FACT is recruited to the +1 nucleosome of transcribed genes and spreads in a Chd1-dependent manner [MNase-ChIP-Seq]

The histone chaperone FACT occupies transcribed regions where it plays prominent roles in maintaining chromatin integrity and preserving epigenetic information. How it is targeted to transcribed regions, however, remains unclear. Proposed models for how FACT finds its way to transcriptionally active chromatin include docking on the RNA polymerase II (RNAPII) C-terminal domain (CTD), recruitment by elongation factors, recognition of modified histone tails and binding partially disassembled nucleosomes. Here, we systematically tested these and other scenarios in Saccharomyces cerevisiae and found that FACT binds transcribed chromatin, not RNAPII. Through a combination of experimental and mathematical modeling evidence, we propose that FACT recognizes the +1 nucleosome, as it is partially unwrapped by the engaging RNAPII, and spreads to downstream nucleosomes aided by the chromatin remodeler Chd1. Our work clarifies how FACT interacts with genes, suggests a processive mechanism for FACT function, and provides a framework to further dissect the molecular mechanisms of transcription-coupled histone chaperoning. Paired-end sequencing of immunoprecipitated micrococcal nuclease-treated chromatin (MNase-ChIP-seq) of both FACT subunits (Spt16 and Pob3) in wild type and chd1Δ cells, with corresponding Inputs. The data also includes MNase-ChIP-seq for Chd1 in wild type and chd1K407R cells, with corresponding Inputs. All data are from two independent biological replicates.