No abstract

SummaryDietary restriction (DR) has the remarkable ability to extend lifespan and healthspan. A variety of DR regimens have been described in species ranging from yeast to mammals. However, whether different DR regimens extend lifespan via universal, distinct, or overlapping pathways is still an open question. Here we examine the genetic pathways that mediate longevity by different DR regimens in Caenorhabditis elegans . We have previously shown that the low-energy sensing AMP-activated protein kinase AMPK/ aak-2 and the Forkhead transcription factor FoxO/ daf-16 are necessary for longevity induced by a DR regimen that we developed (sDR). Here we find that AMPK and FoxO are necessary for longevity induced by another DR regimen, but are dispensable for the lifespan extension induced by two different DR methods. Intriguingly, AMPK is also necessary for the lifespan extension elicited by resveratrol, a natural polyphenol that mimics some aspects of DR. Conversely, we test if genes previously reported to mediate longevity by a variety of DR methods are necessary for sDR-induced longevity. Although clk-1 , a gene involved in ubiquinone biosynthesis, is also required for sDR-induced lifespan extension, we find that four other genes ( sir-2.1 , FoxA/ pha-4 , skn-1 , and hsf-1 ) are all dispensable for longevity induced by sDR. Consistent with the observation that different DR methods extend lifespan by mostly independent genetic mechanisms, we find that the effects on lifespan of two different DR regimens are additive. Understanding the genetic network by which different DR regimens extend lifespan has important implications for harnessing the full benefits of DR on lifespan and healthspan.

Summary The ability to maintain quiescence is critical for the long-term maintenance of a functional stem cell pool. To date, the epigenetic and transcriptional characteristics of quiescent stem cells and how they change with age remain largely unknown. In this study, we explore the chromatin features of adult skeletal muscle stem cells, or satellite cells (SCs), which reside predominantly in a quiescent state in fully developed limb muscles of both young and aged mice. Using a ChIP-seq approach to obtain global epigenetic profiles of quiescent SCs (QSCs), we show that QSCs possess a permissive chromatin state in which few genes are epigenetically repressed by Polycomb group (PcG)-mediated histone 3 lysine 27 trimethylation (H3K27me3), and a large number of genes encoding regulators that specify nonmyogenic lineages are demarcated by bivalent domains at their transcription start sites (TSSs). By comparing epigenetic profiles of QSCs from young and old mice, we also provide direct evidence that, with age, epigenetic changes accumulate and may lead to a functional decline in quiescent stem cells. These findings highlight the importance of chromatin mapping in understanding unique features of stem cell identity and stem cell aging.

The plasticity of aging suggests that longevity may be controlled epigenetically by specific alterations in chromatin state. The link between chromatin and aging has mostly focused on histone deacetylation by the Sir2 family1,2, but less is known about the role of other histone modifications in longevity. Histone methylation plays a crucial role during development and in maintaining stem cell pluripotency in mammals3. Regulators of histone methylation have been associated with aging in worms4,5,6,7 and flies8, but characterization of their role and mechanism of action has been limited. Here we identify the ASH-2 trithorax complex9, which trimethylates histone H3 at lysine 4 (H3K4), as a regulator of lifespan in C. elegans in a directed RNAi screen in fertile worms. Deficiencies in members of the ASH-2 complex–ASH-2 itself, WDR-5, and the H3K4 methyltransferase SET-2 extend worm lifespan. Conversely, the H3K4 demethylase RBR-2 is required for normal lifespan, consistent with the idea that an excess of H3K4 trimethylation–a mark associated with active chromatin–is detrimental for longevity. Lifespan extension induced by ASH-2 complex deficiency requires the presence of an intact adult germline and the continuous production of mature eggs. ASH-2 and RBR-2 act in the germline, at least in part, to regulate lifespan and to control a set of genes involved in lifespan determination. These results suggest that the longevity of the soma is regulated by an H3K4 methyltransferase/demethylase complex acting in the C. elegans germline.

During Drosophila gastrulation, two waves of constriction occur in the apical ventral cells, leading to mesoderm invagination. The first constriction wave is a stochastic process mediated by the constriction of 40% of randomly positioned mesodermal cells and is controlled by the transcription factor Snail. The second constriction wave immediately follows and involves the other 60% of the mesodermal cells. The second wave is controlled by the transcription factor Twist and requires the secreted protein Fog. Complete mesoderm invagination requires redistribution of the motor protein Myosin II to the apical side of the constricting cells. We show that apical redistribution of Myosin II and mesoderm invagination, both of which are impaired in snail homozygous mutants that are defective in both constriction waves, are rescued by local mechanical deformation of the mesoderm with a micromanipulated needle. Mechanical deformation appears to promote Fog-dependent signaling by inhibiting Fog endocytosis. We propose that the mechanical tissue deformation that occurs during the Snail-dependent stochastic phase is necessary for the Fog-dependent signaling that mediates the second collective constriction wave.

The modulation of developmental biochemical pathways by mechanical cues is an emerging feature of animal development, but its evolutionary origins have not been explored. Here we show that a common mechanosensitive pathway involving β-catenin specifies early mesodermal identity at gastrulation in zebrafish and Drosophila. Mechanical strains developed by zebrafish epiboly and Drosophila mesoderm invagination trigger the phosphorylation of β-catenin–tyrosine-667. This leads to the release of β-catenin into the cytoplasm and nucleus, where it triggers and maintains, respectively, the expression of zebrafish brachyury orthologue notail and of Drosophila Twist, both crucial transcription factors for early mesoderm identity. The role of the β-catenin mechanosensitive pathway in mesoderm identity has been conserved over the large evolutionary distance separating zebrafish and Drosophila. This suggests mesoderm mechanical induction dating back to at least the last bilaterian common ancestor more than 570 million years ago, the period during which mesoderm is thought to have emerged.

FoxO transcription factors play a conserved role in longevity and act as tissue-specific tumor suppressors in mammals. Several nodes of interaction have been identified between FoxO transcription factors and p53, a major tumor suppressor in humans and mice. However, the extent and importance of the functional interaction between FoxO and p53 have not been fully explored. Here, we show that p53 transactivates the expression of FoxO3, one of the four mammalian FoxO genes, in response to DNA damaging agents in both mouse embryonic fibroblasts and in thymocytes. We show that p53 transactivates FoxO3 in cells by binding to a site in the second intron of the FoxO3 gene, a genomic region recently found to be associated with extreme longevity in humans. While FoxO3 is not necessary for p53-dependent cell cycle arrest, FoxO3 appears to modulate p53-dependent apoptosis. We also find that FoxO3 loss does not interact with p53 loss for tumor development in vivo, although the tumor spectrum of p53 deficient mice may be affected by FoxO3 loss. Our findings indicate that FoxO3 is a p53 target gene, and suggest that FoxO3 and p53 are part of a regulatory transcriptional network that may play an important role during aging and cancer.

We investigated the modulation of critical transcriptional steps of C2C12 myoblast/osteoblast transdifferentiation triggered by the bone morphogenetic protein 2 (BMP2) signaling protein, in response to epigenetic inhibition of the endocytotic internalization of exogenous BMP2. BMP2 endocytosis was inhibited chemically with polyethylene glycol-50 (PEG-Chol) and cyclodextrin and mechanically by mild hyposmotic treatment. BMP2-dependent nuclear translocation of the mother against Dpp (Smad1) transcription factor was ten times faster if BMP2 endocytosis was inhibited. Smad1-dependent expression of the JunB gene, the first transcriptional step in myoblast dedifferentiation, was increased by a factor of three to four. JunB-dependent levels of myogenin repression, one of the critical markers of terminal myoblastic differentiation, was amplified by a factor of three. Smad1-dependent levels of alkaline phosphatase expression, one of the C2C12 osteoblast differentiation markers, were 3.5 to 5 times higher. The same behavior was observed for osteopontin, the other C2C12 osteoblast differentiation marker. These results suggest that the cell genome could "sense" tissue mechanical deformations by mechanical inhibition of signaling protein endocytosis, thereby translating mechanical strains into transcription events involved in cell differentiation.