Acetylation of histone H3 on lysine 56 occurs during mitotic and meiotic S phase in fungal species. This acetylation blocks a direct electrostatic interaction between histone H3 and nucleosomal DNA, and the absence of this modification is associated with extreme sensitivity to genotoxic agents. We show here that H3-K56 acetylation is catalyzed when Rtt109, a protein that lacks significant homology to known acetyltransferases, forms an active complex with either of two histone binding proteins, Asf1 or Vps75. Rtt109 binds to both these cofactors, but not to histones alone, forming enzyme complexes with kinetic parameters similar to those of known histone acetyltransferase (HAT) enzymes. Therefore, H3-K56 acetylation is catalyzed by a previously unknown mechanism that requires a complex of two proteins: Rtt109 and a histone chaperone. Additionally, these complexes are functionally distinct, with the Rtt109/Asf1 complex, but not the Rtt109/Vps75 complex, being critical for resistance to genotoxic agents.

Histone acetylation affects many nuclear processes including transcription, chromatin assembly, and DNA damage repair. Acetylation of histone H3 lysine 56 (H3 K56ac) in budding yeast occurs during mitotic S phase and persists during DNA damage repair. Here, we show that H3 K56ac is also present during premeiotic S phase and is conserved in fission yeast. Furthermore, the H3 K56ac modification is not observed in the absence of the histone chaperone Asf1. asf1⌬ and H3 K56R mutants exhibit similar sensitivity to DNA damaging agents. Mutational analysis of Asf1 demonstrates that DNA damage sensitivity correlates with (i) decreased levels of H3 K56ac and (ii) a region implicated in histone binding. In contrast, multiple asf1 mutants that are resistant to DNA damage display WT levels of K56ac. These data suggest that maintenance of H3 K56 acetylation is a primary contribution of Asf1 to genome stability in yeast.chromatin assembly ͉ H3 K56ac ͉ DNA damage

Histone acetylation and nucleosome remodeling regulate DNA damage repair, replication and transcription. Rtt109, a recently discovered histone acetyltransferase (HAT) from Saccharomyces cerevisiae, functions with the histone chaperone Asf1 to acetylate lysine K56 on histone H3 (H3K56), a modification associated with newly synthesized histones. In vitro analysis of Rtt109 revealed that Vps75, a Nap1 family histone chaperone, could also stimulate Rtt109-dependent acetylation of H3K56. However, the molecular function of the Rtt109-Vps75 complex remains elusive. Here we have probed the molecular functions of Vps75 and the Rtt109-Vps75 complex through biochemical, structural and genetic means. We find that Vps75 stimulates the kcat of histone acetylation by ∼100-fold relative to Rtt109 alone and enhances acetylation of K9 in the H3 histone tail. Consistent with the In vitro evidence, cells lacking Vps75 showed a substantial reduction (60%) in H3K9 acetylation during S phase. X-ray structural, biochemical and genetic analyses of Vps75 indicate a unique, structurally dynamic Nap1-like fold that suggests a potential mechanism of Vps75-dependent activation of Rtt109. Together, these data provide evidence for a multifunctional HAT-chaperone complex that acetylates histone H3 and deposits H3-H4 onto DNA, linking histone modification and nucleosome assembly.

Cell differentiation is mediated by lineage-determining transcription factors. We show that chromodomain helicase DNA-binding domain 2 (Chd2), a SNF2 chromatin remodelling enzyme family member, interacts with MyoD and myogenic gene regulatory sequences to specifically mark these loci via deposition of the histone variant H3.3 prior to cell differentiation. Directed and genome-wide analysis of endogenous H3.3 incorporation demonstrates that knockdown of Chd2 prevents H3.3 deposition at differentiation-dependent, but not housekeeping, genes and inhibits myogenic gene activation. The data indicate that MyoD determines cell fate and facilitates differentiation-dependent gene expression through Chd2-dependent deposition of H3.3 at myogenic loci prior to differentiation.

Genes specifically expressed in neurons contain members with extended long introns. Longer genes present a problem with respect to fulfilment of gene length transcription, and evidence suggests that dysregulation of long genes is a mechanism underlying neurodegenerative and psychiatric disorders. Here, we report the discovery that RNA-binding protein Sfpq is a critical factor for maintaining transcriptional elongation of long genes. We demonstrate that Sfpq co-transcriptionally binds to long introns and is required for sustaining long-gene transcription by RNA polymerase II through mediating the interaction of cyclin-dependent kinase 9 with the elongation complex. Phenotypically, Sfpq disruption caused neuronal apoptosis in developing mouse brains. Expression analysis of Sfpq-regulated genes revealed specific downregulation of developmentally essential neuronal genes longer than 100 kb in Sfpq-disrupted brains; those genes are enriched in associations with neurodegenerative and psychiatric diseases. The identified molecular machinery yields directions for targeted investigations of the association between long-gene transcriptopathy and neuronal diseases.