The catalytic subunit of phosphatidylinositol 3-kinase is a substrate for the activated platelet-derived growth factor receptor, but not for middle-T antigen pp60(c-src) complexes

S. Roche, R. Dhand, M. D. Waterfield, S. A. Courtneidge

Research output: Contribution to journalArticle

25 Scopus citations

Abstract

The interaction of phosphatidylinositol 3-kinase (PI 3-K) with polyoma-virus middle-T antigen-pp60(c-src) (mT:cSrc) complexes and with the platelet-derived growth factor (PDGF) receptor has been investigated. Firstly, we undertook reconstitution studies, using proteins derived from a baculovirus expression system. The p110 catalytic subunit of the PI 3-K associated with tyrosine kinases only when complexed with the p85α regulatory subunit. Both p85α and p110 were substrates of the PDGF receptor. In contrast, only the p85α subunit was detectably phosphorylated when PI 3-K was associated with mT:cSrc. Secondly, we studied PI 3-K in mammalian cells. In mT-antigen-transformed NIH-3T3 cells neither p85α nor p110 was phosphorylated on tyrosine residues in vivo, even though p85α was a substrate in kinase assays in vitro. In quiescent NIH-3T3 cells, PI 3-K showed detectable activity in vitro; PDGF stimulation resulted in a rapid and transient association of PI 3-K with the receptor, which was correlated with a transient increase in intrinsic PI 3-K activity (approx. 2-fold). The activated PDGF receptor phosphorylated p110 in vitro, at one major site. In vivo, PDGF stimulation induced tyrosine phosphorylation of p110 that persisted for at least 1 h after stimulation. Immunodepletion of the PDGF receptor from stimulated cell lysates showed that p110 was released from the receptor in a tyrosine-phosphorylated form. From these results we conclude that (i) the mT:cSrc complex and the PDGF receptor differ in their association with PI 3-K activity, (ii) PDGF receptor appears to activate PI 3-K in vivo both by relocation of the enzyme and by stimulation of its intrinsic activity, and (iii) tyrosine phosphorylation of the p110 subunit by the PDGF receptor may play a role in PI 3-K regulation in some circumstances.

Original languageEnglish (US)
Pages (from-to)703-711
Number of pages9
JournalBiochemical Journal
Volume301
Issue number3
DOIs
StatePublished - 1994

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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