NMR Dissection of the Detailed Mechanism for Antibiotic Binding to Asite RNA

Abstract

protein complexes containing histone deacetylase (HDAC) activities play key roles in regulating eukaryotic gene transcription by altering 'the histone code' and modulating chromatin structure dynamics. The evolutionarilyconserved, HDAC-associated,~0.5 MDa Sin3S/Rpd3S complex is implicated in repressing transcription from cryptic start sites in the intragenic regions of transcriptionally-active genes and in limiting DNA damage due to genotoxic stress. Unlike the related~2 MDa Sin3L/Rpd3L complex, which is targeted to promoter regions of genes through interactions with sequence-specific DNA-binding repressors, the smaller Sin3S/Rpd3S complex is targeted to intragenic regions through interactions with H3 K36(me 2 /me 3 )-modified histones. The Sin3S/ Rpd3S complex comprises at least five subunits of which the MRG15 and Pf1 subunits are unique to this complex and play important roles in chromatin targeting and complex assembly. Pf1 harbors two PHD zinc fingers of unknown function and two Sin3 interaction domains -SID1 and SID2 -that interact with the PAH2 and PAH1 domains, respectively, of Sin3. Pf1 SID1 overlaps with a segment that we recently identified as being critical for interactions with MRG15 -the subunit that is thought to interact directly with H3 K36(me 2 /me 3 ). We have used solution NMR spectroscopy to investigate the network of protein-protein interactions involving Pf1 and to characterize the structure and potential functions of its PHD domains. These studies provide unexpected insights into the assembly of this corepressor complex besides affording new insights into Sin3 PAH-SID interactions and into the structure and function of PHD fingers. The results of these studies will be presented and their implications in chromatin/transcription biology discussed.