Gene expression changes upon the mutation of individual residues of histone H3 and H4
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ABSTRACT: Nucleosome structure directly influences gene transcription. However, the function of each histone residue remains largely unknown. Here we profiled gene expression changes upon the mutation of individual residues of histone H3 and H4. Histone residues grouped by expression change similarity displayed overall structural relevance. This regulatory functional map of the core histones led to novel findings. First, the residues specific to each histone family tend to be more influential than those commonly found among different histones. Second, unlike histone acetylations, H3K4 trimethylation does not appear to be prerequisite for gene activation. Third, H3Q5 has been newly identified for its putative interactions with many chromatin regulators for transcription control. Lastly, the nucleosome lateral surface seems to play a key role through interactions with the surrounding DNA. Remarkably, we discovered a novel role for H3K56 in chromatin dynamics. The deletion of this residue, but not the alteration of acetylation states, caused a genome-wide decrease in nucleosome mobility and stabilized nucleosome positioning near transcription start and end sites. Occupying the DNA entry/exit site, H3K56 is thought to modulate nucleosome sliding along DNA. Taken together, genomics approaches such as microarray and deep sequencing prove valuable for mapping the function of histone residues. Microarray analysis was performed for 123 histone mutants and four wild-types as two reaplications of H3 and H4 of Saccharomyces ceravisiae.
Project description:Nucleosome structure directly influences gene transcription. However, the function of each histone residue remains largely unknown. Here we profiled gene expression changes upon the mutation of individual residues of histone H3 and H4. Histone residues grouped by expression change similarity displayed overall structural relevance. This regulatory functional map of the core histones led to novel findings. First, the residues specific to each histone family tend to be more influential than those commonly found among different histones. Second, unlike histone acetylations, H3K4 trimethylation does not appear to be prerequisite for gene activation. Third, H3Q5 has been newly identified for its putative interactions with many chromatin regulators for transcription control. Lastly, the nucleosome lateral surface seems to play a key role through interactions with the surrounding DNA. Remarkably, we discovered a novel role for H3K56 in chromatin dynamics. The deletion of this residue, but not the alteration of acetylation states, caused a genome-wide decrease in nucleosome mobility and stabilized nucleosome positioning near transcription start and end sites. Occupying the DNA entry/exit site, H3K56 is thought to modulate nucleosome sliding along DNA. Taken together, genomics approaches such as microarray and deep sequencing prove valuable for mapping the function of histone residues. Performing Mnase-seq for six histone mutants and two wild-types in Saccharomyces cerevisiae
Project description:Nucleosome structure directly influences gene transcription. However, the function of each histone residue remains largely unknown. Here we profiled gene expression changes upon the mutation of individual residues of histone H3 and H4. Histone residues grouped by expression change similarity displayed overall structural relevance. This regulatory functional map of the core histones led to novel findings. First, the residues specific to each histone family tend to be more influential than those commonly found among different histones. Second, unlike histone acetylations, H3K4 trimethylation does not appear to be prerequisite for gene activation. Third, H3Q5 has been newly identified for its putative interactions with many chromatin regulators for transcription control. Lastly, the nucleosome lateral surface seems to play a key role through interactions with the surrounding DNA. Remarkably, we discovered a novel role for H3K56 in chromatin dynamics. The deletion of this residue, but not the alteration of acetylation states, caused a genome-wide decrease in nucleosome mobility and stabilized nucleosome positioning near transcription start and end sites. Occupying the DNA entry/exit site, H3K56 is thought to modulate nucleosome sliding along DNA. Taken together, genomics approaches such as microarray and deep sequencing prove valuable for mapping the function of histone residues. SUBMITTER_CITATION: Global Mapping of the Regulatory Interactions of Histone Residues, Epigenomics Keystone Symposium, January 21th (2012).
Project description:Nucleosome structure directly influences gene transcription. However, the function of each histone residue remains largely unknown. Here we profiled gene expression changes upon the mutation of individual residues of histone H3 and H4. Histone residues grouped by expression change similarity displayed overall structural relevance. This regulatory functional map of the core histones led to novel findings. First, the residues specific to each histone family tend to be more influential than those commonly found among different histones. Second, unlike histone acetylations, H3K4 trimethylation does not appear to be prerequisite for gene activation. Third, H3Q5 has been newly identified for its putative interactions with many chromatin regulators for transcription control. Lastly, the nucleosome lateral surface seems to play a key role through interactions with the surrounding DNA. Remarkably, we discovered a novel role for H3K56 in chromatin dynamics. The deletion of this residue, but not the alteration of acetylation states, caused a genome-wide decrease in nucleosome mobility and stabilized nucleosome positioning near transcription start and end sites. Occupying the DNA entry/exit site, H3K56 is thought to modulate nucleosome sliding along DNA. Taken together, genomics approaches such as microarray and deep sequencing prove valuable for mapping the function of histone residues. SUBMITTER_CITATION: Global Mapping of the Regulatory Interactions of Histone Residues, Epigenomics Keystone Symposium, January 21th (2012).
Project description:Genomic profiling of (1) histone H3K56 acetylation and (2) meiotic DSBs flanked by replication protein A (RPA). The major goal was to reveal the genome-wide distribution DSBs when the H3K56 residue was mutated to alanine (H3K56A).
Project description:The S. cerevisiae Rpd3 large (Rpd3L) and small (Rpd3S) histone deacetylase (HDAC) complexes are prototypes for understanding transcriptional repression in eukaryotes [1]. The current view is that they function by deacetylating chromatin, thereby limiting accessibility of transcriptional factors to the underlying DNA. However, an Rpd3 catalytic mutant retains substantial repression capability when targeted to a promoter as a LexA fusion protein [2]. We investigated the HDAC-independent properties of the Rpd3 complexes biochemically and discovered a chaperone function, which promotes histone deposition onto DNA, and a novel activity, which prevents nucleosome eviction but not remodeling mediated by the ATP-dependent RSC complex. These HDAC-independent activities inhibit Pol II transcription on a nucleosomal template. The functions of the endogenous Rpd3 complexes can be recapitulated with recombinant Rpd3 core complex comprising Sin3, Rpd3, and Ume1. To test the hypothesis that Rpd3 contributes to chromatin stabilization in vivo, we measured histone H3 density genomewide and found that it was reduced at promoters in an Rpd3 deletion mutant but partially restored in a catalytic mutant. Importantly, the effects on H3 density are most apparent on RSC-enriched genes [3]. Our data suggest that the Rpd3 core complex could contribute to repression via a novel nucleosome stabilization function. H3 were ChIP'd from yeast strains and normalized to input.
Project description:DNA replication ensures the accurate transmission of genetic information during cell cycle. The interaction between histone methyltransferase SUV420H1 and histone variant H2A.Z plays a critical role in the licensing of early replication origins. However, the mechanism by which SUV420H1 preferentially recognizes H2A.Z-nucleosome and deposits H4 lysine 20 dimethylation (H4K20me2) on DNA replication origins remains elusive. Here, we determined the cryo-EM structures of SUV420H1 bound to H2A.Z-nucleosome or H2A-nucleosome. Our structures show that the SUV420H1 catalytic domain (CD) directly interacts with histone H4 and nucleosomal DNA, whereas a SUV420H1 arginine-rich motif (ARM) anchors to the acidic patch of the nucleosome. The N-terminal aminal acid residues of H4 (Aa 1-24) forms a lasso-shaped structure sandwiched between SUV420H1 CD and nucleosome. The lasso-shaped structure stabilizes the SUV420H1-nucleosome interaction and precisely projects the H4 K20 residue into the SUV420H1 catalytic center. Further analyses revealed a crucial role of SUV420H1 KR-loop (aminal acid residues 214-223), which spatially lies closely to H2A.Z specific residues D97/S98, in dictating the preference for H2A.Z-nucleosome. SUV420H1 K219A/R220A mutations reduced the activity of SUV420H1 for H4K20me2 modification, the preference of SUV420H1 for H2A.Z-nucleosome, and the efficiency of DNA replication initiation. Collectively, our findings elucidate how SUV420H1 preferentially recognizes H2A.Z-nucleosome to deposit H4K20me2 modification and shed light on therapeutic strategies targeting the DNA replication initiation.
Project description:Here we report V. parahaemolyticus (serotype O3:K6) infection induced histone deacetylation in intestinal epithelial cells, wherein we have shown deacetylation of H3K9, H3K56, H3K18 and H4K16 residues. Interestingly we observed concomitant downregulation of host deacetylases.
Project description:Protein methylation catalyzed by SAM-dependent methyltransferase represents a major PTM involved in important biological processes. Because methylation can occur on nitrogen, oxygen and sulfur centers and multiple methylation states exist on the nitrogen centers, methylproteome remains poorly documented. Here we present the methylation by isotope labeled SAM (MILS) strategy for a highly-confident analysis of the methylproteome of the SAM-auxotrophic Saccharomyces cerevisiae based on the online multidimensional μHPLC/MS/MS technology. We identified 117 methylated proteins, containing 182 methylation events associated with 174 methylation sites. About 90% of these methylation events were previously unknown. Our results indicated, 1) over 6% of the yeast proteome are methylated, 2) the amino acid residue preference of protein methylation follows the order Lys >> Arg > Asp > Glu ≈ Gln ≈ Asp > His > Cys, 3) the methylation state on nitrogen center is largely exclusive, and 4) the methylated proteins are located mainly in nucleus/ribosome associated with translation/transcription and DNA/RNA processing. Our dataset is the most comprehensive methylproteome known-to-date of all living organisms, and should significantly contribute to the field of protein methylation and related research.
Project description:Histone acetylation and deacetylation are among the principal mechanisms by which chromatin is regulated during transcription, DNA silencing, and DNA repair. We analyzed patterns of genetic interactions uncovered during comprehensive genome-wide analyses in yeast to probe how histone acetyltransferase (HAT) and histone deacetylase (HDAC) protein complexes interact. The genetic interaction data unveil an underappreciated role of HDACs in maintaining cellular viability, and led us to show that deacetylation of the histone variant Htz1p at lysine 14 is mediated by Hda1p. Studies of the essential nucleosome acetyltransferase of H4 (NuA4) revealed acetylation-dependent protein stabilization of Yng2p, a potential nonhistone substrate of NuA4 and Rpd3C, and led to a new functional organization model for this critical complex. We also found that DNA double-stranded breaks (DSBs) result in local recruitment of the NuA4 complex, followed by an elaborate NuA4 remodeling process concomitant with Rpd3p recruitment and histone deacetylation. These new characterizations of the HDA and NuA4 complexes demonstrate how systematic analyses of genetic interactions may help illuminate the mechanisms of intricate cellular processes. Keywords: genetic modification The 44 datasets in this Series profiled the genome-wide genetic interactions for query genes encoding either HAT and HDAC catalytic subunits or subunits of the associated protein complexes. Of the 32 query genes, 5 were essential and were tested as temperature-sensitive (ts) alleles at three or more temperatures. (ESA1 was also tested as a hypomorphic allele.) The other query genes were tested as null deletion alleles derived from the Yeast Knockout strain collection.