Mitogenic Roles of Gab 1 and Grb 10 as Direct Cellular Partners in the Regulation of MAP Kinase Signaling

Functions of signaling mediators Grb 10 or Gab 1 have been described in mitogenesis but remained disconnected. Here, we report the peptide hormone-dependent direct association between Grb 10 and Gab 1 and their functional connection in mitogenic signaling via MAP kinase using cultured fibroblasts as a model. In response to PDGF-, IGF-I, or insulin increased levels of Grb 10 potentiated cell proliferation or survival whereas dominant-negative, domain-specific Grb 10 peptide mimetics attenuated cell proliferation. This response was sensitive to p44/42 MAPK inhibitor but not to p38 MAPK inhibitor. In response to IGF-I or insulin Raf-1, MEK 1/2, and p44/42 MAPK were regulated by Grb 10 but not Ras or p38 MAPK. In response to PDGF MEK 1/2, p44/42 MAPK and p38 MAPK were regulated by Grb 10 but not Ras or Raf-1. Peptide hormone-dependent co-immunoprecipitation of Grb 10 and Gab 1 was demonstrated and specifically blocked by a Grb 10 SH2 domain peptide mimetic. This domain was sufficient for direct, peptide hormone-dependent association with Gab 1 via the Crk binding region. In response to PDGF, IGF-I, or insulin, in a direct comparison, elevated levels of mouse Grb 10 delta, or human Grb 10 beta or zeta equally potentiated fibroblast proliferation. Proliferation was severely reduced by Gab 1 gene disruption whereas an elevated Gab 1 gene dose proportionally stimulated Grb 10-potentiated cell proliferation. In conclusion, Gab 1 and Grb 10 function as direct binding partners in the regulation of the mitogenic MAP kinase signal. In cultured fibroblasts, elevated levels of human Grb 10 beta, zeta or mouse Grb 10 delta comparably potentiate mitogenesis in response to PDGF, IGF-I, or insulin. J. Cell. Biochem. 105: 1172-1182, 2008. (c) 2008 Wiley-Liss, Inc.