The participation of ras gene products in models of eukaryotic proliferation (meiotic maturation of Xenopus oocytes) and differentiation (3T3 L1 preadipocytes) was analyzed. It was demonstrated in 3T3 L1 cells that Ras proteins are obligatory intermediates in insulin signaling pathways of mammalian cells. Activation of the receptor for insulin or IGF-1 triggers the immediate formation of the active Ras-GTP complex and subsequent activation of cytosolic serine-threonine kinases including mitogen activated protein (MAP) and S6 kinases. Ras activation by insulin does not involve GTPase activating protein (GAP) phosphorylation but is associated with tyrosine phosphorylation of the GAP-associated p62 protein. Surprisingly. Raf1 kinase activation is dissociated from activation of the MAP kinases by insulin. This indicates that at least two separate parallel signals emerge from Ras after insulin stimulation of these cells. Microinjection of p21ras into Xenopus oocytes induces maturation promoter factor activation and G2 to M progression. even in the absence of protein synthesis. through a cascade of phosphorylations involving cytosolic MAP kinases prior to cdc2 kinase activation. Purified Ras proteins can also activate the cytoplasmic ERK kinases when added to highly concentrated cell-free extracts of untreated oocytes. Besides the previous role of Ras proteins in G phase of the cell cycle, a role for Ras proteins in the M phase of the cell cycle was also demonstrated. both in oocytes and embryos. Current work includes analysis of the functional role of specific domains (SH2, SH3 and PH) of Ras regulatory proteins and phosphatases in phosphorylation cascades transducing signals leading to cell division in oocytes.
