Sin3A recruits Tet1 to the PAH1 domain via a highly conserved Sin3-Interaction Domain.

The Sin3A complex acts as a transcriptional hub, integrating the function of diverse transcription factors with histone modifying enzymes, notably, histone deacetylases (HDAC) 1 and 2. The Sin3A protein sits at the centre of the complex, mediating multiple simultaneous protein-protein interactions via its four paired-amphipathic helix (PAH) domains (PAH1-4). The PAH domains contain a conserved four helical bundle, generating a hydrophobic cleft into which the single-helix of a Sin3-interaction domain (SID) is able to insert and bind with high affinity. Although they share a similar mode of interaction, the SIDs of different repressor proteins bind to only one of four potential PAH domains, due to the specific combination of hydrophobic residues at the interface. Here we report the identification of a highly conserved SID in the 5-methylcytosine dioxygenase, Tet1 (Tet1-SID), which interacts directly with the PAH1 domain of Sin3A. Using a combination of NMR spectroscopy and homology modelling we present a model of the PAH1/Tet1-SID complex, which binds in a Type-II orientation similar to Sap25. Mutagenesis of key residues show that the 11-amino acid Tet1-SID is necessary and sufficient for the interaction with Sin3A and is absolutely required for Tet1 to repress transcription in cells.

Sin3A复合物作为一个转录枢纽，将多种转录因子的功能与组蛋白修饰酶，特别是组蛋白去乙酰化酶（HDAC）1和2整合在一起。Sin3A蛋白位于该复合物的中心，通过其四个成对的两亲性螺旋（PAH）结构域（PAH1 - 4）介导多种同时发生的蛋白质 - 蛋白质相互作用。PAH结构域包含一个保守的四螺旋束，形成一个疏水裂缝，Sin3相互作用结构域（SID）的单螺旋能够插入其中并高亲和力结合。尽管它们具有相似的相互作用模式，但由于界面处疏水残基的特定组合，不同阻遏蛋白的SID仅与四个潜在的PAH结构域中的一个结合。在此我们报道在5 - 甲基胞嘧啶双加氧酶Tet1中鉴定出一个高度保守的SID（Tet1 - SID），它直接与Sin3A的PAH1结构域相互作用。通过结合核磁共振波谱和同源建模，我们提出了PAH1/Tet1 - SID复合物的模型，其以类似于Sap25的II型取向结合。关键残基的诱变表明，11个氨基酸的Tet1 - SID对于与Sin3A的相互作用是必要且充分的，并且对于Tet1在细胞中抑制转录是绝对必需的。