Tyrosine 508 of the 85-kilodalton subunit of phosphatidylinositol 3-kinase is phosphorylated by the platelet-derived growth factor receptor.
The mechanisms by which growth factors and oncogenic agents activate phosphatidylinositol 3-kinase (PI3 kinase) are unknown. Previously, we reported that the 85-kDa regulatory subunit of PI3 kinase is tyrosine-phosphorylated both in vitro by the platelet-derived growth factor beta-receptor (PDGFR) tyrosine kinase and in fibroblasts in response to PDGF. As a first step in determining the role of tyrosine phosphorylation in PDGF signaling through PI3 kinase, we investigated which tyrosines on p85 are phosphorylated by the PDGFR. Recombinant p85 was phosphorylated with recombinant PDGF receptors, and tryptic phosphopeptides were purified by HPLC and analyzed by Edman degradation. By this approach and by mutational analysis, Y508 was identified as the major in vitro phosphorylation site. Tryptic phosphopeptide mapping demonstrated Y508 to also be phosphorylated in vivo in COS cells. Comparison of these data with a previous report [Hayashi, H., Nishioka, Y., Kamohara, S., Kanai, F., Ishii, K., Fukui, Y., Shibasaki, F., Takenawa, T., Kido, H., Katsunuma, N., & Ebina, Y. (1993) J. Biol. Chem. 268, 7107-7117] suggests that p85 is phosphorylated differently by the PDGF and insulin receptor tyrosine kinases. Therefore, p85 may be regulated differently by PDGF and insulin. Mapping of phosphorylation sites on p85 may lead to new insights into the regulation of signal transduction through PI3 kinase.
Kavanaugh
W M
WM
Department of Medicine, University of California, San Francisco 94143.
Turck
C W
CW
Klippel
A
A
Williams
L T
LT
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D051379:Mice; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D019869:Phosphatidylinositol 3-Kinases; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry / Q000378:metabolism*; D017479:Receptors, Platelet-Derived Growth Factor / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism*
D011994:Recombinant Proteins; D014443:Tyrosine; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D017479:Receptors, Platelet-Derived Growth Factor