Reverse genetic analysis of the yeast RSC chromatin remodeler reveals a role for RSC3 and SNF5 homolog 1 in ploidy maintenance.
Reverse genetic analysis of the yeast RSC chromatin remodeler reveals a role for RSC3 and SNF5 homolog 1 in ploidy maintenance.
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DOI：
10.1371/journal.pgen.0030092
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发表时间：
2007-06
期刊：
影响因子：
4.5
通讯作者：
Logie C
中科院分区：
文献类型：
--
作者：
Campsteijn C;Wijnands-Collin AM;Logie C
酵母“重塑染色质结构”(RSC)复合体是一种多亚基“开关缺陷/蔗糖非发酵”型ATP依赖的核小体重构体，与人类相应的核小体重构体是公认的肿瘤抑制因子。使用温度诱导的所有RSC亚基的降解融合，我们开始绘制RSC需求作为有丝分裂细胞周期的函数。我们发现RSC在G1、G2和有丝分裂过程中执行必要的功能。值得注意的是，当RSC亚单位SFH1、酵母hSNF5肿瘤抑制基因同源基因和RSC3的退化等位基因结合时，我们观察到染色体互补增加了一倍。通过敲除S期细胞周期蛋白CLB5和瞬时耗尽复制起点许可因子CDC6P，取消了同时解除SFH1和RSC3的调控以诱导这些倍体转变的需要。此外，结合sfh1和rsc3的退化等位基因和9个与cdc28/cdk1相关的细胞周期蛋白的缺失等位基因，揭示了S期细胞周期蛋白基因clb5和rsc3之间强烈而特异的遗传相互作用，表明Rsc3p在适当的S期调控中发挥了作用。综上所述，我们的结果表明RSC参与了G1/S相变的调节，并确立了RSC介导的染色质重塑在倍性维持中迄今未被预料到的作用。一些负责改变染色体三维结构的分子以十多种不同蛋白质的复合体的形式工作，其中许多在真菌、植物和动物中都是保守的。有两个这样的复合体在酵母中被称为“重塑染色质结构”(RSC)，在人类中被称为“开关缺陷/蔗糖非发酵”(SWI/SNF)。已知SWI/SNF可抑制多种人类癌症的发生。由于癌症是一种细胞过度分裂的疾病，我们试图确定在酵母细胞分裂周期中，RSC何时执行必要的功能。使用一种通用的方法在其他健康的酵母细胞中诱导必需蛋白的失活，我们发现RSC复合体在染色体复制之前和染色体分离之前都是重要的。有趣的是，将我们产生的两个突变结合在一起，会导致酵母的整个染色体组加倍。众所周知，细胞染色体互补的这种倍增导致遗传遗传的可塑性增加，这本身就是人类癌症一些侵袭性特征的基础，我们的发现为为什么SWI/SNF是如此有效的肿瘤抑制因子提供了新的模型，这反过来可能为癌症治疗提供有价值的新途径。
The yeast “remodels the structure of chromatin” (RSC) complex is a multi-subunit “switching deficient/sucrose non-fermenting” type ATP-dependent nucleosome remodeler, with human counterparts that are well-established tumor suppressors. Using temperature-inducible degron fusions of all the essential RSC subunits, we set out to map RSC requirement as a function of the mitotic cell cycle. We found that RSC executes essential functions during G1, G2, and mitosis. Remarkably, we observed a doubling of chromosome complements when degron alleles of the RSC subunit SFH1, the yeast hSNF5 tumor suppressor ortholog, and RSC3 were combined. The requirement for simultaneous deregulation of SFH1 and RSC3 to induce these ploidy shifts was eliminated by knockout of the S-phase cyclin CLB5 and by transient depletion of replication origin licensing factor Cdc6p. Further, combination of the degron alleles of SFH1 and RSC3, with deletion alleles of each of the nine Cdc28/Cdk1-associated cyclins, revealed a strong and specific genetic interaction between the S-phase cyclin genes CLB5 and RSC3, indicating a role for Rsc3p in proper S-phase regulation. Taken together, our results implicate RSC in regulation of the G1/S-phase transition and establish a hitherto unanticipated role for RSC-mediated chromatin remodeling in ploidy maintenance. Some molecules responsible for altering the 3-D organization of chromosomes work as complexes of more than ten different proteins, and many are conserved in fungi, plants, and animals. Two such complexes are called “remodels the structure of chromatin” (RSC) in yeast and “switching deficient/sucrose non-fermenting” (SWI/SNF) in man. SWI/SNF is known to inhibit the advent of multiple types of human cancers. Since cancer is a disease whereby cells unduly divide, we sought to define when in the yeast cell division cycle RSC executes essential functions. Using a generic method to induce inactivation of essential proteins in otherwise healthy yeast cells, we found that the RSC complex is important before chromosome replication as well as before chromosome segregation. Interestingly, combining two of the mutations we had generated caused doubling of the entire chromosome complement of yeast. As it is known that such multiplication of the cellular chromosome complements results in an increased malleability of the genetic patrimony, which itself is known to underlie some of the aggressive traits of human cancers, our discovery suggests new models as to why SWI/SNF is such a potent tumor suppressor, and this may in turn provide valuable new inroads for cancer treatment.
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