Mapping of effective protein domains is a demanding stride to disclose the functional relationship between regulatory complexes. Domain analysis of protein interactions is requisite for understanding the pleiotropic responses of the respective partners. Cti6 is a multifunctional regulator for which we could show recruitment of co-repressors Sin3, Cyc8 and Tup1. However, the responsible core domain tethering Cti6 to these co-repressors is poorly understood. Here, we report the pivotal domain of Cti6 that is indispensable for co-repressor recruitment. We substantiated that amino acids 450–506 of Cti6 bind PAH2 of Sin3. To analyse this Cti6–Sin3 Interaction Domain (CSID) in more detail, selected amino acids within CSID were replaced by alanine. It is revealed that hydrophobic amino acids V467, L481 and L491 L492 L493 are important for Cti6–Sin3 binding. In addition to PAH2 of Sin3, CSID also binds to tetratricopeptide repeats (TPR) of Cyc8. Indeed, we could demonstrate Cti6 recruitment to promoters of genes, such as RNR3 and SMF3, containing iron-responsive elements (IRE). Importantly, Sin3 is also recruited to these promoters but only in the presence of functional Cti6. Our findings provide novel insights toward the critical interaction domain in the co-regulator Cti6, which is a component of regulatory complexes that are closely related to chromatin architecture and the epigenetic status of genes that are regulated by pleiotropic co-repressors.

有效蛋白质结构域的定位对于揭示调节复合物之间的功能关系是一项艰巨的任务。蛋白质相互作用的域分析是理解各个伴侣的多效性反应所必需的。Cti6是一个多功能调节器，我们可以为它显示辅助阻遏物Sin3，Cyc8和Tup1的募集。但是，对将Cti6绑定到这些共阻遏物的负责任的核心域了解甚少。在这里，我们报告了Cti6的关键域，这对于共阻遏物募集是必不可少的。我们证实了Cti6的450-506位氨基酸与Sin3的PAH2结合。为了更详细地分析此Cti6-Sin3相互作用域（CSID），将CSID中选定的氨基酸替换为丙氨酸。据揭示疏水性氨基酸V467，L481和L491 L492 L493对于Cti6–Sin3结合很重要。除Sin3的PAH2外，CSID还与Cyc8的四三肽重复序列（TPR）结合。确实，我们可以证明Cti6募集到基因的启动子，例如RNR3和SMF3，包含铁响应元素（IRE）。重要的是，Sin3也被募集到这些启动子中，但仅在功能性Cti6存在的情况下。我们的发现为共调节子Cti6中的关键相互作用域提供了新颖的见解，Cti6是与染色体结构和多效性共阻遏物调节的基因的表观遗传状态密切相关的调节复合物的一个组成部分。