The p85alpha subunit of phosphatidylinositol 3'-kinase binds to and stimulates the GTPase activity of Rab proteins.
Rab5 and Rab4 are small monomeric GTPases localized on early endosomes and function in vesicle fusion events. These Rab proteins regulate the endocytosis and recycling or degradation of activated receptor tyrosine kinases such as the platelet-derived growth factor receptor (PDGFR). The p85alpha subunit of phosphatidylinositol 3'-kinase contains a BH domain with sequence homology to GTPase activating proteins (GAPs), but has not previously been shown to possess GAP activity. In this report, we demonstrate that p85alpha has GAP activity toward Rab5, Rab4, Cdc42, Rac1 and to a lesser extent Rab6, with little GAP activity toward Rab11. Purified recombinant Rab5 and p85alpha can bind directly to each other and not surprisingly, the p85alpha-encoded GAP activity is present in the BH domain. Because p85alpha stays bound to the PDGFR during receptor endocytosis, p85alpha will also be localized to the same early endosomal compartment as Rab5 and Rab4. Taken together, the physical co-localization and the ability of p85alpha to preferentially stimulate the down-regulation of Rab5 and Rab4 GTPases suggests that p85alpha regulates how long Rab5 and Rab4 remain in their GTP-bound active state. Cells expressing BH domain mutants of p85 show a reduced rate of PDGFR degradation as compared with wild type p85 expressing cells. These cells also show sustained activation of the mitogen-activated protein kinase and Akt pathways. Thus, the p85alpha protein may play a role in the down-regulation of activated receptors through its temporal control of the GTPase cycles of Rab5 and Rab4.
Chamberlain
M Dean
MD
Cancer Research Unit, Health Research Division, Saskatchewan Cancer Agency, Saskatoon, Saskatchewan S7N 4H4, Canada.
Berry
Tangyne R
TR
Pastor
M Chris
MC
Anderson
Deborah H
DH
D000818:Animals; D019556:COS Cells; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism; D015151:Immunoblotting; D047468:Immunoprecipitation; D051379:Mice; D009154:Mutation; D041681:NIH 3T3 Cells; D019869:Phosphatidylinositol 3-Kinases / Q000737:chemistry* / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017479:Receptors, Platelet-Derived Growth Factor / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D013997:Time Factors; D014162:Transfection; D020695:rab4 GTP-Binding Proteins / Q000737:chemistry*; D020696:rab5 GTP-Binding Proteins / Q000737:chemistry*
D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D005982:Glutathione Transferase; D019869:Phosphatidylinositol 3-Kinases; D017479:Receptors, Platelet-Derived Growth Factor; D020558:GTP Phosphohydrolases; D020695:rab4 GTP-Binding Proteins; D020696:rab5 GTP-Binding Proteins