Histone chaperones FACT and Spt6 prevent histone variants from turning into histone deviants.

Histone variants are specialized histones which replace their canonical counterparts in specific nucleosomes. Together with histone post-translational modifications and DNA methylation, they contribute to the epigenome. Histone variants are incorporated at specific locations by the concerted action of histone chaperones and ATP-dependent chromatin remodelers. Recent studies have shown that the histone chaperone FACT plays key roles in preventing pervasive incorporation of two histone variants: H2A.Z and CenH3/CENP-A. In addition, Spt6, another histone chaperone, was also shown to be important for appropriate H2A.Z localization. FACT and Spt6 are both associated with elongating RNA polymerase II. Based on these two examples, we propose that the establishment and maintenance of histone variant genomic distributions depend on a transcription-coupled epigenome editing (or surveillance) function of histone chaperones.

Jeronimo C ，Robert F 《-》

Histone Recycling by FACT and Spt6 during Transcription Prevents the Scrambling of Histone Modifications.

Genomic DNA is framed by additional layers of information, referred to as the epigenome. Epigenomic marks such as DNA methylation, histone modifications, and histone variants are concentrated on specific genomic sites, where they can both instruct and reflect gene expression. How this information is maintained, notably in the face of transcription, is not completely understood. Specifically, the extent to which modified histones themselves are retained through RNA polymerase II passage is unclear. Here, we show that several histone modifications are mislocalized when the transcription-coupled histone chaperones FACT or Spt6 are disrupted in Saccharomyces cerevisiae. In the absence of functional FACT or Spt6, transcription generates nucleosome loss, which is partially compensated for by the increased activity of non-transcription-coupled histone chaperones. The random incorporation of transcription-evicted modified histones scrambles epigenomic information. Our work highlights the importance of local recycling of modified histones by FACT and Spt6 during transcription in the maintenance of the epigenomic landscape.

Jeronimo C ，Poitras C ，Robert F 《Cell Reports》

The Histone Chaperones FACT and Spt6 Restrict H2A.Z from Intragenic Locations.

H2A.Z is a highly conserved histone variant involved in several key nuclear processes. It is incorporated into promoters by SWR-C-related chromatin remodeling complexes, but whether it is also actively excluded from non-promoter regions is not clear. Here we provide genomic and biochemical evidence that the RNA polymerase II (RNA Pol II) elongation-associated histone chaperones FACT and Spt6 both contribute to restricting H2A.Z from intragenic regions. In the absence of FACT or Spt6, the lack of efficient nucleosome reassembly coupled to pervasive incorporation of H2A.Z by mislocalized SWR-C alters chromatin composition and contributes to cryptic initiation. Therefore, chaperone-mediated H2A.Z confinement is crucial for restricting the chromatin signature of gene promoters that otherwise may license or promote cryptic transcription.

Jeronimo C ，Watanabe S ，Kaplan CD ，Peterson CL ，Robert F ... -  《-》

The Abundant Histone Chaperones Spt6 and FACT Collaborate to Assemble, Inspect, and Maintain Chromatin Structure in Saccharomyces cerevisiae.

Saccharomyces cerevisiae Spt6 protein is a conserved chromatin factor with several distinct functional domains, including a natively unstructured 30-residue N-terminal region that binds competitively with Spn1 or nucleosomes. To uncover physiological roles of these interactions, we isolated histone mutations that suppress defects caused by weakening Spt6:Spn1 binding with the spt6-F249K mutation. The strongest suppressor was H2A-N39K, which perturbs the point of contact between the two H2A-H2B dimers in an assembled nucleosome. Substantial suppression also was observed when the H2A-H2B interface with H3-H4 was altered, and many members of this class of mutations also suppressed a defect in another essential histone chaperone, FACT. Spt6 is best known as an H3-H4 chaperone, but we found that it binds with similar affinity to H2A-H2B or H3-H4. Like FACT, Spt6 is therefore capable of binding each of the individual components of a nucleosome, but unlike FACT, Spt6 did not produce endonuclease-sensitive reorganized nucleosomes and did not displace H2A-H2B dimers from nucleosomes. Spt6 and FACT therefore have distinct activities, but defects can be suppressed by overlapping histone mutations. We also found that Spt6 and FACT together are nearly as abundant as nucleosomes, with ∼24,000 Spt6 molecules, ∼42,000 FACT molecules, and ∼75,000 nucleosomes per cell. Histone mutations that destabilize interfaces within nucleosomes therefore reveal multiple spatial regions that have both common and distinct roles in the functions of these two essential and abundant histone chaperones. We discuss these observations in terms of different potential roles for chaperones in both promoting the assembly of nucleosomes and monitoring their quality.

McCullough L ，Connell Z ，Petersen C ，Formosa T ... -  《-》

FACT is recruited to the +1 nucleosome of transcribed genes and spreads in a Chd1-dependent manner.

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 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 test these and other scenarios in Saccharomyces cerevisiae and find that FACT binds transcribed chromatin, not RNAPII. Through a combination of high-resolution genome-wide mapping, single-molecule tracking, and mathematical modeling, 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.

Jeronimo C ，Angel A ，Nguyen VQ ，Kim JM ，Poitras C ，Lambert E ，Collin P ，Mellor J ，Wu C ，Robert F ... -  《-》