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International audience; Histone modifications are key epigenetic regulators that control chromatin structure and gene transcription, thereby impacting on various important cellular phenotypes. Over the past decade, a growing number of studies have indicated that changes in various histone modifications have a significant influence on the aging process. Furthermore, it has been revealed that the abundance and localization of histone modifications are responsive to various environmental stimuli, such as diet, which can also affect gene expression and lifespan. This supports the notion that histone modifications can serve as a main cellular platform for signal integration. Hence, in this review we focus on the role of histone modifications during aging, report the data indicating that diet affects histone modification levels and explore the idea that histone modifications may function as an intersection through which diet regulates lifespan. A greater understanding of the epigenetic mechanisms that link environmental signals to longevity may provide new strategies for therapeutic intervention in age-related diseases and for promoting healthy aging.

Transcriptional memory of gene expression enables adaptation to repeated stimuli across many organisms. However, the regulation and heritability of transcriptional memory in single cells and through divisions remains poorly understood. Here, we combined microfluidics with single-cell live imaging to monitor Saccharomyces cerevisiae galactokinase 1 (GAL1) expression over multiple generations. By applying pedigree analysis, we dissected and quantified the maintenance and inheritance of transcriptional reinduction memory in individual cells through multiple divisions. We systematically screened for loss- and gain-of-memory knockouts to identify memory regulators in thousands of single cells. We identified new loss-of-memory mutants, which affect memory inheritance into progeny. We also unveiled a gain-of-memory mutant, elp6Δ, and suggest that this new phenotype can be mediated through decreased histone occupancy at the GAL1 promoter. Our work uncovers principles of maintenance and inheritance of gene expression states and their regulators at the single-cell level. 78 5 915 925.e7

International audience; It is becoming increasingly evident that aging is controlled by both genetic and epigenetic factors. Histone modifying enzymes and their modifications comprise one of the main components of epigenetic mechanisms which have been directly linked to lifespan regulation in many organisms. Studies in diverse species have highlighted changes in the distribution or abundance of certain histone marks during the lifespan of a cell or an organism, leading to alterations in gene expression. In some cases, this aging-dependent patterning of histone modifications affects the expression of key longevity genes while, in other cases, they drive large-scale transcriptome changes that eventually contribute to functional decline and other hallmarks of aging. Since epigenetic factors, including histone modifications, are malleable to environmental signals, it was reasonably hypothesized that the epigenome could act as a platform through which external signals control lifespan via gene regulation. However, evidence connecting an environmental stimulus to a specific histone modification and the subsequent alteration of a particular gene expression program influencing lifespan was lacking. Using the genetically tractable eukaryote Saccharomyces cerevisiae we have recently reported that histone H4 N-terminal acetylation (N-acH4), a modification catalyzed by the N-terminal acetyltransferase Nat4, responds to calorie restriction (CR) in order to enable the expression of genes which directly delay aging

Changes in histone modifications are an attractive model through which environmental signals, such as diet, could be integrated in the cell for regulating its lifespan. However, evidence linking dietary interventions with specific alterations in histone modifications that subsequently affect lifespan remains elusive. We show here that deletion of histone N-alpha-terminal acetyltransferase Nat4 and loss of its associated H4 N-terminal acetylation (N-acH4) extend yeast replicative lifespan. Notably, nat4Δ-induced longevity is epistatic to the effects of calorie restriction (CR). Consistent with this, (i) Nat4 expression is downregulated and the levels of N-acH4 within chromatin are reduced upon CR, (ii) constitutive expression of Nat4 and maintenance of N-acH4 levels reduces the extension of lifespan mediated by CR, and (iii) transcriptome analysis indicates that nat4Δ largely mimics the effects of CR, especially in the induction of stress-response genes. We further show that nicotinamidase Pnc1, which is typically upregulated under CR, is required for nat4Δ-mediated longevity. Collectively, these findings establish histone N-acH4 as a regulator of cellular lifespan that links CR to increased stress resistance and longevity. © 2016 The Authors. Published under the terms of the CC BY 4.0 license 17 1829 1843 Cited By :2

Background Transcriptome studies have revealed that many eukaryotic genomes are pervasively transcribed producing numerous long non-coding RNAs (lncRNAs). However, only a few lncRNAs have been ascribed a cellular role thus far, with most regulating the expression of adjacent genes. Even less lncRNAs have been annotated as essential hence implying that the majority may be functionally redundant. Therefore, the function of lncRNAs could be illuminated through systematic analysis of their synthetic genetic interactions (GIs). Results Here, we employ synthetic genetic array (SGA) in Saccharomyces cerevisiae to identify GIs between long intergenic non-coding RNAs (lincRNAs) and protein-coding genes. We first validate this approach by demonstrating that the telomerase RNA TLC1 displays a GI network that corresponds to its well-described function in telomere length maintenance. We subsequently performed SGA screens on a set of uncharacterised lincRNAs and uncover their connection to diverse cellular processes. One of these lincRNAs, SUT457, exhibits a GI profile associating it to telomere organisation and we consistently demonstrate that SUT457 is required for telomeric overhang homeostasis through an Exo1-dependent pathway. Furthermore, the GI profile of SUT457 is distinct from that of its neighbouring genes suggesting a function independent to its genomic location. Accordingly, we show that ectopic expression of this lincRNA suppresses telomeric overhang accumulation in sut457Δ cells assigning a trans-acting role for SUT457 in telomere biology. Conclusions Overall, our work proposes that systematic application of this genetic approach could determine the functional significance of individual lncRNAs in yeast and other complex organisms. Electronic supplementary material The online version of this article (doi:10.1186/s12915-016-0325-7) contains supplementary material, which is available to authorized users.

Protein N-terminal acetylation is an abundant post-translational modification in eukaryotes implicated in various fundamental cellular and biochemical processes. This modification is catalysed by evolutionarily conserved N-terminal acetyltransferases (NATs) whose deregulation has been linked to cancer development and thus, are emerging as useful diagnostic and therapeutic targets. Naa40 is a highly selective NAT that acetylates the amino-termini of histones H4 and H2A and acts as a sensor of cell growth in yeast. In the present study, we examine the role of Naa40 in cancer cell survival. We demonstrate that depletion of Naa40 in HCT116 and HT-29 colorectal cancer cells decreases cell survival by enhancing apoptosis, whereas Naa40 reduction in non-cancerous mouse embryonic fibroblasts has no effect on cell viability. Specifically, Naa40 knockdown in colon cancer cells activates the mitochondrial caspase-9-mediated apoptotic cascade. Consistent with this, we show that caspase-9 activation is required for the induced apoptosis because treatment of cells with an irreversible caspase-9 inhibitor impedes apoptosis when Naa40 is depleted. Furthermore, the effect of Naa40-depletion on cell-death is mediated through a p53-independent mechanism since p53-null HCT116 cells still undergo apoptosis upon reduction of the acetyltransferase. Altogether, these findings reveal an anti-apoptotic role for Naa40 and exhibit its potential as a therapeutic target in colorectal cancers. Electronic supplementary material The online version of this article (doi:10.1007/s10495-015-1207-0) contains supplementary material, which is available to authorized users.

Induction of gene expression in yeast and human cells involves changes in the histone modifications associated with promoters. Here we identify a histone H3 endopeptidase activity in Saccharomyces cerevisiae that may regulate these events. The endopeptidase cleaves H3 after Ala21, generating a histone that lacks the first 21 residues and shows a preference for H3 tails carrying repressive modifications. In vivo, the H3 N terminus is clipped, specifically within the promoters of genes following the induction of transcription. H3 clipping precedes the process of histone eviction seen when genes become fully active. A truncated H3 product is not generated in yeast carrying a mutation of the endopeptidase recognition site (H3 Q19A L20A) and gene induction is defective in these cells. These findings identify clipping of H3 tails as a previously uncharacterized modification of promoter-bound nucleosomes, which may result in the localized clearing of repressive signals during the induction of gene expression. © 2009 Nature America, Inc. All rights reserved. 16 17 22 Cited By :70

Polycomb group (PcG) complexes 2 and 3 are involved in transcriptional silencing. These complexes contain a histone lysine methyltransferase (HKMT) activity that targets different lysine residues on histones H1 or H3 in vitro. However, it is not known if these histones are methylation targets in vivo because the human PRC2/3 complexes have not been studied in the context of a natural promoter because of the lack of known target genes. Here we report the use of RNA expression arrays and CpG-island DNA arrays to identify and characterize human PRC2/3 target genes. Using oligonucleotide arrays, we first identified a cohort of genes whose expression changes upon siRNA-mediated removal of Suz12, a core component of PRC2/3, from colon cancer cells. To determine which of the putative target genes are directly bound by Suz12 and to precisely map the binding of Suz12 to those promoters, we combined a high-resolution chromatin immunoprecipitation (ChIP) analysis with custom oligonucleotide promoter arrays. We next identified additional putative Suz12 target genes by using ChIP coupled to CpG-island microarrays. We showed that HKMT-Ezh2 and Eed, two other components of the PRC2/3 complexes, colocalize to the target promoters with Suz12. Importantly, recruitment of Suz12, Ezh2 and Eed to target promoters coincides with methylation of histone H3 on Lys 27. 18 1592 1605 Cited By :364

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Gene ontology (GO) terms enriched by the negative genetic interactions of SUTs and of their flanking genes. Heatmap of GO terms enriched with Pâ â ¤â 0.05 (generated using the Fisher exact test) by genes classified as negative genetic interactions for the indicated SUTs and for their corresponding adjacent protein-coding genes. The negative genetic interactions of the flanking protein-coding genes were extracted from the Drygin database ( http://drygin.ccbr.utoronto.ca/ ). (PDF 95 kb)

Lack of SUT457 accelerates senescence in tlc1Δ cells. a, b Senescence assays performed using liquid cultures of the indicated isogenic wild-type and mutant strains. The strains were generated through tetrad dissection of the heterozygous diploid double mutant SUT457/sut457ΔTLC1/tlc1Δ. Each plot represents one of three independent tetrads examined (see also Fig. 3e). (PDF 65 kb)

Deletion of EXO1 rescues the growth arrest of gbp2Î sut457Î mutant cells. Isogenic strains of the indicated genotype (top panels) were streaked repeatedly on solid rich medium and their growth at passage 5 is shown. (PDF 68 kb)

Detection of SUT457 RNA. Total RNA was isolated from wild-type and sut457Î cells and analysed by northern blotting using a probe (Additional file 14: Table S7) against the SUT457 sequence (Additional file 6: Table S5). A band of 345Â bp corresponding to the size of SUT457 is detected only in wild-type cells (left panel). A probe against actin (Additional file 14: Table S7) was used as a loading control (right panel). (PDF 26 kb)

Overlap between TLC1 and EST1 negative genetic interactions. a Venn diagram showing the overlap (P = 0.001 generated using the hypergeometric test) between genes whose deletions result in negative genetic interactions with TLC1 in this study and with EST1 as reported in [43]. GO terms enriched by common negative genetic interactions with P

Methylation at particular residues on histone tails has been associated with various functions and, in the case of dimethylated histone H3 arginine 2 (H3R2), cross-talk with methylation of a nearby lysine has been shown to be linked to transcriptional repression. Budding yeast monomethylated H3R2 is now shown to be associated with active loci and involved in activation of meiotic genes upon induction of sporulation. Dimethylation of histone H3 Arg2 (H3R2me2) maintains transcriptional silencing by inhibiting Set1 mediated trimethylation of H3K4. Here we demonstrate that Arg2 is also monomethylated (H3R2me1) in yeast but that its functional characteristics are distinct from H3R2me2: (i) H3R2me1 does not inhibit histone H3 Lys4 (H3K4) methylation; (ii) it is present throughout the coding region of genes; and (iii) it correlates with active transcription. Collectively, these results indicate that different H3R2 methylation states have defined roles in gene expression.

Epigenetic modifications, including those occurring on DNA and on histone proteins, control gene expression by establishing and maintaining different chromatin states. In recent years, it has become apparent that epigenetic modifications do not function alone, but work together in various combinations, and crossregulate each other in a manner that diversifies their functional states. Arginine methylation is one of the numerous PTMs (post-translational modifications) occurring on histones, catalysed by a family of PRMTs (protein arginine methyltransferases). This modification is involved in the regulation of the epigenome largely by controlling the recruitment of effector molecules to chromatin. Histone arginine methylation associates with both active and repressed chromatin states depending on the residue involved and the configuration of the deposited methyl groups. The present review focuses on the increasing number of cross-talks between histone arginine methylation and other epigenetic modifications, and describe how these cross-talks influence factor binding to regulate transcription. Furthermore, we present models of general cross-talk mechanisms that emerge from the examples of histone arginine methylation and allude to various techniques that help decipher the interplay among epigenetic modifications. © 2013 Biochemical Society. 41 751 759 Cited By :24

Post-translational modifications of histones play a key role in DNA-based processes, like transcription, by modulating chromatin structure. N-terminal acetylation is unique among the numerous histone modifications because it is deposited on the N-alpha amino group of the first residue instead of the side-chain of amino acids. The function of this modification and its interplay with other internal histone marks has not been previously addressed. Here, we identified N-terminal acetylation of H4 (N-acH4) as a novel regulator of arginine methylation and chromatin silencing in Saccharomyces cerevisiae. Lack of the H4 N-alpha acetyltransferase (Nat4) activity results specifically in increased deposition of asymmetric dimethylation of histone H4 arginine 3 (H4R3me2a) and in enhanced ribosomal-DNA silencing. Consistent with this, H4 N-terminal acetylation impairs the activity of the Hmt1 methyltransferase towards H4R3 in vitro. Furthermore, combinatorial loss of N-acH4 with internal histone acetylation at lysines 5, 8 and 12 has a synergistic induction of H4R3me2a deposition and rDNA silencing that leads to a severe growth defect. This defect is completely rescued by mutating arginine 3 to lysine (H4R3K), suggesting that abnormal deposition of a single histone modification, H4R3me2a, can impact on cell growth. Notably, the cross-talk between N-acH4 and H4R3me2a, which regulates rDNA silencing, is induced under calorie restriction conditions. Collectively, these findings unveil a molecular and biological function for H4 N-terminal acetylation, identify its interplay with internal histone modifications, and provide general mechanistic implications for N-alpha-terminal acetylation, one of the most common protein modifications in eukaryotes., Author Summary The genome of eukaryotic cells is packaged into nucleosomes consisting of an octamer of histone proteins that is wrapped around by DNA. Histone proteins are often modified with chemical groups that can influence the arrangement of nucleosomes and thereby affect DNA-based processes like transcription. Histone N-terminal acetylation, which comprises the addition of a chemical group at the tip of the histone tail, is an abundant modification whose function is unknown. In this work, we show that N-terminal acetylation of histone H4 can strongly inhibit the occurrence of a neighboring modification, namely dimethylation at the third arginine. To do this, N-terminal acetylation cooperates with other internal lysine acetylation marks. We find that the communication amongst these histone modifications is necessary for controlling the expression of ribosomal RNA genes that are required for protein synthesis and cell growth. Our experiments show that in the absence of both N-terminal acetylation and lysine acetylation there is a strong increase in H4 arginine 3 dimethylation levels leading to cell lethality. This growth defect can be rescued by a point mutation on H4 that blocks methylation at position 3. Together, our results unveil a molecular and biological function for the previously uncharacterized N-terminal acetylation of histones.

Post-translational modifications (PTMs) of histones have been implicated in cellular processes such as transcription, replication and DNA repair. These processes normally involve dynamic changes in chromatin structure and DNA accessibility. Most of the PTMs reported so far map on the histone tails and essentially affect chromatin structure indirectly by recruiting effector proteins. A recent study by Schneider and colleagues published in Cell (1) has uncovered the function of H3K122 acetylation found within the histone globular domain and specifically positioned on the DNA-bound surface of the nucleosome. Their findings demonstrate a direct effect of histone PTMs on chromatin dynamics, and propose that modifications located in different parts of the nucleosome employ distinct regulatory mechanisms. 4 343 348 Cited By :4

Regulation of histone methylation is critical for proper gene expression and chromosome function. Suppressor of Zeste 12 (SUZ12) is a requisite member of the EED/EZH2 histone methyltransferase complexes, and is required for full activity of these complexes in vitro. In mammals and flies, SUZ12/Su(z)12 is necessary for trimethylation of histone H3 on lysine 27 (H3K27me3) on facultative heterochromatin. However, Su(z)12 is unique among Polycomb Group Proteins in that Su(z)12 mutant flies exhibit gross defects in position effect variegation, suggesting a role for Su(z)12 in constitutive heterochromatin formation. We investigated the role of Suz12 in constitutive heterochromatin and discovered that Suz12 is required for histone H3 lysine 9 tri-methylation (H3K9me3) in differentiated but not undifferentiated mouse embryonic stem cells. Knockdown of SUZ12 in human cells caused a reduction in H3K27me3 and H3K9me3, and altered the distribution of HP1α. In contrast, EZH2 knockdown caused loss of H3K27me3 but not H3K9me3, indicating that SUZ12 regulates H3-K9 methylation in an EZH2-independent fashion. This work uncovers a role for SUZ12 in H3-K9 methylation. © 2007 Springer. 15 299 314 Cited By :26

It is well-established that deregulation of the transcriptional activity of many different genes has been causatively linked to human diseases. In cancer, altered patterns of gene expression are often the result of the inappropriate expression of a specific transcriptional activator or repressor. Functional studies of cancer-specific transcription factors have relied upon the study of candidate target genes. More recently, gene expression profiling using DNA microarrays that contain tens of thousands of cDNAs corresponding to human mRNAs has allowed for a large-scale identification of genes that respond to increased or decreased levels of a particular transcription factor. However, such experiments do not distinguish direct versus indirect target genes. Coupling chromatin immunoprecipitation to microarrays that contain genomic regions (ChIP-chip) has provided investigators with the ability to identify, in a high-throughput manner, promoters directly bound by specific transcription factors. Clearly, knowledge gained from both types of arrays provides complementary Information, allowing greater confidence that a transcription factor regulates a particular gene. In this review, we focus on Polycomb group (PcG) complexes as an example of transcriptional regulators that are implicated In various cellular processes but about which very little is known concerning their target gene specificity. We provide examples of how both expression arrays and ChIP-chip microarray-based assays can be used to identify target genes of a particular PcG complex and suggest improvements in the application of array technology for faster and more comprehensive identification of directly regulated target genes. 229 705 721 Cited By :53

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