Department of Nanobiology, Graduate School of Advanced Integration Science, Chiba University, Chiba 263-8522, Japan.
Mol Biol Cell. 2013 Jun;24(12):1933-46. doi: 10.1091/mbc.E12-12-0851. Epub 2013 Apr 24.
We identify Xenopus ADF/cofilin (XAC) and its activator, Slingshot phosphatase (XSSH), as key regulators of actin dynamics essential for spindle microtubule assembly during Xenopus oocyte maturation. Phosphorylation of XSSH at multiple sites within the tail domain occurs just after germinal vesicle breakdown (GVBD) and is accompanied by dephosphorylation of XAC, which was mostly phosphorylated in immature oocytes. This XAC dephosphorylation after GVBD is completely suppressed by latrunculin B, an actin monomer-sequestering drug. On the other hand, jasplakinolide, an F-actin-stabilizing drug, induces dephosphorylation of XAC. Effects of latrunculin B and jasplakinolide are reconstituted in cytostatic factor-arrested extracts (CSF extracts), and XAC dephosphorylation is abolished by depletion of XSSH from CSF extracts, suggesting that XSSH functions as an actin filament sensor to facilitate actin filament dynamics via XAC activation. Injection of anti-XSSH antibody, which blocks full phosphorylation of XSSH after GVBD, inhibits both meiotic spindle formation and XAC dephosphorylation. Coinjection of constitutively active XAC with the antibody suppresses this phenotype. Treatment of oocytes with jasplakinolide also impairs spindle formation. These results strongly suggest that elevation of actin dynamics by XAC activation through XSSH phosphorylation is required for meiotic spindle assembly in Xenopus laevis.
我们鉴定出非洲爪蟾 ADF/cofilin（XAC）及其激活剂 Slingshot 磷酸酶（XSSH）是纺锤体微管组装所必需的肌动蛋白动力学的关键调节因子，这对于非洲爪蟾卵母细胞成熟至关重要。XSSH 在尾部结构域的多个位点的磷酸化发生在生发泡破裂（GVBD）之后，并且伴随着 XAC 的去磷酸化，XAC 在未成熟卵母细胞中主要被磷酸化。GVBD 后 XAC 的这种去磷酸化完全被 Latrunculin B 抑制，Latrunculin B 是一种肌动蛋白单体隔离药物。另一方面，jasplakinolide 是一种稳定 F-肌动蛋白的药物，诱导 XAC 的去磷酸化。Latrunculin B 和 jasplakinolide 的作用在细胞停滞因子（CSF）提取物中得到重建，并且通过从 CSF 提取物中耗尽 XSSH 消除了 XAC 的去磷酸化，这表明 XSSH 作为肌动蛋白丝传感器发挥作用，通过 XAC 激活促进肌动蛋白丝动力学。注射抗 XSSH 抗体可阻止 GVBD 后 XSSH 的完全磷酸化，从而抑制减数分裂纺锤体的形成和 XAC 的去磷酸化。与抗体共注射组成性激活的 XAC 可抑制该表型。用 jasplakinolide 处理卵母细胞也会损害纺锤体的形成。这些结果强烈表明，通过 XSSH 磷酸化激活 XAC 引起的肌动蛋白动力学升高对于非洲爪蟾减数分裂纺锤体的组装是必需的。
