Abstract
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.
Highlights
The Sin3A protein is the central component of the multi-protein Sin3A complex, which along with its core components, histone deacetylase 1 and 2 (HDAC1/2), regulates chromatin structure and gene expression in all eukaryotic cells[1]
Similar to the Sin[3] interaction domain (SID) of other transcriptional repressors (Mxd[14,5], REST18, HBP23, etc.), the Ten-eleven translocation 1 (Tet1)-Sin3 interaction domain (SID) consists of a single helix with an arrangement of specific hydrophobic residues which form a lock and key interaction with one of four potential Sin3A paired-amphipathic helix (PAH) domains (PAH1-4)
Mapping experiments using Sin3A truncation and point-mutants revealed that the Tet1-SID binds to the PAH1 115–212 (PAH1) domain
Summary
The Sin3A protein is the central component of the multi-protein Sin3A complex, which along with its core components, histone deacetylase 1 and 2 (HDAC1/2), regulates chromatin structure and gene expression in all eukaryotic cells[1]. The complex lacks intrinsic DNA binding activity and is recruited to specific loci by transcription factors, including Mxd[1], PLZF and HBP12,3. These factors ‘plug-in’ to the Sin3A complex via interactions mediated by four conserved paired amphipathic helix (PAH) domains (PAH1-4). The repressor protein, Mxd[1], for example, will bind tightly to Sin3A-PAH2, but not PAH1, 3 or 46,7 This modular array of protein docking domains enables Sin3A to form a hub of transcription factors and chromatin modifying activities in the cell. The 11-amino acid Tet1-SID is necessary and sufficient for the interaction with Sin3A and are absolutely required for Tet[1] to repress transcription in cells
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