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HD-PTP is important for platelet-derived growth factor beta-receptor downregulation and mitogenicity
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
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(English)Manuscript (preprint) (Other (popular science, discussion, etc.))
Abstract [en]

HD-PTP (His domain-protein tyrosine phosphatase) is a ubiquitously expressed, putative protein tyrosine phosphatase that has been implicated in regulation of cell migration and endosomal sorting of epidermal growth factor receptor (EGFR), and in multivesicular bodies formation. In this study we show that silencing of HD-PTP negatively regulates the PDGF- induced phosphorylation of the ubiquitin ligase c-Cbl, which was associated with lower PDGFRβ ubiquitination. We also showed that in cells depleted of HD-PTP, PDGFRβ degradation but not cell-surface clearance was decreased. Interestingly, our data showed that lack HD-PTP in cells negatively affected PDGF- induced phosphorylation of Erk1/2, Akt and p38 pathways. However, reduced HD-PTP expression resulted in increased proliferative response of cell treated with PDGF. Our data suggests that HD-PTP functions downstream of the PDGFR, affecting c-Cbl phosphorylation, PDGFRβ degradation and signaling.

Keyword [en]
PDGF, HD-PTP, ubiquitination, downregulation, degradation, c-Cbl, proliferation
Identifiers
URN: urn:nbn:se:uu:diva-123093OAI: oai:DiVA.org:uu-123093DiVA: diva2:312021
Available from: 2010-04-23 Created: 2010-04-23 Last updated: 2010-05-12
In thesis
1. Regulation of PDGFRβ signaling 
Open this publication in new window or tab >>Regulation of PDGFRβ signaling 
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Platelet-derived growth factor (PDGF) isoforms, which bind to closely related a- and b-tyrosine kinase receptors, induce migration, proliferation, survival and differentiation of mesenchymal cells. They signal by the active receptor attracting Src homology 2 (SH2) domain containing proteins, which subsequently initiate a set of signaling pathways. The aim of this thesis was to elucidate regulatory mechanisms involved in PDGFRb signaling.

In the first two projects we investigated the roles in downregulation of PDGFRb of two related adaptor proteins, i.e. ALG-2 interacting protein X (Alix) and His-domain containing protein tyrosine phosphatase (HD-PTP) functions of. We found that Alix and HD-PTP influence ubiquitination of PDGFRb following PDGF stimulation, by affecting the E3 ligase c-Cbl. Alix enhances complex formation between c-Cbl and PDGFRb, increases c-Cbl phosphorylation and decreases its stability. Interestingly, while both HD-PTP and Alix participate in degradation of PDGFRb, only Alix affects receptor internalization. Moreover, we demonstrated that absence of HD-PTP promotes cell proliferation. In conclusion, we suggest that both Alix and HD-PTP are important adaptor proteins in regulation of PDGFRb downregulation, although the observed differences between their actions suggest that Alix and HD-PTP exert their functions via different mechanisms.

The third study explored the importance of tyrosine residue 857 in the activation loop of PDGFRb. We report that, in vitro the tyrosine residue 857 to phenylalanine (Y857F) mutant receptor kinase activity is diminished while in vivo it does not affect the phosphorylation of PDGFRb. The phosphorylation pattern of PDGFRb revealed that most sites in the Y857F mutant receptor were phosphorylated similarly as in the wild-type receptor. However, tyrosine residue 771 was found to be hyperphosphorylated in the Y857F mutant receptor. This may be due to defective phosphorylation and activation of SHP-2, since it has been shown to dephosphorylate the receptor at Y771. In addition, activation of the Erk1/2 and Akt pathways was defective downstream of the Y857F mutant receptor. Interestingly, the Y857F mutant receptor was able to mediate cell migration, but not proliferation.

The last study investigated a role of the tyrosine kinase Fer in PDGF signaling. We showed that Fer interacted with and was activated by PDGFRb in a ligand-dependent manner. In cells depleted of Fer, receptor phosphorylation was decreased and phosphorylation of Stat3 was abolished, whereas Stat5, Erk1/2 and Akt were activated normally. Colony formation in soft agar was abolished in cells depleted of Fer, but no effect was seen on cell proliferation and migration. Since Stat3 has been shown to be involved in transformation, we speculate that phosphorylation of Stat3 in Fer-depleted cells, affects the ability of cells to form colonies.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2010. 59 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 563
Keyword
PDGF, HD-PTP, Alix, Cbl, Ub, Fer, Y857, Y771, downregulation, degradation, ubiquitination, activation loop
National Category
Biochemistry and Molecular Biology
Research subject
Medical Cell Biology
Identifiers
urn:nbn:se:uu:diva-123045 (URN)978-91-554-7813-1 (ISBN)
Public defence
2010-06-02, B42, BMC, BMC, 09:00 (English)
Opponent
Supervisors
Available from: 2010-05-11 Created: 2010-04-22 Last updated: 2010-05-24

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