MUTATIONS IN THE KINESIN-LIKE PROTEIN EG5 DISRUPTING LOCALIZATION TO THE MITOTIC SPINDLE
MUTATIONS IN THE KINESIN-LIKE PROTEIN EG5 DISRUPTING LOCALIZATION TO THE MITOTIC SPINDLE
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DOI：
10.1073/pnas.92.10.4289
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发表时间：
1995-05-09
影响因子：
11.1
通讯作者：
MITCHISON, TJ
中科院分区：
文献类型：
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作者：
SAWIN, KE;MITCHISON, TJ
Eg5是驱动蛋白样微管马达蛋白bimC亚家族的成员，在有丝分裂中定位于纺锤体微管，但不定位于间期微管。我们研究了纺锤体定位的分子基础，通过瞬时转染爪蟾A6细胞与myc标记的Eg5衍生物。mycEg5蛋白在巨细胞病毒启动子的组成性高水平表达，在整个间期细胞质中，在早期前期开始结合微管，并通过有丝分裂到末期结束保持定位于纺锤体和/或中间体微管。Eg5的N端和C端区域都是这种细胞周期调节靶向所必需的。Eg5在其C-末端结构域中还包含bimC亚家族蛋白中保守的序列，其包括潜在的p34（cdc2）磷酸化位点。我们发现，一个单一的苏氨酸（T937）在这个网站nonphosphorylatable丙氨酸突变废除本地化的突变蛋白质的纺锤体，而突变的T937丝氨酸保留纺锤体本地化。我们推测Eg5的磷酸化可能调节其在细胞周期中的纺锤体定位。
Eg5, a member of the bimC subfamily of kinesin-like microtubule motor proteins, localizes to spindle microtubules in mitosis but not to interphase microtubules. We investigated the molecular basis for spindle localization by transient transfection of Xenopus A6 cells with myc-tagged derivatives of Eg5. Expressed at constitutively high levels from a cytomegalovirus promoter, mycEg5 protein is cytoplasmic throughout interphase, begins to bind microtubules in early prophase, and remains localized to spindle and/or midbody microtubules through mitosis to the end of telophase. Both Nand C-terminal regions of Eg5 are required for this cell-cycle-regulated targeting. Eg5 also contains within its C-terminal domain a sequence conserved among bimC subfamily proteins that includes a potential p34(cdc2) phosphorylation site. We show that mutation of a single threonine (T937) within this Site to nonphosphorylatable alanine abolishes localization of the mutant protein to the spindle, whereas mutation of T937 to serine preserves spindle localization. We hypothesize that phosphorylation of Eg5 may regulate its localization to the spindle in the cell cycle.