Structure And Function Of Human Paf1 Complex

The conserved Pafl complex is a multifunctional complex. It is comprised of the Pafl, Ctr9, Leol, Cdc73, and Rtfl subunits in yeast. Ski8is also the component of human Pafl complex. In the transcription elongation, Pafl complex is associated with the promoter and coding sequences (CDS) of active genes. Cdc73and Rtfl are both required for the association of Pafl complex with chromation. In addition, interactions between Pafl complex and FACT, Spt4-Spt5and TFIIS have also been shown to modulate recruitment of the Pafl complex to chromation.Besides the role of Pafl complex in transcription elongation, it also plays critical role in promoting transcription-coupled histone modifications, including H2B monoubiquitylation and histone H3methylation. Pafl complex engages in the recruitment of Brel/Rad6ubiquitin conjugase-ligase complex, which can catalyse H2B monoubiquitylation. Monoubiquitylation of H2B is a prerequisite for the methylation of H3K4and H3K79by the Setl/COMPASS and Dotl methyltransferases, respectively. In addition, various studies have revealed function of Pafl complex in other diverse processes, including participation in RNA3’-end formation during posttranscriptional events, involvement in stem cell identity maintenance and mammalian embryonic development, and immunological responses to disease states and cancer progression.Thus far, it remains elusive that assembly process of Pafl complex and the molecular mechanism how Pafl complex carried out funtions. In this thesis, we confirmed the interaction between the Pafl and Leol subunits in vitro and mapped each minimal corresponding binding region in Pafl and Leol. We resolved the structure of Pafl/Leol subcomplex within the human Pafl complex at2.5A resolution. Anylasis of structure reveals that antiparallel beta-sheet between the Pafl and Leol was the main interaction, which mediates the formation of a tightly associated Pafl/Led heterodimer. Deletion mutagenesis and co-immunoprecipitation experiments indicate that Leol subunit binds to Pafl complex through Pafl and that the Ctr9is the key scaffold subunit which plays key role in Pafl complex assembly. Additionally, we demonstrate that the heterodimer of Pafl/Leol subcomplex is necessary to specifically recognize histone H3, the histone octamer, and nucleosome in vitro. Our results uncover the assembly process of Pafl complex and substrate recognition during Pafl complex-coordinated histone modifications.