Elevated histone expression promotes life span extension.
ABSTRACT: Changes to the chromatin structure accompany aging, but the molecular mechanisms underlying aging and the accompanying changes to the chromatin are unclear. Here, we report a mechanism whereby altering chromatin structure regulates life span. We show that normal aging is accompanied by a profound loss of histone proteins from the genome. Indeed, yeast lacking the histone chaperone Asf1 or acetylation of histone H3 on lysine 56 are short lived, and this appears to be at least partly due to their having decreased histone levels. Conversely, increasing the histone supply by inactivation of the histone information regulator (Hir) complex or overexpression of histones dramatically extends life span via a pathway that is distinct from previously known pathways of life span extension. This study indicates that maintenance of the fundamental chromatin structure is critical for slowing down the aging process and reveals that increasing the histone supply extends life span.
Project description:Aging is the main risk factor for many prevalent diseases. However, the molecular mechanisms regulating aging at the cellular level are largely unknown. Using single cell yeast as a model organism, we found that reducing yeast histone proteins accelerates chronological aging and increasing histone supply extends chronological life span. We sought to identify pathways that regulate chronological life span by controlling intracellular histone levels. Thus, we screened the histone H3/H4 mutant library to uncover histone residues and posttranslational modifications that regulate histone gene expression. We discovered 15 substitution mutations with reduced histone proteins and 5 mutations with increased histone proteins. Among these mutations, we found Set1 complex-catalyzed H3K4me3 promotes histone gene transcription and maintains normal chronological life span. Unlike the canonical functions of H3K4me3 in gene expression, H3K4me3 facilitates histone gene transcription by acting as a boundary to restrict the spread of the repressive HIR/Asf1/Rtt106 complex from histone gene promoters. Collectively, our study identified a novel mechanism by which H3K4me3 antagonizes the HIR/Asf1/Rtt106 repressor complex to promote histone gene expression and extend chronological life span.
Project description:The orderly deposition of histones onto DNA is mediated by conserved assembly complexes, including chromatin assembly factor-1 (CAF-1) and the Hir proteins . CAF-1 and the Hir proteins operate in distinct but functionally overlapping histone deposition pathways in vivo . The Hir proteins and CAF-1 share a common partner, the highly conserved histone H3/H4 binding protein Asf1, which binds the middle subunit of CAF-1 as well as to Hir proteins . Asf1 binds to newly synthesized histones H3/H4 , and this complex stimulates histone deposition by CAF-1 . In yeast, Asf1 is required for the contribution of the Hir proteins to gene silencing . Here, we demonstrate that Hir1, Hir2, Hir3, and Hpc2 comprise the HIR complex, which copurifies with the histone deposition protein Asf1. Together, the HIR complex and Asf1 deposit histones onto DNA in a replication-independent manner. Histone deposition by the HIR complex and Asf1 is impaired by a mutation in Asf1 that inhibits HIR binding. These data indicate that the HIR complex and Asf1 proteins function together as a conserved eukaryotic pathway for histone replacement throughout the cell cycle.
Project description:Transcription by RNA polymerase II is accompanied by dynamic changes in chromatin, including the eviction/deposition of nucleosomes or the covalent modification of histone subunits. This study examined the role of the histone H3/H4 chaperones, Asf1 and HIR, in histone mobility during transcription, with particular focus on the histone exchange pathway, using a dual histone expression system. The results showed that the exchange of H3/H4 normally occurs during transcription by the histone chaperones. Both Asf1 and HIR are important for histone deposition but have a different effect on histone exchange. While Asf1 mediated incorporation of external H3/H4 and renewal of pre-existing histones, HIR opposed it. The balance of two opposing activities might be an important mechanism for determining current chromatin states.
Project description:Efficient supply of new histones during DNA replication is critical to restore chromatin organization and maintain genome function. The histone chaperone anti-silencing function 1 (Asf1) serves a key function in providing H3.1-H4 to CAF-1 for replication-coupled nucleosome assembly. We identify Codanin-1 as a novel interaction partner of Asf1 regulating S-phase histone supply. Mutations in Codanin-1 can cause congenital dyserythropoietic anaemia type I (CDAI), characterized by chromatin abnormalities in bone marrow erythroblasts. Codanin-1 is part of a cytosolic Asf1-H3.1-H4-Importin-4 complex and binds directly to Asf1 via a conserved B-domain, implying a mutually exclusive interaction with the chaperones CAF-1 and HIRA. Codanin-1 depletion accelerates the rate of DNA replication and increases the level of chromatin-bound Asf1, suggesting that Codanin-1 guards a limiting step in chromatin replication. Consistently, ectopic Codanin-1 expression arrests S-phase progression by sequestering Asf1 in the cytoplasm, blocking histone delivery. We propose that Codanin-1 acts as a negative regulator of Asf1 function in chromatin assembly. This function is compromised by two CDAI mutations that impair complex formation with Asf1, providing insight into the molecular basis for CDAI disease.
Project description:While histone chaperones have been intensely studied, the roles of components of the Hir-Asf1 histone chaperone complex such as Hir3p and Hpc2p are poorly understood. Using sensitive protein sequence profile analyses we investigated the evolution of these proteins and showed that Hir3p and Hpc2p have a much wider phyletic pattern than was previously known. We established the animal histone-deacetylase-complex-interacting proteins, CAIN/CABIN, to be orthologs of Hir3p. They contain a conserved core of around 30 TPR-like bi-helical repeats that are likely to form a super-helical scaffold. We identified a conserved domain, the HUN domain, in all Hpc2p homologs, including animal ubinuclein/yemanuclein and the recently discovered vertebrate cell-cycle regulator FLJ25778. The HUN domain has a characteristic pattern of conserved acidic residues based on which we predict that it is a previously unrecognized histone-tail-binding chaperone. By analyzing various high-throughput data sets, such as RNAi knock-downs, genetic and protein interaction maps and cell-cycle-specific gene expression data, we present evidence that Hpc2p homologs might be deployed in specific processes of chromatin dynamics relating to cell-cycle progression in vertebrates and schizogony in Plasmodium. Beyond the conserved HUN domain these proteins show extensive divergence patterns in different eukaryotic lineages. Hence, we propose that Hpc2p homologs are probably involved in recruitment of the ancient conserved histone-loading Hir-Asf1 complex to different lineage-specific chromatin reorganization processes.
Project description:In Saccharomyces cerevisiae, the histone chaperone Rtt106 binds newly synthesized histone proteins and mediates their delivery into chromatin during transcription, replication, and silencing. Rtt106 is also recruited to histone gene regulatory regions by the HIR histone chaperone complex to ensure S-phase-specific expression. Here we showed that this Rtt106:HIR complex included Asf1 and histone proteins. Mutations in Rtt106 that reduced histone binding reduced Rtt106 enrichment at histone genes, leading to their increased transcription. Deletion of the chromatin boundary element Yta7 led to increased Rtt106:H3 binding, increased Rtt106 enrichment at histone gene regulatory regions, and decreased histone gene transcription at the HTA1-HTB1 locus. These results suggested a unique regulatory mechanism in which Rtt106 sensed the level of histone proteins to maintain the proper level of histone gene transcription. The role of these histone chaperones and Yta7 differed markedly among the histone gene loci, including the two H3-H4 histone gene pairs. Defects in silencing in rtt106 mutants could be partially accounted for by Rtt106-mediated changes in histone gene repression. These studies suggested that feedback mediated by histone chaperone complexes plays a pivotal role in regulating histone gene transcription.
Project description:Epigenetic mechanisms, including histone post-translational modifications, control longevity in diverse organisms. Relatedly, loss of proper transcriptional regulation on a global scale is an emerging phenomenon of shortened life span, but the specific mechanisms linking these observations remain to be uncovered. Here, we describe a life span screen in Saccharomyces cerevisiae that is designed to identify amino acid residues of histones that regulate yeast replicative aging. Our results reveal that lack of sustained histone H3K36 methylation is commensurate with increased cryptic transcription in a subset of genes in old cells and with shorter life span. In contrast, deletion of the K36me2/3 demethylase Rph1 increases H3K36me3 within these genes, suppresses cryptic transcript initiation, and extends life span. We show that this aging phenomenon is conserved, as cryptic transcription also increases in old worms. We propose that epigenetic misregulation in aging cells leads to loss of transcriptional precision that is detrimental to life span, and, importantly, this acceleration in aging can be reversed by restoring transcriptional fidelity.
Project description:We previously reported a novel affinity purification (AP) method termed modified chromatin immunopurification (mChIP), which permits selective enrichment of DNA-bound proteins along with their associated protein network. In this study, we report a large-scale study of the protein network of 102 chromatin-related proteins from budding yeast that were analyzed by mChIP coupled to mass spectrometry. This effort resulted in the detection of 2966 high confidence protein associations with 724 distinct preys. mChIP resulted in significantly improved interaction coverage as compared with classical AP methodology for ?75% of the baits tested. Furthermore, mChIP successfully identified novel binding partners for many lower abundance transcription factors that previously failed using conventional AP methodologies. mChIP was also used to perform targeted studies, particularly of Asf1 and its associated proteins, to allow for a understanding of the physical interplay between Asf1 and two other histone chaperones, Rtt106 and the HIR complex, to be gained.
Project description:Anti-silencing function 1 (Asf1) is a highly conserved chaperone of histones H3/H4 that assembles or disassembles chromatin during transcription, replication, and repair. The structure of the globular domain of Asf1 bound to H3/H4 determined by X-ray crystallography to a resolution of 1.7 Angstroms shows how Asf1 binds the H3/H4 heterodimer, enveloping the C terminus of histone H3 and physically blocking formation of the H3/H4 heterotetramer. Unexpectedly, the C terminus of histone H4 that forms a mini-beta sheet with histone H2A in the nucleosome undergoes a major conformational change upon binding to Asf1 and adds a beta strand to the Asf1 beta sheet sandwich. Interactions with both H3 and H4 were required for Asf1 histone chaperone function in vivo and in vitro. The Asf1-H3/H4 structure suggests a "strand-capture" mechanism whereby the H4 tail acts as a lever to facilitate chromatin disassembly/assembly that may be used ubiquitously by histone chaperones.
Project description:Changes in chromatin organization occur during aging. Overexpression of histones partially alleviates these changes and promotes longevity. We report that deletion of the histone H3-H4 minor locus HHT1-HHF1 extended the replicative life span of Saccharomyces cerevisiae. This longevity effect was mediated through TOR signaling inhibition. We present evidence for evolutionarily conserved transcriptional and phenotypic responses to defects in chromatin structure, collectively termed the chromatin architectural defect (CAD) response. Promoters of the CAD response genes were sensitive to histone dosage, with HHT1-HHF1 deletion, nucleosome occupancy was reduced at these promoters allowing transcriptional activation induced by stress response transcription factors Msn2 and Gis1, both of which were required for the life-span extension of hht1-hhf1?. Therefore, we conclude that the CAD response induced by moderate chromatin defects promotes longevity.