Biochemical Characterization of the Yeast Set1 Complex

Abstract

Previous studies have shown that H2B ubiquitylation-dependent H3K4 methylation is highly enriched at the actively transcribing region of chromatin. Although cell based studies proposed that the Set1 complex (Set1C), a sole enzyme complex responsible for H3K4 methylation in yeast, interacts with RNA polymerase II and the Paf1 complex, detailed physical interaction between Set1C and transcription machinery has not been clearly understood. To detail molecular basis of recruitment of Set1C to the actively transcribing genes and underlying mechanism of H2B ubiquitylation-dependent H3K4 methylation, we deployed several biochemical analyses using reconstituted/purified Set1C. Here, we found that Set1C can directly interact with RNA, suggesting a direct physical link of Set1C to nascent RNA emerging from the transcribing RNA polymerase II. In addition, an in vitro histone methyltransferase assay employing purified Set1C and a recombinant chromatin template allowed us to find out a specific region within Spp1 required for H2B ubiquitylation-dependent H3K4 methylation.