The Sos1-Rac1 signaling - Possible involvement of a vacuolar H+ -ATPase E subunit

We have purified and identified a 32-kDa protein interacting with the Dbl o ncogene homology domain of mSos1(Sos-DH) from rat brains by glutathione S-t ransferase-Sos-DH affinity chromatography. Peptide sequencing revealed that the protein is identical to a positive regulatory E subunit (V-ATPase E) o f a vacuolar H+-ATPase, which is responsible for acidification of endosome and alkalinization of intracellular pH. The interaction between V-ATPase E and Sos-DH was confirmed by yeast two-hybrid assay. A coimmunoprecipitation assay demonstrated that a V-ATPase E protein physiologically bound to mSos 1, and the protein was colocalized with mSos1 in the cytoplasm, as determin ed by immunohistochemistry. mSos1 was found in the early endosome fraction together with V-ATPase E and Rac1, suggesting the functional involvement of mSos1/V-ATPase E complexes in the Rac1 activity at endosomes. Overexpressi on of V-ATPase E in COS cells enhanced the ability of mSos1 to promote the guanine nucleotide exchange activity for Rac1 and stimulated the kinase act ivity of Jun kinase, a downstream target of Rac1. Thus, the data indicate t hat V-ATPase E may participate in the regulation of the mSos1-dependent Rac 1 signaling pathway involved in growth factor receptor-mediated cell growth control.