Samel Anke, Rudner Adam, Ehrenhofer-Murray Ann E
Institut für Biologie, Humboldt-Universität zu Berlin, Berlin 10099, Germany.
Department of Biochemistry, Microbiology and Immunology, Ottawa Institute of Systems Biology, University of Ottawa, Ontario ON K1H 8M5, Canada.
G3 (Bethesda). 2017 Apr 3;7(4):1117-1126. doi: 10.1534/g3.116.037739.
Heterochromatin formation in the yeast is characterized by the assembly of the Silent Information Regulator (SIR) complex, which consists of the histone deacetylase Sir2 and the structural components Sir3 and Sir4, and binds to unmodified nucleosomes to provide gene silencing. Sir3 contains an AAA ATPase-like domain, and mutations in an exposed loop on the surface of this domain abrogate Sir3 silencing function , as well binding to the Sir2/Sir4 subcomplex. Here, we found that the removal of a single methyl group in the C-terminal coiled-coil domain (mutation T1314S) of Sir4 was sufficient to restore silencing at the silent mating-type loci and to a Sir3 version with a mutation in this loop. Restoration of telomeric silencing required further mutations of Sir4 (E1310V and K1325R). Significantly, these mutations in Sir4 restored complex formation between Sir3 and the Sir4 coiled-coil, indicating that the improved affinity between Sir3 and Sir4 is responsible for the restoration of silencing. Altogether, these observations highlight remarkable properties of selected amino-acid changes at the Sir3-Sir4 interface that modulate the affinity of the two proteins.
酵母中的异染色质形成以沉默信息调节因子（SIR）复合物的组装为特征，该复合物由组蛋白去乙酰化酶Sir2以及结构成分Sir3和Sir4组成，并与未修饰的核小体结合以实现基因沉默。Sir3包含一个类似AAA ATP酶的结构域，该结构域表面一个暴露环上的突变会消除Sir3的沉默功能，以及与Sir2/Sir4亚复合物的结合。在这里，我们发现，在Sir4的C末端卷曲螺旋结构域中去除单个甲基（T1314S突变）足以恢复沉默交配型位点的沉默，并恢复该环中存在突变的Sir3版本的沉默。端粒沉默的恢复需要Sir4进一步的突变（E1310V和K1325R）。值得注意的是，Sir4中的这些突变恢复了Sir3与Sir4卷曲螺旋之间的复合物形成，表明Sir3和Sir4之间亲和力的提高是沉默恢复的原因。总之，这些观察结果突出了Sir3-Sir4界面处选定氨基酸变化的显著特性，这些变化调节了两种蛋白质的亲和力。
