RSC-mediated nucleosome positioning and GRFs form barriers in promoters to limit divergent noncoding transcription

Abstract The directionality of gene promoters - the ratio of protein-coding over divergent noncoding transcription - is highly variable and regulated. How promoter directionality is controlled remains poorly understood. We show that the chromatin remodelling complex RSC and general regulatory factors (GRFs) dictate promoter directionality by attenuating divergent transcription. Depletion of RSC increased divergent noncoding transcription and decreased protein-coding transcription at promoters with strong directionality. Consistent with RSC’s role in regulating chromatin, RSC depletion impacts nucleosome occupancy upstream of the nucleosome depleted region where divergent transcription initiates, suggesting that nucleosome positioning at the 5’ border of gene promoters physically blocks the recruitment of the transcription machinery and inhibits initiation of divergent transcription. Highly directional promoters were also enriched for the binding of GRFs such as Reb1 and Abf1. Furthermore, ectopic targeting of divergent transcription initiation sites with GRFs or the dCas9 protein can suppress divergent transcription. Our data suggest that RSC-mediated nucleosome positioning and GRFs play a pervasive role in repressing divergent transcription. We propose that any DNA binding factor, when stably associated with cryptic transcription start sites, can form a barrier for repressing divergent transcription. Our study provides an explanation as to why certain promoters are more directional than others.