Role of CBP and SATB-1 in aging, dietary restriction, and insulin-like signaling.
ABSTRACT: How dietary restriction (DR) increases lifespan and decreases disease burden are questions of major interest in biomedical research. Here we report that hypothalamic expression of CREB-binding protein (CBP) and CBP-binding partner Special AT-rich sequence binding protein 1 (SATB-1) is highly correlated with lifespan across five strains of mice, and expression of these genes decreases with age and diabetes in mice. Furthermore, in Caenorhabditis elegans, cbp-1 is induced by bacterial dilution DR (bDR) and the daf-2 mutation, and cbp-1 RNAi specifically in adults completely blocks lifespan extension by three distinct protocols of DR, partially blocks lifespan extension by the daf-2 mutation but not of cold, and blocks delay of other age-related pathologies by bDR. Inhibiting the C. elegans ortholog of SATB-1 and CBP-binding partners daf-16 and hsf-1 also attenuates lifespan extension by bDR, but not other protocols of DR. In a transgenic Abeta42 model of Alzheimer's disease, cbp-1 RNAi prevents protective effects of bDR and accelerates Abeta42-related pathology. Furthermore, consistent with the function of CBP as a histone acetyltransferase, drugs that enhance histone acetylation increase lifespan and reduce Abeta42-related pathology, protective effects completely blocked by cbp-1 RNAi. Other factors implicated in lifespan extension are also CBP-binding partners, suggesting that CBP constitutes a common factor in the modulation of lifespan and disease burden by DR and the insulin/IGF1 signaling pathway.
Project description:Dietary restriction (DR) is the most effective environmental intervention to extend lifespan in a wide range of species. However, the molecular mechanisms underlying the benefits of DR on longevity are still poorly characterized. AMP-activated protein kinase (AMPK) is activated by a decrease in energy levels, raising the possibility that AMPK might mediate lifespan extension by DR.By using a novel DR assay that we developed and validated in C. elegans, we find that AMPK is required for this DR method to extend lifespan and delay age-dependent decline. We find that AMPK exerts its effects in part via the FOXO transcription factor DAF-16. FOXO/DAF-16 is necessary for the beneficial effects of this DR method on lifespan. Expression of an active version of AMPK in worms increases stress resistance and extends longevity in a FOXO/DAF-16-dependent manner. Lastly, we find that AMPK activates FOXO/DAF-16-dependent transcription and phosphorylates FOXO/DAF-16 at previously unidentified sites, suggesting a possible direct mechanism of regulation of FOXO/DAF-16 by AMPK.Our study shows that an energy-sensing AMPK-FOXO pathway mediates the lifespan extension induced by a novel method of dietary restriction in C. elegans.
Project description:The effects of antioxidants on lifespan have been widely studied. Our previous study showed supplementation with N-acetyl-l-cysteine (NAC) extends the lifespan of <i>Caenorhabditis elegans</i>. Here we aimed to determine the lifespan-extending mechanism involved with NAC and the effect of NAC on Alzheimer's disease (AD). NAC further increased the lifespan of <i>age</i>-<i>1</i> and <i>clk</i>-<i>1</i> mutants, which have increased lifespan owing to reduced insulin/IGF-1-like signaling and mitochondrial function, respectively. There was no additional lifespan extension in <i>eat</i>-<i>2</i> background, a genetic model of dietary restriction (DR), by NAC. Gene knockdown experiments revealed that the effect of NAC is not dependent on SKN-1, a protein-sensing DR status, whereas DAF-16, a transcription factor regulating stress-responsive genes, is required for lifespan extension by NAC. NAC delayed paralysis caused by amyloid beta. Our results show that NAC mimics the effect of DR on lifespan, possibly through the induction of DAF-16 nuclear localization and may retard the incidence of AD.
Project description:Screening a library of drugs with known safety profiles in humans yielded 30 drugs that reliably protected mammalian neurons against glucose toxicity. Subsequent screening demonstrated that 6 of these 30 drugs increase lifespan in C. elegans: caffeine, ciclopirox olamine, tannic acid, acetaminophen, bacitracin, and baicalein. Every drug significantly reduced the age-dependent acceleration of mortality rate. These protective effects were blocked by RNAi inhibition of cbp-1 in adults only, which also blocks protective effects of dietary restriction. Only 2 drugs, caffeine and tannic acid, exhibited a similar dependency on DAF-16. Caffeine, tannic acid, and bacitracin also reduced pathology in a transgenic model of proteotoxicity associated with Alzheimer's disease. These results further support a key role for glucose toxicity in driving age-related pathologies and for CBP-1 in protection against age-related pathologies. These results also provide novel lead compounds with known safety profiles in human for treatment of age-related diseases, including Alzheimer's disease and diabetic complications.
Project description:Caloric/dietary restriction (CR/DR) can promote longevity and protect against age-associated disease across species. The molecular mechanisms coordinating food intake with health-promoting metabolism are thus of significant medical interest. We report that conserved Caenorhabditis elegans microRNA-80 (mir-80) is a major regulator of the DR state. mir-80 deletion confers system-wide healthy aging, including maintained cardiac-like and skeletal muscle-like function at advanced age, reduced accumulation of lipofuscin, and extended lifespan, coincident with induction of physiological features of DR. mir-80 expression is generally high under ad lib feeding and low under food limitation, with most striking food-sensitive expression changes in posterior intestine. The acetyltransferase transcription co-factor cbp-1 and interacting transcription factors daf-16/FOXO and heat shock factor-1 hsf-1 are essential for mir-80(?) benefits. Candidate miR-80 target sequences within the cbp-1 transcript may confer food-dependent regulation. Under food limitation, lowered miR-80 levels directly or indirectly increase CBP-1 protein levels to engage metabolic loops that promote DR.
Project description:Interventions that slow aging and prevent chronic disease may come from an understanding of how dietary restriction (DR) increases lifespan. Mechanisms proposed to mediate DR longevity include reduced mTOR signaling, activation of the NAD⁺ -dependent deacylases known as sirtuins, and increases in NAD⁺ that derive from higher levels of respiration. Here, we explored these hypotheses in Caenorhabditis elegans using a new liquid feeding protocol. DR lifespan extension depended upon a group of regulators that are involved in stress responses and mTOR signaling, and have been implicated in DR by some other regimens [DAF-16 (FOXO), SKN-1 (Nrf1/2/3), PHA-4 (FOXA), AAK-2 (AMPK)]. Complete DR lifespan extension required the sirtuin SIR-2.1 (SIRT1), the involvement of which in DR has been debated. The nicotinamidase PNC-1, a key NAD⁺ salvage pathway component, was largely required for DR to increase lifespan but not two healthspan indicators: movement and stress resistance. Independently of pnc-1, DR increased the proportion of respiration that is coupled to ATP production but, surprisingly, reduced overall oxygen consumption. We conclude that stress response and NAD⁺ -dependent mechanisms are each critical for DR lifespan extension, although some healthspan benefits do not require NAD⁺ salvage. Under DR conditions, NAD⁺ -dependent processes may be supported by a DR-induced shift toward oxidative metabolism rather than an increase in total respiration.
Project description:Dietary 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.
Project description:Caenorhabditis elegans SKN-1 (ortholog of mammalian Nrf1/2/3) is critical for oxidative stress resistance and promotes longevity under reduced insulin/IGF-1-like signaling (IIS), dietary restriction (DR), and normal conditions. SKN-1 inducibly activates genes involved in detoxification, protein homeostasis, and other functions in response to stress. Here we used genome-scale RNA interference (RNAi) screening to identify mechanisms that prevent inappropriate SKN-1 target gene expression under non-stressed conditions. We identified 41 genes for which knockdown leads to activation of a SKN-1 target gene (gcs-1) through skn-1-dependent or other mechanisms. These genes correspond to multiple cellular processes, including mRNA translation. Inhibition of translation is known to increase longevity and stress resistance and may be important for DR-induced lifespan extension. One model postulates that these effects derive from reduced energy needs, but various observations suggest that specific longevity pathways are involved. Here we show that translation initiation factor RNAi robustly induces SKN-1 target gene transcription and confers skn-1-dependent oxidative stress resistance. The accompanying increases in longevity are mediated largely through the activities of SKN-1 and the transcription factor DAF-16 (FOXO), which is required for longevity that derives from reduced IIS. Our results indicate that the SKN-1 detoxification gene network monitors various metabolic and regulatory processes. Interference with one of these processes, translation initiation, leads to a transcriptional response whereby SKN-1 promotes stress resistance and functions together with DAF-16 to extend lifespan. This stress response may be beneficial for coping with situations that are associated with reduced protein synthesis.
Project description:Mutations in the daf-2 gene of the conserved Insulin/Insulin-like Growth Factor (IGF-1) pathway double the lifespan of the nematode Caenorhabditis elegans. This phenotype is completely suppressed by deletion of Forkhead transcription factor daf-16. To uncover regulatory mechanisms coordinating this extension of life, we employed a quantitative proteomics strategy with daf-2 mutants in comparison with N2 and daf-16; daf-2 double mutants. This revealed a remarkable longevity-specific decrease in proteins involved in mRNA processing and transport, the translational machinery, and protein metabolism. Correspondingly, the daf-2 mutants display lower amounts of mRNA and 20S proteasome activity, despite maintaining total protein levels equal to that observed in wild types. Polyribosome profiling in the daf-2 and daf-16;daf-2 double mutants confirmed a daf-16-dependent reduction in overall translation, a phenotype reminiscent of Dietary Restriction-mediated longevity, which was independent of germline activity. RNA interference (RNAi)-mediated knockdown of proteins identified by our approach resulted in modified C. elegans lifespan confirming the importance of these processes in Insulin/IGF-1-mediated longevity. Together, the results demonstrate a role for the metabolism of proteins in the Insulin/IGF-1-mediated extension of life.
Project description:Preferably, lifespan-extending therapies should work when applied late in life without causing undesired pathologies. Reducing insulin/insulin-like growth factor (IGF)-1 signaling (IIS) increases lifespan across species, but the effects of reduced IIS interventions in extreme geriatric ages remains unknown. Using the nematode <i>Caenorhabditis elegans</i>, we engineered the conditional depletion of the DAF-2/insulin/IGF-1 transmembrane receptor using an auxin-inducible degradation (AID) system. This allowed for the temporal and spatial reduction in DAF-2 protein levels at time points after which interventions such as RNAi become ineffective. Using this system, we found that AID-mediated depletion of DAF-2 protein surpasses the longevity of <i>daf-2</i> mutants. Depletion of DAF-2 during early adulthood resulted in multiple adverse phenotypes, including growth retardation, germline shrinkage, egg retention, and reduced brood size. By contrast, AID-mediated depletion of DAF-2 post-reproduction, or specifically in the intestine in early adulthood, resulted in an extension of lifespan without these deleterious effects. Strikingly, at geriatric ages, when 75% of the population had died, AID-mediated depletion of DAF-2 protein resulted in a doubling in lifespan. Thus, we provide a proof-of-concept that even close to the end of an individual's lifespan, it is possible to slow aging and promote longevity.
Project description:Dietary restriction (DR) is the most effective and reproducible intervention to extend lifespan in divergent species1. In mammals, two regimens of DR, intermittent fasting (IF) and caloric restriction (CR), have proven to extend lifespan and reduce the incidence of age-related disorders2. An important characteristic of IF is that it can increase lifespan, even when there is little or no overall decrease in calorie intake2. The molecular mechanisms underlying IF-induced longevity, however, remain largely unknown. Here we establish an IF regimen that effectively extends the lifespan of Caenorhabditis elegans, and show that a nutrient-related signalling molecule, the low molecular weight GTPase Cel-Rheb, has a dual role in lifespan regulation; Cel-Rheb is required for the IF-induced longevity, whereas inhibition of Cel-Rheb mimics the CR effects. We also show that Cel-Rheb exerts its effects in part via the insulin/IGF-like signalling effector DAF-16 in IF, and that Cel-Rheb is required for fasting-induced nuclear translocation of DAF-16. We find that HSP-12.6, a DAF-16 target, functions to mediate the IF-induced longevity. Furthermore, our analyses demonstrate that most of fasting-induced upregulated genes require Cel-Rheb function for their induction, and that Cel-Rheb/Cel-TOR signalling is required for the fasting-induced downregulation of an insulin-like peptide, INS-7. These findings identify the essential role of signalling via Cel-Rheb in IF-induced longevity and gene expression changes, and suggest a molecular link between the IF-induced longevity and the insulin/IGF-like signalling pathway. Experiment Overall Design: We examined fasting-induced changes of the gene expression profiles in Caenorhabditis elegans. We performed the genome-wide analysis by using Affymetrix GeneChip oligonucleotide microarrays, and examined the effect of downregulation of Cel-Rheb and Cel-TOR by RNAi on the expression profiles. Five independent experiments were performed with wild type N2. Synchronized worms under six conditions (control-fed, control-fasting, Rheb RNAi-fed, Rheb RNAi-fasting, TOR RNAi-fed, and TOR RNAi-fasting) were collected and frozen with liquid nitrogen at day 4 of adulthood. Total RNA was extracted with Sepasol(R)-RNA â Super (Nacalai tesque), and purified with RNeasy Mini Kit (Qiagen), according to manufactureâs instructions. Synthesis of cDNA, in vitro transcription and biotin labelling cRNA, and hybridization to the C. elegans Genome Array (Affymetrix) were performed according to Affymetrix protocols. Hybridized arrays were scanned using an Affymetrix GeneChip Scanner. Scanned chip images were analyzed with GeneSpring GX 7.3.1 (Agilent Technologies).