The spindle pole body assembly component Mps3p/Nep98p functions in sister chromatid cohesion
The spindle pole body assembly component Mps3p/Nep98p functions in sister chromatid cohesion
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DOI：
10.1074/jbc.m404324200
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发表时间：
2004-11-19
影响因子：
4.8
通讯作者：
Skibbens, RV
中科院分区：
文献类型：
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作者：
Antoniacci, LM;Kenna, MA;Skibbens, RV
在有丝分裂过程中，为了实现染色体的成功分离，在细胞周期的早期必须发生几个过程，包括纺锤体极体复制、DNA复制以及新生姐妹染色单体之间内聚力的建立。纺锤体极体复制在G₁期开始，并在S期早期随着纺锤体极体成熟并开始分离而持续进行。纺锤体极体复制的关键步骤是核膜跨膜蛋白Msp3p/Nep98p（在此称为Mps3p）依次招募Cdc31p和Spc42p。与DNA复制同时，Ctf7p/Eco1p（在此称为Ctf7p）确保新生姐妹染色单体配对在一起，将复制产物识别为姐妹染色单体。在此，我们提供了第一个证据，即核膜纺锤体极体组装成分Mps3p对姐妹染色单体的内聚力起着关键作用。通过全基因组双杂交筛选，Mps3p被鉴定为与Ctf7p相互作用，并且通过体内（免疫共沉淀）和体外（GST pull - down）实验证实了这种物理相互作用。对新的mps3/nep98等位基因进行的体内内聚力测定表明，Mps3p的缺失导致姐妹染色单体过早分离，并且Mps3p在G₁期之后起作用，与Ctf7p的作用时间一致。在有丝分裂期间，内聚力的维持不需要Mps3p，这表明Mps3p在内聚力的建立而非维持中起作用。导致内聚力缺陷的突变Mps3p在体外不再与Ctf7p结合，这表明Mps3p和Ctf7p之间的相互作用具有生理相关性。为支持这一模型，mps3 ctf7双突变细胞表现出条件性合成致死。这些发现证明了Mps3p在姐妹染色单体的内聚力方面的新作用，并为纺锤体极体成分发生突变时导致非整倍性的机制提供了新的见解。
For successful chromosome segregation during mitosis, several processes must occur early in the cell cycle, including spindle pole duplication, DNA replication, and the establishment of cohesion between nascent sister chromatids. Spindle pole body duplication begins in G(1) and continues during early S-phase as spindle pole bodies mature and start to separate. Key steps in spindle pole body duplication are the sequential recruitment of Cdc31p and Spc42p by the nuclear envelope transmembrane protein Msp3p/Nep98p (herein termed Mps3p). Concurrent with DNA replication, Ctf7p/Eco1p (herein termed Ctf7p) ensures that nascent sister chromatids are paired together, identifying the products of replication as sister chromatids. Here, we provide the first evidence that the nuclear envelope spindle pole body assembly component Mps3p performs a function critical to sister chromatid cohesion. Mps3p was identified as interacting with Ctf7p from a genome-wide two-hybrid screen, and the physical interaction was confirmed by both in vivo (co-immunoprecipitation) and in vitro (GST pull-down) assays. An in vivo cohesion assay on new mps3/nep98 alleles revealed that loss of Mps3p results in precocious sister chromatid separation and that Mps3p functions after G(1), coincident with Ctf7p. Mps3p is not required for cohesion during mitosis, revealing that Mps3p functions in cohesion establishment and not maintenance. Mutated Mps3p that results in cohesion defects no longer binds to Ctf7p in vitro, demonstrating that the interaction between Mps3p and Ctf7p is physiologically relevant. In support of this model, mps3 ctf7 double mutant cells exhibit conditional synthetic lethality. These findings document a new role for Mps3p in sister chromatid cohesion and provide novel insights into the mechanism by which a spindle pole body component, when mutated, contributes to aneuploidy.