Effect of genistein alone and in combination with okadaic acid on the cellcycle resumption of mouse oocytes

Our biopharmacological approach suggests that the now well-documented inhib itory effects of genistein on the maturation of mammalian oocytes do not se em to be related to its effect on tyrosine kinases. Indeed, we show that bo th tyrphostin B46 and Lavendustin A, two selective inhibitors of protein ty rosine kinases, fail to inhibit meiosis reinitiation. According to recent f indings, the G2/M arrest induced by genistein could be due to inhibition of the kinase activity of cdc2. We were therefore mainly interested in dissec ting the cytological effects of genistein on mouse primary and secondary oo cytes. Genistein exerts the same cytological effects as IBMX on primary ooc ytes: their germinal vesicle is maintained in a central position, the cytop lasmic microtubule network is stabilized, the central GV immobilization is overcome by demecolcine and they complete normal maturation after their tra nsfer to culture medium. The GV-arresting activity of genistein is also byp assed by OA but combination of both drugs results in a dramatic reorganizat ion of the cytoskeleton leading to a huge membrane bulging, which is quite different to apoptotic-related blebbing. MAP Kinase activation is correlate d with meiosis reinitiation. When applied after GVBD has taken place, genis tein does not inhibit MAPK activation, metaphase spindle formation and meta phase-to-anaphase transition. but prevents the barrel-shaped MI spindle fro m undergoing its peripheral migration and the oocytes from extruding their first polar body. It may thus be concluded that the checkpoint control for anaphase onset is unaffected by the drug. On the contrary, our results sugg est that spindle anaphase A to spindle anaphase B transition, spindle degra dation, mid-body formation and cytokinesis are triggered by a genistein-sen sitive mechanism that might be a mid-anaphase checkpoint. Finally, we confi rm that genistein induces transition to interphase in metaphase II oocytes but never induces cortical granule exocytosis, the cytoplasmic hallmark of activation.