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Ligand stimulation reduces platelet-derived growth factor beta-receptor susceptibility to tyrosine dephosphorylation
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.
2001 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 276, no 30, 27749-27752 p.Article in journal (Refereed) Published
Abstract [en]

Ligand binding to the platelet-derived growth factor (PDGF) beta-receptor leads to increased receptor tyrosine phosphorylation as a consequence of dimerization-induced activation of the intrinsic receptor tyrosine kinase activity. In this study we asked whether ligand-stimulated PDGF beta-receptor tyrosine phosphorylation, to some extent, also involved reduced susceptibility to tyrosine dephosphorylation. To investigate this possibility we compared the sensitivity of ligand-stimulated and non-stimulated forms of tyrosine-phosphorylated PDGF beta-receptors to dephosphorylation using various preparations containing protein-tyrosine phosphatase activity. Ligand-stimulated or unstimulated tyrosine-phosphorylated receptors were obtained after incubation of cells with pervanadate only or pervanadate, together with PDGF-BB, respectively. Dephosphorylation of receptors immobilized on wheat germ agglutinin-Sepharose, as well as of receptors in intact cell membranes, was investigated under conditions when rephosphorylation did not occur. As compared with unstimulated receptors the ligand-stimulated PDGF beta-receptors showed about 10-fold reduced sensitivity to dephosphorylation by cell membranes, a recombinant form of the catalytic domain of density-enhanced phosphatase-1, or recombinant protein-tyrosine phosphatase 1B. We conclude that ligand-stimulated forms of the PDGF beta-receptor display a reduced susceptibility to dephosphorylation. Our findings suggest a novel mechanism whereby ligand stimulation of PDGF beta-receptor, and possibly other tyrosine kinase receptors, leads to a net increase in receptor tyrosine phosphorylation.

Place, publisher, year, edition, pages
The American Society for Biochemistry and Molecular Biology, Inc. , 2001. Vol. 276, no 30, 27749-27752 p.
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-90184DOI: 10.1074/jbc.C100286200PubMedID: 11390370OAI: oai:DiVA.org:uu-90184DiVA: diva2:162449
Available from: 2003-03-14 Created: 2003-03-14 Last updated: 2017-12-14Bibliographically approved
In thesis
1. Protein Tyrosine Phosphatases as Regulators of Receptor Ryrosine Kinases
Open this publication in new window or tab >>Protein Tyrosine Phosphatases as Regulators of Receptor Ryrosine Kinases
2003 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Tyrosine phosphorylation is a crucial mechanism in cellular signaling and regulates proliferation, differentiation, migration and adhesion. The phosphorylation reaction is reversible and is governed by two families of enzymes: protein tyrosine kinases and protein tyrosine phosphatases (PTPs). This thesis investigates the role of PTPs in regulating receptor protein tyrosine kinases (RTKs), and explores a mechanism for regulation of phosphatase activity.

Most receptor tyrosine kinases are activated by ligand induced dimerization, which results in an increase in receptor phosphorylation. Preparations of ligand-stimulated dimeric PDGF β-receptors were shown to be less susceptible to dephosphorylation compared with unstimulated receptors. This revealed that reduced receptor dephosphorylation contributes to ligand-induced increase in RTK phosphorylation.

The receptor-like phosphatase DEP-1 site-selectively dephosphorylates the PDGF β-receptor. One of the most preferred sites is the PLC-γ binding phosphotyrosine pY1021, and the autoregulatory pY857 is one of the least preferred sites. By using chimeric phospho-peptides derived from these two sites as substrate for DEP-1, it was shown that a lysine residue at position +3 acts as a negative determinant for DEP-1 and that an aspartic acid residue at position –1 is a positive determinant.

The modulatory effect of TC-PTP on PDGF β-receptor signaling was explored by using mouse embryonic fibroblasts derived from TC-PTP knockout mice. PDGF β-receptors derived from knockout cells exhibited a higher level of ligand-induced phosphorylation compared to receptors from wildtype cells. The increase was unevenly distributed between different autophosphorylation sites. The PLC-γ binding site, previously implicated in chemotactic response, displayed the largest increase. Consistently, a cell migration assay revealed hyper-responsiveness to PDGF of TC-PTP knockout cells as compared to wildtype cells.

Reversible oxidation of the active site cysteine in PTPs is a mechanism, which have been postulated to regulate phosphatase specific activity. An antibody-based generic method for detection of oxidized PTPs was developed. Using this method it was revealed for the first time that UV-induced inactivation of PTPs involves oxidation of the active site cysteine.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2003. 54 p.
Series
Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 0282-7476 ; 1232
Keyword
Cell and molecular biology, protein tyrosine phosphatase, tyrosine kinase, Cell- och molekylärbiologi
National Category
Biochemistry and Molecular Biology
Research subject
Molecular Cellbiology
Identifiers
urn:nbn:se:uu:diva-3345 (URN)91-554-5556-5 (ISBN)
Public defence
2003-04-04, B42, BMC, Uppsala, 13:15
Opponent
Supervisors
Available from: 2003-03-14 Created: 2003-03-14Bibliographically approved

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