The budding yeast heterochromatic SIR complex resets upon exit from stationary phase

The budding yeast SIR complex (Silent Information Regulator) is the principal actor in heterochromatin formation, which causes epigenetically regulated gene silencing phenotypes. The maternal chromatin bound SIR complex is disassembled during replication. Consequently, if heterochromatin is to be restored on both daughter strands, the SIR complex has to be reformed on both strands to pre-replication levels. The dynamics of SIR complex maintenance and reformation during the cell-cycle and in different growth conditions are however not clear. Understanding exchange rates of SIR subunits during the cell cycle and their distribution pattern to daughter chromatids after replication has important implications for how heterochromatic states may be inherited and therefore how epigenetic states are maintained from one cellular generation to the next. We used the tag switch RITE system to measure genome wide turnover rates of the SIR subunit Sir3 before and after exit from stationary phase and show that maternal Sir3 subunits are completely replaced with newly synthesized Sir3 at subtelomeric regions during the first cell cycle after release from stationary phase. The SIR complex is therefore not "inherited" and the silenced state has to be established de novo upon exit from stationary phase. Additionally, our analysis of genome-wide transcription dynamics shows that precise Sir3 dosage is needed for the optimal up-regulation of "growth" genes during the first cell-cycle after release from stationary phase

Distinct transcriptional roles for Histone H3-K56 acetylation during the cell cycle in Yeast

Dynamic disruption and reassembly of promoter-proximal nucleosomes is a conserved hallmark of transcriptionally active chromatin. Histone H3-K56 acetylation (H3K56Ac) enhances these turnover events and promotes nucleosome assembly during S phase. Here we sequence nascent transcripts to investigate the impact of H3K56Ac on transcription throughout the yeast cell cycle. We find that H3K56Ac is a genome-wide activator of transcription. While H3K56Ac has a major impact on transcription initiation, it also appears to promote elongation and/or termination. In contrast, H3K56Ac represses promiscuous transcription that occurs immediately following replication fork passage, in this case by promoting efficient nucleosome assembly. We also detect a stepwise increase in transcription as cells transit S phase and enter G2, but this response to increased gene dosage does not require H3K56Ac. Thus, a single histone mark can exert both positive and negative impacts on transcription that are coupled to different cell cycle events

A rare natural lipid induces neuroglobin expression to prevent amyloid oligomers toxicity and retinal neurodegeneration

Most neurodegenerative diseases such as Alzheimer's disease are proteinopathies linked to the toxicity of amyloid oligomers. Treatments to delay or cure these diseases are lacking. Using budding yeast, we report that the natural lipid tripentadecanoin induces expression of the nitric oxide oxidoreductase Yhb1 to prevent the formation of protein aggregates during aging and extends replicative lifespan. In mammals, tripentadecanoin induces expression of the Yhb1 orthologue, neuroglobin, to protect neurons against amyloid toxicity. Tripentadecanoin also rescues photoreceptors in a mouse model of retinal degeneration and retinal ganglion cells in a Rhesus monkey model of optic atrophy. Together, we propose that tripentadecanoin affects p-bodies to induce neuroglobin expression and offers a potential treatment for proteinopathies and retinal neurodegeneration.Peer reviewe

Inheritance of Chromatin Proteins in Budding Yeast: metabolic gene regulators TUP1, FPR4 and Rpd3L are retained in the mother cell

Asymmetric division is a prerequisite for cellular differentiation. Phenotypic transformation during differentiation is a poorly understood epigenetic phenomenon, in which chromatin theoretically plays a role. The assumption that chromatin components segregate asymmetrically in asymmetric divisions has however not been systematically tested. We have developed a live cell imaging method to measure how 18 chromatin proteins are inherited in asymmetric divisions of budding yeast. We show that abundant and moderately abundant maternal proteins segregate stochastically and symmetrically between the two cells with the exception of Rxt3, Fpr4 and Tup1, which are retained in the mother. Mother retention seems to be the norm for low abundance proteins with the exception of Sir2 and the linker histone H1. Our in vivo analysis of chromatin protein behavior in single cells highlights general trends in protein biology during the cell cycle such as coupled protein synthesis and decay, and a correlation between half-lives and cell cycle duration

A key role for chd1 in histone h3 dynamics at the 3\u27 ends of long genes in yeast

Chd proteins are ATP-dependent chromatin remodeling enzymes implicated in biological functions from transcriptional elongation to control of pluripotency. Previous studies of the Chd1 subclass of these proteins have implicated them in diverse roles in gene expression including functions during initiation, elongation, and termination. Furthermore, some evidence has suggested a role for Chd1 in replication-independent histone exchange or assembly. Here, we examine roles of Chd1 in replication-independent dynamics of histone H3 in both Drosophila and yeast. We find evidence of a role for Chd1 in H3 dynamics in both organisms. Using genome-wide ChIP-on-chip analysis, we find that Chd1 influences histone turnover at the 5\u27 and 3\u27 ends of genes, accelerating H3 replacement at the 5\u27 ends of genes while protecting the 3\u27 ends of genes from excessive H3 turnover. Although consistent with a direct role for Chd1 in exchange, these results may indicate that Chd1 stabilizes nucleosomes perturbed by transcription. Curiously, we observe a strong effect of gene length on Chd1\u27s effects on H3 turnover. Finally, we show that Chd1 also affects histone modification patterns over genes, likely as a consequence of its effects on histone replacement. Taken together, our results emphasize a role for Chd1 in histone replacement in both budding yeast and Drosophila melanogaster, and surprisingly they show that the major effects of Chd1 on turnover occur at the 3\u27 ends of genes

Patterns and Mechanisms of Ancestral Histone Protein Inheritance in Budding Yeast

Tracking of ancestral histone proteins over multiple generations of genome replication in yeast reveals that old histones move along genes from 3′ toward 5′ over time, and that maternal histones move up to around 400 bp during genomic replication

Replication and Active Demethylation Represent Partially Overlapping Mechanisms for Erasure of H3K4me3 in Budding Yeast

Histone modifications affect DNA–templated processes ranging from transcription to genomic replication. In this study, we examine the cell cycle dynamics of the trimethylated form of histone H3 lysine 4 (H3K4me3), a mark of active chromatin that is viewed as “long-lived” and that is involved in memory during cell state inheritance in metazoans. We synchronized yeast using two different protocols, then followed H3K4me3 patterns as yeast passed through subsequent cell cycles. While most H3K4me3 patterns were conserved from one generation to the next, we found that methylation patterns induced by alpha factor or high temperature were erased within one cell cycle, during S phase. Early-replicating regions were erased before late-replicating regions, implicating replication in H3K4me3 loss. However, nearly complete H3K4me3 erasure occurred at the majority of loci even when replication was prevented, suggesting that most erasure results from an active process. Indeed, deletion of the demethylase Jhd2 slowed erasure at most loci. Together, these results indicate overlapping roles for passive dilution and active enzymatic demethylation in erasing ancestral histone methylation states in yeast

The small DNA binding domain of λ integrase is a context-sensitive modulator of recombinase functions

λ Integrase (Int) has the distinctive ability to bridge two different and well separated DNA sequences. This heterobivalent DNA binding is facilitated by accessory DNA bending proteins that bring flanking Int sites into proximity. The regulation of λ recombination has long been perceived as a structural phenomenon based upon the accessory protein-dependent Int bridges between high-affinity arm-type (bound by the small N-terminal domain) and low-affinity core-type DNA sites (bound by the large C-terminal domain). We show here that the N-terminal domain is not merely a guide for the proper positioning of Int protomers, but is also a context-sensitive modulator of recombinase functions. In full-length Int, it inhibits C-terminal domain binding and cleavage at the core sites. Surprisingly, its presence as a separate molecule stimulates the C-terminal domain functions. The inhibition in full-length Int is reversed or overcome in the presence of arm-type oligonucleotides, which form specific complexes with Int and core-type DNA. We consider how these results might influence models and experiments pertaining to the large family of heterobivalent recombinases

Mechanics of DNA Replication and Transcription Guide the Asymmetric Distribution of RNAPol2 and New Nucleosomes on Replicated Daughter Genomes

Replication of the eukaryotic genome occurs in the context of chromatin. Chromatin is commonly thought to carry epigenetic information from one generation to the next, although it is unclear how such information survives the disruptions of nucleosomal architecture occurring during genomic replication. In order to better understand the transmission of gene expression states from one cell generation to the next we have developed a method for following chromatin structure dynamics during replication-ChIP-NChAP-Chromatin Immuno-Precipitation-Nascent Chromatin Avidin Pulldown-which we used to monitor RNAPol2 and new nucleosome binding to newly-replicated daughter genomes in S. Cerevisiae. The strand specificity of our libraries allowed us to uncover the inherently asymmetric distribution of RNAPol2 and H3K56ac-a mark of new histones-on daughter chromatids after replication. Our results show a range of distributions on thousands of genes from symmetric to asymmetric with enrichment shifts from one replicated strand to the other throughout S-phase. We propose a two-step model of chromatin assembly on nascent DNAwhich provides a mechanistic framework for the regulation of asymmetric segregation of maternal histones, and discuss our model for chromatin assembly in the context of a mechanism for gene expression buffering without a direct role for H3K56ac

The asymmetric distribution of RNA polymerase II and nucleosomes on replicated daughter genomes is caused by differences in replication timing between the lagging and the leading strand

International audienceChromatin features are thought to have a role in the epigenetic transmission of transcription states from one cell generation to the next. It is unclear how chromatin structure survives disruptions caused by genomic replication or whether chromatin features are instructive of the transcription state of the underlying gene. We developed a method to monitor budding yeast replication, transcription, and chromatin maturation dynamics on each daughter genome in parallel, with which we identified clusters of secondary origins surrounding known origins. We found a difference in the timing of lagging and leading strand replication on the order of minutes at most yeast genes. We propose a model in which the majority of old histones and RNA polymerase II (RNAPII) bind to the gene copy that replicated first, while newly synthesized nucleosomes are assembled on the copy that replicated second. RNAPII enrichment then shifts to the sister copy that replicated second. The order of replication is largely determined by genic orientation: If transcription and replication are codirectional, the leading strand replicates first; if they are counterdirectional, the lagging strand replicates first. A mutation in the Mcm2 subunit of the replicative helicase Mcm2-7 that impairs Mcm2 interactions with histone H3 slows down replication forks but does not qualitatively change the asymmetry in nucleosome distribution observed in the WT. We propose that active transcription states are inherited simultaneously and independently of their underlying chromatin states through the recycling of the transcription machinery and old histones, respectively. Transcription thus actively contributes to the reestablishment of the active chromatin state