Division of Biochemistry, Faculty of Pharmaceutical Sciences, Tohoku Medical and Pharmaceutical University, 4-4-1 Komatsushima, Aoba-ku, Sendai, Miyagi, 981-8558, Japan.
Division of Biochemistry, Faculty of Pharmaceutical Sciences, Tohoku Medical and Pharmaceutical University, 4-4-1 Komatsushima, Aoba-ku, Sendai, Miyagi, 981-8558, Japan.
Biochem Biophys Res Commun. 2022 Jan 1;586:150-156. doi: 10.1016/j.bbrc.2021.11.077. Epub 2021 Nov 25.
Bi-oriented attachment of microtubules to the centromere is a pre-requisite for faithful chromosome segregation during mitosis. Budding yeast have point centromeres containing the cis-element proteins CDE-I, -II, and -III, which interact with trans-acting factors such as Cbf1, Cse4, and Ndc10. Our previous genetic screens, using a comprehensive library of histone point mutants, revealed that the TBS-I, -II, and -III regions of nucleosomes are required for faithful chromosome segregation. In TBS-III deficient cells, peri-centromeric nucleosomes containing the H2A.Z homolog Htz1 are lacking, however, it is unclear why chromosome segregation is defective in these cells. Here, we show that, in cells lacking TBS-III, both chromatin binding at the centromere and the total amount of some of the centromere proteins are reduced, and transcription through the centromere is up-regulated during M-phase. Moreover, the chromatin binding of Cse4, Mif2, Cbf1, Ndc10, and Scm3 was reduced upon ectopic transcription through the centromere in wild-type cells. These results suggest that transcription through the centromere displaces key centromere proteins and, consequently, destabilizes the interaction between centromeres and microtubules, leading to defective chromosome segregation. The identification of new roles for histone binding residues in TBS-III will shed new light on nucleosome function during chromosome segregation.
微管双定向附着到着丝粒是有丝分裂中染色体正确分离的必要条件。芽殖酵母具有点状着丝粒，包含顺式作用元件蛋白 CDE-I、-II 和 -III，这些蛋白与 Cbf1、Cse4 和 Ndc10 等反式作用因子相互作用。我们之前的遗传筛选使用了组蛋白点突变的综合文库，结果表明核小体的 TBS-I、-II 和 -III 区域对于染色体的正确分离是必需的。在 TBS-III 缺失的细胞中，缺乏含有 H2A.Z 同源物 Htz1 的着丝粒周围核小体，然而，尚不清楚为什么这些细胞的染色体分离出现缺陷。在这里，我们表明，在缺乏 TBS-III 的细胞中，着丝粒处的染色质结合和一些着丝粒蛋白的总量都减少了，并且在 M 期，着丝粒处的转录被上调。此外，在野生型细胞中，通过着丝粒异位转录后，Cse4、Mif2、Cbf1、Ndc10 和 Scm3 的染色质结合减少。这些结果表明，着丝粒处的转录会取代关键的着丝粒蛋白，从而破坏着丝粒与微管之间的相互作用，导致染色体分离缺陷。新发现的 TBS-III 中组蛋白结合残基的作用将为染色体分离过程中核小体的功能提供新的认识。
