Interleukin-3 induces association of the protein-tyrosine phosphatase SHP2 and phosphatidylinositol 3-kinase with a 100-kDa tyrosine-phosphorylated protein in hemopoietic cells.
We have observed previously the co-immunoprecipitation of the p85 subunit of phosphatidylinositol-3 kinase (PI3K) and SHP2 in murine lymphohemopoietic cells after stimulation with interleukin-3. We have investigated this interaction in more detail and now report the identification of a potentially novel 100-kDa protein (termed p100), which is inducibly phosphorylated on tyrosine after interleukin-3 treatment and which co-immunoprecipitates with both p85 PI3K and SHP2. The Src homology region 2 domains of both p85 and SHP2 appear to mediate their interactions with p100. Sequential precipitation analyses suggest that these interactions are direct and do not involve Grb2, and that the same p100 protein, or a portion of it, interacts with both p85 and SHP2, implying that p100 may serve to link these two proteins. Far Western blotting with both full-length p85 and isolated p85 Src homology region 2 domains supports this view. Interestingly, p100 also appears to be a substrate for the SHP2 phosphatase activity. In addition, p100 is precipitated by Grb2-glutathione S-transferase fusion proteins, an interaction largely mediated by the Grb2 SH3 domains. p100 appears to be distinct from JAK2, Vav, STAT5, and c-Cbl. Although largely cytosolic, p100 can be detected associated with SHP2 and PI3K in crude membrane fractions after interleukin-3 stimulation. We propose that p100 plays a role as an adaptor molecule, linking PI3K and SHP2 in IL-3 signaling.
Craddock
B L
BL
Pharmacology Group, Department of Pharmacy and Pharmacology, University of Bath, Bath BA2 7AY, United Kingdom.
Welham
M J
MJ
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D001854:Bone Marrow Cells / Q000201:enzymology*; D002478:Cells, Cultured; D051380:GRB2 Adaptor Protein; D007377:Interleukin-3 / Q000494:pharmacology*; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism*; D011485:Protein Binding; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D013379:Substrate Specificity
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495949:Grb2 protein, mouse; D007377:Interleukin-3; D047908:Intracellular Signaling Peptides and Proteins; D011506:Proteins; D019000:Phosphotyrosine; D019869:Phosphatidylinositol 3-Kinases; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases; C516725:Ptpn11 protein, mouse; C509248:Ptpn6 protein, mouse