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Epigenetic alterations occur as organisms age, and lead to chromatin deterioration, loss of transcriptional silencing and genomic instability. Dysregulation of the epigenome has been associated with increased susceptibility to age-related disorders. In this study, we aimed to characterize the age-dependent changes of the epigenome and, in turn, to understand epigenetic processes that drive aging phenotypes. We focused on the aging-associated changes in the repressive histone marks H3K9me3 and H3K27me3 in C. elegans. We observed region-specific gain and loss of both histone marks, but the changes are more evident for H3K9me3. We further found alteration of heterochromatic boundaries in aged somatic tissues. Interestingly, we discovered that the most statistically significant changes reflected H3K9me3-marked regions that are formed during aging, and are absent in developing worms, which we termed “aging-specific repressive regions” (ASRRs). These ASRRs preferentially occur in genic regions that are marked by high levels of H3K9me2 and H3K36me2 in larval stages. Maintenance of high H3K9me2 levels in these regions have been shown to correlate with a longer lifespan. Next, we examined whether the changes in repressive histone marks lead to de-silencing of repetitive DNA elements, as reported for several other organisms. We observed increased expression of active repetitive DNA elements but not global re-activation of silent repeats in old worms, likely due to the distributed nature of repetitive elements in the C. elegans genome. Intriguingly, CELE45, a putative short interspersed nuclear element (SINE), was greatly overexpressed at old age and upon heat stress. SINEs have been suggested to regulate transcription in response to various cellular stresses in mammals. It is likely that CELE45 RNAs also play roles in stress response and aging in C. elegans. Taken together, our study revealed significant and specific age-dependent changes in repressive histone modifications and repetitive elements, providing important insights into aging biology., Author summary Heterochromatin refers to the portion of the genome that is tightly packed where genes stay silent. Heterochromatin is typically decorated by particular chemical groups called histone modifications, such as trimethylation of lysine 9 or lysine 27 on histone 3 (H3K9me3 or H3K27me3). To understand how the heterochromatin landscape may change from a "youthful" to an "aged" state, we monitored the genome-wide patterns of H3K9me3 and H3K27me3 during aging using the genetic model soil worm C. elegans. We found that while H3K27me3 remained relatively stable with age, H3K9me3 showed substantial gain and loss at specific loci in aged worms. We observed that new H3K9me3-marked heterochromatin preferentially formed in specific gene-rich regions in aged worms. Interestingly, these particular regions were marked by high levels of three other histone modifications when worms were young. This result suggested that H3K9me3 gain during aging is influenced by the gene-specific landscape of histone modifications established at young age rather than that it occurs in a stochastic manner. In summary, our study discovered reproducible and gene-specific changes in histone modifications that likely contribute to the aging phenotypes.

The transcriptional co-regulator host cell factor-1 (HCF-1) plays critical roles in promoting cell cycle progression in diverse cell types, and in maintaining self-renewal of embryonic stem cells, but its role in pancreatic β-cell function has not been investigated. Immunhistochemistry of mouse pancreas revealed nuclear expression of HCF-1 in pancreatic islets. Reducing HCF-1 expression in the INS-1 pancreatic β-cell line resulted in reduced cell proliferation, reduced glucose-stimulated insulin secretion, and reduced expression of the critical β-cell transcription factor Pdx1. HCF-1 is a known co-activator of the E2F1 transcription factor, and loss of E2F1 results in pancreatic β-cell dysfunction and reduced expression of Pdx1. Therefore we wondered whether HCF-1 might be required for E2F1 regulation of Pdx1. Chromatin immunoprecipitation experiments revealed that HCF-1 and E2F1 co-localize to the Pdx1 promoter. These results indicate that HCF-1 represents a novel transcriptional regulator required for maintaining pancreatic β-cell function.

Larvae of the nematode Caenorhabditis elegans must choose between reproductive development and dauer diapause. This decision is based on sensing of environmental inputs and dauer pheromone, a small molecule signal that serves to monitor population density. These signals are integrated via conserved neuroendocrine pathways that converge on steroidal ligands of the nuclear receptor DAF-12, a homolog of the mammalian vitamin D receptor and liver X receptor. DAF-12 acts as the main switch between gene expression programs that drive either reproductive development or dauer entry. Extensive studies in the past two decades demonstrated that biosynthesis of two bile acid-like DAF-12 ligands, named dafachronic acids (DA), controls developmental fate. In this issue of PLoS Biology, Wollam et al. showed that a conserved steroid-modifying enzyme, DHS-16, introduces a key feature in the structures of the DAF-12 ligands, closing a major gap in the DA biosynthesis pathway. The emerging picture of DA biosynthesis in C. elegans enables us to address a key question in the field: how are complex environmental signals integrated to enforce binary, organism-wide decisions on developmental fate? Schaedel et al. demonstrated that pheromone and DA serve as competing signals, and that a positive feedback loop based on regulation of DA biosynthesis ensures organism-wide commitment to reproductive development. Considering that many components of DA signaling are highly conserved, ongoing studies in C. elegans may reveal new aspects of bile acid function and lifespan regulation in mammals.

Recent findings indicate that perturbations of the mitochondrial electron transport chain (METC) can cause extended longevity in evolutionarily diverse organisms. To uncover the molecular basis of how altered METC increases lifespan in C. elegans, we performed an RNAi screen and revealed that three predicted transcription factors are specifically required for the extended longevity of mitochondrial mutants. In particular, we demonstrated that the nuclear homeobox protein CEH-23 uniquely mediates the longevity but not the slow development, reduced brood size, or resistance to oxidative stress associated with mitochondrial mutations. Furthermore, we showed that ceh-23 expression levels are responsive to altered METC, and enforced overexpression of ceh-23 is sufficient to extend lifespan in wild-type background. Our data point to mitochondria-to-nucleus communications to be key for longevity determination and highlight CEH-23 as a novel longevity factor capable of responding to mitochondrial perturbations. These findings provide a new paradigm for how mitochondria impact aging and age-dependent diseases., Author Summary Mitochondria have long been associated with aging and age-related diseases. Recent research has shown that a slight dampening of mitochondrial function can dramatically increase the lifespan of a wide range of organisms, suggesting that a similar mechanism likely operates in humans. The molecular basis of this observation is largely unknown, however. Uncovering the genes that allow altered mitochondrial function to impact longevity will give us important new insights into how mitochondria affect the aging process and will pave the way for future therapeutic developments aiming to improve healthy aging and to treat age-related diseases. Here, we used an RNAi screen in the genetic model organism C. elegans, a nematode worm, to uncover how altered mitochondrial function can modulate longevity. We found that in order for mitochondria to affect lifespan, they must communicate with several unique transcription factors in the nucleus. Notably, we discovered that the putative homeobox transcription factor CEH-23, which has not previously been implicated in longevity determination, is able to respond to changes in mitochondrial function and in turn causes an extension in lifespan.

The conserved DAF-16/FOXO transcription factors and SIR-2.1/SIRT1 deacetylases are critical for diverse biological processes, particularly longevity and stress response; and complex regulation of DAF-16/FOXO by SIR-2.1/SIRT1 is central to appropriate biological outcomes. Caenorhabditis elegans Host Cell Factor 1 (HCF-1) is a longevity determinant previously shown to act as a co-repressor of DAF-16. We report here that HCF-1 represents an integral player in the regulatory loop linking SIR-2.1/SIRT1 and DAF-16/FOXO in both worms and mammals. Genetic analyses showed that hcf-1 acts downstream of sir-2.1 to influence lifespan and oxidative stress response in C. elegans. Gene expression profiling revealed a striking 80% overlap between the DAF-16 target genes responsive to hcf-1 mutation and sir-2.1 overexpression. Subsequent GO-term analyses of HCF-1 and SIR-2.1-coregulated DAF-16 targets suggested that HCF-1 and SIR-2.1 together regulate specific aspects of DAF-16-mediated transcription particularly important for aging and stress responses. Analogous to its role in regulating DAF-16/SIR-2.1 target genes in C. elegans, the mammalian HCF-1 also repressed the expression of several FOXO/SIRT1 target genes. Protein–protein association studies demonstrated that SIR-2.1/SIRT1 and HCF-1 form protein complexes in worms and mammalian cells, highlighting the conservation of their regulatory relationship. Our findings uncover a conserved interaction between the key longevity determinants SIR-2.1/SIRT1 and HCF-1, and they provide new insights into the complex regulation of FOXO proteins., Author Summary The nematode C. elegans has been instrumental in identifying and characterizing genetic components that influence aging. Studies in worms have been successfully extended to complex mammalian organisms allowing for the identification of genetic factors that impact longevity in mammals. DAF-16/FOXO transcription factors are among the best characterized longevity factors, and their increased activity leads to a longer lifespan and improved stress resistance in many organisms. Elucidating how the activities of DAF-16/FOXO are regulated will provide new insights into the basic biology of aging and will aid future therapeutic developments aiming to improve healthy aging and alleviate age-related diseases in humans. We utilized both C. elegans and mammalian cell culture systems to dissect the functional and molecular interactions between two important DAF-16 regulators, HCF-1 and SIR-2.1/SIRT1. We demonstrated that HCF-1 and SIR-2.1/SIRT1 physically associate and antagonize each other to properly regulate DAF-16/FOXO-mediated expression of genes important for longevity and stress response. We further showed that the functional relationships among these three proteins are conserved in mammals. Our work implicates HCF-1 as an important player in the regulation of FOXO by SIRT1, and thereby a potential longevity determinant in humans, and prompts further characterization of HCF-1's functions in aging and age-related pathologies.

The transcription factor DAF-16/forkhead box O (FOXO) is a critical longevity determinant in diverse organisms, however the molecular basis of how its transcriptional activity is regulated remains largely unknown. We report that the Caenorhabditis elegans homolog of host cell factor 1 (HCF-1) represents a new longevity modulator and functions as a negative regulator of DAF-16. In C. elegans, hcf-1 inactivation caused a daf-16-dependent lifespan extension of up to 40% and heightened resistance to specific stress stimuli. HCF-1 showed ubiquitous nuclear localization and physically associated with DAF-16. Furthermore, loss of hcf-1 resulted in elevated DAF-16 recruitment to the promoters of its target genes and altered expression of a subset of DAF-16-regulated genes. We propose that HCF-1 modulates C. elegans longevity and stress response by forming a complex with DAF-16 and limiting a fraction of DAF-16 from accessing its target gene promoters, and thereby regulates DAF-16-mediated transcription of selective target genes. As HCF-1 is highly conserved, our findings have important implications for aging and FOXO regulation in mammals., Author Summary One of the key molecules that modulate longevity in evolutionarily diverse organisms is the transcription factor DAF-16/FOXO. Despite its importance in aging and other biological processes, how DAF-16/FOXO activity is regulated in the nucleus is largely unknown. We report a new player important for aging modulation, the nematode homolog of host cell factor 1 (HCF-1), and show that it functions as a negative regulator of DAF-16. In worms, HCF-1 inactivation extends lifespan up to 40% and increases resistance to specific stress stimuli. To affect lifespan and stress response, HCF-1 requires the activity of DAF-16. We show that the HCF-1 protein is expressed in the nucleus and partners with DAF-16 in worms. Furthermore, we demonstrate that loss of HCF-1 results in elevated levels of DAF-16 at the promoters of its target genes and altered expression of a subset of DAF-16-regulated genes. We propose that HCF-1 modulates longevity and stress response by binding to DAF-16 and preventing the transcription factor from accessing its target gene promoters, thereby regulating the expression of DAF-16 target genes. As HCF-1 is highly conserved, our findings have important implications for aging and FOXO regulation in humans., Caenorhabditis elegans HCF-1 is a new longevity factor that functions to antagonize the transcriptional activities of DAF-16/FOXO by forming a complex with DAF-16/FOXO and preventing a fraction of DAF-16/FOXO from accessing its target gene promoters.