDGF-BB-induced activation of Erk5, Stat3, Src and PKC
    Open this publication in new window or tab >>Depletion of DUSP4 results in enhanced PDGFRβ cell surface clearance and suppression of PDGF-BB-induced activation of Erk5, Stat3, Src and PKC
    (English)Manuscript (preprint) (Other academic)
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-301761 (URN)
    Available from: 2016-08-24 Created: 2016-08-24 Last updated: 2016-08-25
    3. MKP3 negatively modulates PDGF-induced Akt and Erk5 phosphorylation as well as chemotaxis
    Open this publication in new window or tab >>MKP3 negatively modulates PDGF-induced Akt and Erk5 phosphorylation as well as chemotaxis
    Show others...
    2012 (English)In: Cellular Signalling, ISSN 0898-6568, E-ISSN 1873-3913, Vol. 24, no 3, p. 635-640Article in journal (Refereed) Published
    Abstract [en]

    MAP kinase phosphatase-3 (MKP3), also known as DUSP6 or Pyst1, is a dual specificity phosphatase considered to selectively dephosphorylate extracellular-signal-regulated kinase 1/2 (Erk1/2). Here, we report that in NIH3T3 cells, MKP3 is induced in response to platelet-derived growth factor (PDGF)-BB treatment in an Erk1/2- and phosphatidylinositol 3-kinase (PI3K)-dependent manner, but independently of Erk5 expression. Silencing of MKP3 expression did not affect PDGF-BB-induced Erk1/2 or p38 phosphorylation; however, their basal level of phosphorylation was elevated. Furthermore, we found that PDGF-BB-mediated activation of Erk5 and Akt was enhanced when the MKP3 expression was reduced. Interfering with Mek1/2 or PI3K using the inhibitors CI-1040 and LY-294002, respectively, inhibited PDGF-BB-induced MKP3 expression. Functionally, we found that MKP3 silencing did not affect cell proliferation, but enhanced the chemotactic response toward PDGF-BB. Although both Akt and Erk5 have been linked to increased cell survival, downregulation of MKP3 did not alter the ability of PDGF-BB to protect NIH3T3 cells from starvation-induced apoptosis. However, we observed an increased apoptosis in untreated cells with reduced MKP3 expression. In summary, our data indicate that there is negative cross-talk between Erk1/2 and Erk5 that involves regulation of MKP3 expression, and that PI3K in addition to promoting Akt phosphorylation also negatively modulates Akt, through MKP3 expression.

    Keywords
    Non-Smads, Smads, TAK1, TGF beta, TRAF6
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-168808 (URN)10.1016/j.cellsig.2011.11.001 (DOI)000300478400007 ()22100392 (PubMedID)
    Available from: 2012-02-15 Created: 2012-02-15 Last updated: 2017-12-07Bibliographically approved
    4. Erk5 promotes prolonged PDGFRβ activation by limiting ligand-induced receptor internalization and degradation
    Open this publication in new window or tab >>Erk5 promotes prolonged PDGFRβ activation by limiting ligand-induced receptor internalization and degradation
    (English)Manuscript (preprint) (Other academic)
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-301762 (URN)
    Available from: 2016-08-24 Created: 2016-08-24 Last updated: 2016-08-25
  • 128.
    Eger, Glenda
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Papadopoulos, Natalia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Lennartsson, Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    NR4A1 Promotes PDGF-BB-Induced Cell Colony Formation in Soft Agar2014In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 9, no 9, p. e109047-Article in journal (Refereed)
    Abstract [en]

    The fibroblast mitogen platelet-derived growth factor -BB (PDGF-BB) induces a transient expression of the orphan nuclear receptor NR4A1 (also named Nur77, TR3 or NGFIB). The aim of the present study was to investigate the pathways through which NR4A1 is induced by PDGF-BB and its functional role. We demonstrate that in PDGF-BB stimulated NIH3T3 cells, the MEK1/2 inhibitor CI-1040 strongly represses NR4A1 expression, whereas Erk5 downregulation delays the expression, but does not block it. Moreover, we report that treatment with the NF-κB inhibitor BAY11-7082 suppresses NR4A1 mRNA and protein expression. The majority of NR4A1 in NIH3T3 was found to be localized in the cytoplasm and only a fraction was translocated to the nucleus after continued PDGF-BB treatment. Silencing NR4A1 slightly increased the proliferation rate of NIH3T3 cells; however, it did not affect the chemotactic or survival abilities conferred by PDGF-BB. Moreover, overexpression of NR4A1 promoted anchorage-independent growth of NIH3T3 cells and the glioblastoma cell lines U-105MG and U-251MG. Thus, whereas NR4A1, induced by PDGF-BB, suppresses cell growth on a solid surface, it increases anchorage-independent growth.

  • 129.
    Eger, Glenda
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Rorsman, Charlotte
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Lennartsson, Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Depletion of DUSP4 results in enhanced PDGFRβ cell surface clearance and suppression of PDGF-BB-induced activation of Erk5, Stat3, Src and PKCManuscript (preprint) (Other academic)
  • 130. Eichner, Annegret
    et al.
    Brock, Josef
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Souchelnytskyi, Serhiy
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Bone morphogenetic protein-7 (OP1) and transforming growth factor-beta1 modulate 1,25(OH)2-vitamin D3-induced differentiation of human osteoblasts2002In: Experimental Cell Research, ISSN 0014-4827, E-ISSN 1090-2422, Vol. 275, no 1, p. 132-142Article in journal (Refereed)
    Abstract [en]

    Bone morphogenetic proteins (BMPs) and transforming growth factor-beta (TGFbeta) are potent regulators of osteoblast differentiation and proliferation, processes that are crucial in bone remodeling. BMPs and TGFbeta act in concert with other local factors and hormones, among them 1,25(OH)2-vitamin D3 and insulin. Here we show that BMP7 inhibits 1,25(OH)2-vitamin D3-induced differentiation of human osteoblasts, whereas TGFbeta1 stimulates it, as assessed by assays for alkaline phosphatase (ALP) induction, matrix mineralization, and morphology changes. BMP7 or TGFbeta1 alone affects the differentiation of human osteoblasts. Similar results were obtained in assays for ALP induction using conditionally immortalized human osteoblasts (hFOB) and primary osteoblasts obtained from trabecular bone of the femoral head after hip replacement surgery. BMP7 stimulation led to a decrease of 1,25(OH)2-vitamin D3-induced binding of nuclear proteins to a vitamin D response element, as shown by electrophoretic mobility shift assay. Our results suggest that 1,25(OH)2-vitamin D3 modulates in opposite ways the effects of BMP7 and TGFbeta1 on osteoblast differentiation.

  • 131.
    Ekerljung, Lina
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Lennartsson, Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Gedda, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    The HER2-binding Affibody Molecule (ZHER2:342)2 Increases Radiosensitivity in SKBR-3 cells2012In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 7, no 11, p. e49579-Article in journal (Refereed)
    Abstract [en]

    We have previously shown that the HER2-specific affibody molecule (ZHER2:342)2 inhibits proliferation of SKBR-3 cells. Here, we continue to investigate its biological effects in vitro by studying receptor dimerization and clonogenic survival following irradiation. We found that (ZHER2:342)2 sensitizes the HER2-overexpressing cell line SKBR-3 to ionizing radiation. The survival after exposure to (ZHER2:342)2 and 8 Gy (S8Gy 0.006) was decreased by a factor of 4 compared to the untreated (S8Gy 0.023). The low HER2-expressing cell line MCF-7 was more radiosensitive than SKBR-3 but did not respond to (ZHER2:342)2. Treatment by (ZHER2:342)2 strongly increased the levels of dimerized and phosphorylated HER2 already after 5 minutes of stimulation. The monomeric ZHER2:342 does not seem to be able to induce receptor phosphorylation and dimerization or sensitize cells to irradiation.