ACTIVATION OF THE XENOPUS OOCYTE MITOGEN-ACTIVATED PROTEIN-KINASE PATHWAY BY MOS IS INDEPENDENT OF RAF

Synthesis of the protein kinase Mos is required for progesterone-induc ed activation of MAP kinase, M-phase promoting factor (MPF), and meiot ic maturation of Xenopus oocytes. Mos can function as a MAP kinase kin ase kinase, leading to activation of MAP kinase; how Mos causes activa tion of MPF is not yet known, The protein kinase Raf, which acts as a MAP kinase kinase kinase in somatic cells, also appears to be involved in meiotic maturation, but recent work has suggested that the Raf act s downstream of Mos activity during oocyte maturation. Using an oocyte cell-free system, we report here that a dominant negative Raf, which inhibits Pas-induced MAP kinase activation, does not block Mos-induced activation in vitro. These results indicate that, in contrast to prev ious conclusions, Mos-induced oocyte MAP kinase activation proceeds in dependently of Raf. Using a dominant-negative MAP kinase construct, we also show that most of the mitogen-induced hyperphosphorylation and d ramatic gel retardation of Raf, which is often taken as a marker for t he activation of Raf by upstream components, is actually dependent on, and thus downstream of, MAP kinase activation.