The histone chaperone Spt6 controls chromatin structure through its conserved N-terminal domain [MNase-Seq]
The disassembly and reassembly of nucleosomes by histone chaperones is an essential activity during eukaryotic transcription elongation. This highly conserved process maintains chromatin integrity by transiently removing nucleosomes as barriers and then restoring them in the wake of transcription. While transcription elongation requires multiple histone chaperones, there is little understanding of how most of them function and why so many are required. Here, we show that the histone chaperone Spt6 acts through its acidic, intrinsically disordered N-terminal domain (NTD) to bind histones and control chromatin structure. The Spt6 NTD is essential for viability and its histone binding activity is conserved between yeast and humans. The essential nature of the Spt6 NTD can be bypassed by changes in another histone chaperone, FACT, revealing a close functional connection between the two. Our results have led to a mechanistic model for dynamic cooperation between multiple histone chaperones during transcription elongation. Overall design: Micrococcal nuclease sequencing (MNase-seq) analysis to determine whether the Spt6 NTD is required for normal chromatin structure. Since the Spt6?2-238 protein does not support viability in Saccharomyces cerevisiae, we used a strain in which the genome-encoded Spt6 could be rapidly depleted using an auxin-inducible degron while expressing either no (empty vector), wild-type (Spt6), or mutant (Spt6?2-238) versions of Spt6 from a plasmid. We then performed MNase digestion on cells grown in non-depleted (DMSO) and depleted (IAA) conditions. Experiments were performed in biological triplicate (rep1, rep2, and rep3).