Phosphatidylinositol 3-kinase associates, via its Src homology 2 domains, with the activated erythropoietin receptor.
The erythropoietin receptor (EpR) belongs to a family of hematopoietin receptors whose members lack tyrosine kinase activity. Nonetheless, within minutes of binding Ep, a number of cellular proteins become transiently phosphorylated on tyrosine residues. One of these proteins, as we and others have shown previously, is the EpR itself. To identify the remaining protein substrates, we have examined the antiphosphotyrosine immunoprecipitates of lysates from Ba/F3 cells expressing high levels of cell surface EpRs. We now present data showing that, in response to Ep, the 85-Kd regulatory subunit of phosphatidylinositol 3-kinase (PI 3-kinase) becomes immunoprecipitable with antiphosphotyrosine antibodies. This appears to be due, in large part, to the specific association of PI 3-kinase with the tyrosine-phosphorylated EpR, either directly or through a 93- or 70-Kd tyrosine-phosphorylated intermediate. The activity of this EpR associated PI 3-kinase, assessed in anti-EpR immunoprecipitates, is maximal within 2 minutes of incubation with Ep and returns almost to baseline levels by 10 minutes. In vitro studies suggest that the interaction between PI 3-kinase and the activated EpR is mediated by the N- and C-terminal SH2 domains of p85 and tyrosine-phosphorylated motifs on the EpR.
Damen
J E
JE
Terry Fox Laboratory, B.C. Cancer Research Centre, Vancouver, Canada.
Mui
A L
AL
Puil
L
L
Pawson
T
T
Krystal
G
G
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D002460:Cell Line; D007163:Immunosorbent Techniques; D051379:Mice; D008969:Molecular Sequence Data; D019869:Phosphatidylinositol 3-Kinases; D010770:Phosphotransferases / Q000737:chemistry / Q000378:metabolism*; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000378:metabolism*; D017467:Receptors, Erythropoietin / Q000378:metabolism*; D014443:Tyrosine / Q000031:analogs & derivatives / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D017467:Receptors, Erythropoietin; D019000:Phosphotyrosine; D014443:Tyrosine; D010770:Phosphotransferases; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases