Spatial regulation of transcription and histone occupancy by histone chaperones FACT and Spt6

The FACT complex and Spt6 are conserved histone chaperones that are recruited to the open reading frames of transcribed genes. In this study, we provide evidence that FACT interaction with acetylated H3 tail is important for its localization to the coding sequences. Pol II CTD kinase Kin28 additionally stimulates FACT recruitment to a subset of genes. Pol II occupancies in the 5’ ends of transcribed genes are greatly reduced on depleting FACT, whereas reduced occupancies at the 3’ ends were observed upon Spt6 depletion indicating that these factors modulate Pol II progression through distinct regions of transcribed coding sequences. While FACT is largely responsible for reassembling histones, we uncover a role for Spt6 in promoting histone eviction in addition to widely-accepted role for Spt6 in histone reassembly. Consistent with their localization in the coding regions, simultaneously impairing FACT and Spt6 function severely dampens histone eviction and impairs transcription genome-wide. ChIP-chip experiments to measure Spt16 occupancies in WT and kin28as mutant, as well Rpb3 and histone H3 occupancies in undepleted or depleted cells for Spt16 and Spt6, and also in the strain lacking Spt6 tandem SH2 domain The WT and mutant strains were grown in Synthetic complete and cells were induced for Gcn4 by treating with Sulfometuron methyl for 30 minutes and processed for chromatin immunoprecipitation using antibodies again Myc, Rpb3 or histone H3.