The yeast protein kinase Mps1p is required for assembly of the integral spindle pole body component Spc42p
The yeast protein kinase Mps1p is required for assembly of the integral spindle pole body component Spc42p
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DOI：
10.1083/jcb.200111025
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发表时间：
2002-02-04
影响因子：
7.8
通讯作者：
Winey, M
中科院分区：
文献类型：
--
作者：
Castillo, AR;Meehl, JB;Winey, M
酿酒酵母Mps1编码一种必需的蛋白激酶，在纺锤体极体(SPB)复制和纺锤体检查点中发挥作用。以前描述的Mps1突变体在这两个功能上都失败了，导致了异常的DNA分离，并造成了致命的后果。在这里，我们报告了一个独特的条件等位基因Mps1-8的鉴定，该等位基因在SPB复制中存在缺陷，但不存在纺锤体检查点。Mps1-8的突变位于Mps1的非催化区域，对突变蛋白的分析表明，Mps1-8p在体外具有野生型激酶活性。对Mps1-8条件生长表型的剂量抑制因子的筛选确定了编码完整的SPB组分SPC42的基因。进一步的分析表明，当与SPC42的某些突变等位基因结合时，Mps1-8显示出合成生长缺陷。通过免疫荧光显微镜和免疫ENA分析，Mps1p的一个表位标记的版本(Mps1p-myc)定位于SPBS和动蛋白中心。这与我们通过免疫共沉淀检测到的Mps1p和Spc42p之间的物理相互作用是一致的。Spc42p在体外是Mps1p磷酸化的底物，而在体内Spc42p的磷酸化依赖于Mps1p。最后，在Mps1-1突变株中，Spc42p组装异常。我们得出结论，Mps1p调控SPB复制过程中完整SPB组件Spc42p的组装。
Saccharomyces cerevisiae MPS1 encodes an essential protein kinase that has roles in spindle pole body (SPB) duplication and the spindle checkpoint. previously characterized MPS1 mutants fail in both functions, leading to aberrant DNA segregation with lethal consequences. Here, we report the identification of a unique conditional allele, mps1-8, that is defective in SPB duplication but not the spindle checkpoint. The mutations in mps1-8 are in the noncatalytic region of MPS1, and analysis of the mutant protein indicates that Mps1-8p has wild-type kinase activity in vitro. A screen for dosage suppressors of the mps1-8 conditional growth phenotype identified the gene encoding the integral SPB component SPC42. Additional analysis revealed that mps1-8 exhibits synthetic growth defects when combined with certain mutant alleles of SPC42. An epitope-tagged version of Mps1p (Mps1p-myc) localizes to SPBs and kinetochores by immunofluorescence microscopy and immuno-ENA analysis. This is consistent with the physical interaction we detect between Mps1p and Spc42p by coimmunoprecipitation. Spc42p is a substrate for Mps1p phosphorylation in vitro, and Spc42p phosphorylation is dependent on Mps1p in vivo. Finally, Spc42p assembly is abnormal in a mps1-1 mutant strain. We conclude that Mps1p regulates assembly of the integral SPB component Spc42p during SPB duplication.