Actin assembly mediated by a nucleation promoting factor WASH is involved in MTOC-TMA formation during Xenopus oocyte...

During Xenopus oocyte maturation, the microtubule organizing center and transient microtubule array (MTOC-TMA) forms at the vegetal side of the nucleus and we have previously demonstrated that this formation requires actin reorganization. However, the molecular mechanism that coordinates actin filaments with microtubules (MTs) during oocyte maturation is unknown. Here we show that the actin nucleation promoting factor WASH is involved in MTOC-TMA assembly. WASH depletion disrupted the alignment of actin filaments and MT at the base of MTOC-TMA. In vitro cosedimentation assays revealed that the Xenopus WASH VCA domain, which comprises the actin and Arp2/3 complex binding region, directly bound to MT but that the WHD2 domain, which had been previously reported to be an MT-binding domain of mammalian WASH, did not. Injection of the WHD2-deleted mutant into oocytes caused a drastic accumulation of actin filaments in the cytoplasm and malformation of MTOC-TMA, suggesting that the WHD2 domain negatively regulates the VCA domain activity during oocyte maturation. Our results suggest that WASH complex-dependent actin nucleation is involved in the assembly of MTOC-TMA.