Molecular Genetics of Ageing
Ageing is defined as a gradual decline of organismal homeostasis and of physiologic functions throughout the body, and is associated with an increased risk of age-related disease. Yet work from model genetic organisms reveals that animals possess natural defense mechanisms that help stave off ageing, increase organismal homeostasis and stimulate longevity. In particular, all species have the ability to assess environmental and physiological information, and mediate between programs geared towards growth and reproduction, versus survival and maintenance, which ultimately influence organismal life span. By using the model genetic system of C. elegans, we aim to unravel phyletically conserved features of animal life span regulation. We specifically study how endocrine and signal transduction pathways coordinate such processes, as well as how changes in metabolism, quality control mechanisms, and stress resistance impact longevity.
In C. elegans many of the conserved pathways that stimulate adult longevity were first identified for their influence on formation of a long-lived larval stage called the dauer diapause, which animals enter under harsh conditions. Endocrine pathways play a critical role in mediating this life stage choice, and include steroid hormone, insulin/IGF, and TGF-beta signaling. We have focused in particular on how steroid hormone and insulin/IGF signaling coordinate longevity. DAF-12 is a steroid receptor transcription factor most related to vertebrate Vitamin-D, Liver-X, and Farnesoid-X receptors. Like its vertebrate cousins, DAF-12 transcriptional activity is turned on by bile acid-like steroids, in this case called the dafachronic acids (DA). Genetic and molecular studies suggest a model whereby in favorable environmental conditions, cues detected by the nervous system result in activation of insulin/IGF and TGF-beta signaling, thereby stimulating DA production and DAF-12 activation. Consequently animals progress through larval development, culminating in maturation and short life. Conversely in unfavorable environments, endocrine systems are shut down, and the unliganded DAF-12 receptor specifies dauer formation, organismal arrest and longevity.
Interestingly, the DA/DAF-12 module is conserved amongst parasitic nematodes, whose infective stage is analogous to dauer. Thus the study of these pathways has not only shed light on the biology of ageing, but may have therapeutic implications for treatment of pathogenic nematodes. By using a combination of genetics and biochemistry we are elucidating the dafachronic acid biosynthetic pathways and their regulation by physiologic signaling and environmental inputs during organismal commitments. Additionally we continue to genetically dissect components of dauer formation in order to identify novel molecular pathways that promote long life.
Removal of C. elegans germline stem cells by laser microsurgery or by mutations that halt their proliferation results in a 60% extension of life span. Importantly, this type of longevity may be conserved, since removal of germline stem cells in fruitflies also extends life. Gonadal longevity depends on the steroid receptor DAF-12/FXR and its ligands, the forkhead transcription factor DAF-16/FOXO, as well as a number of other transcriptional regulators. These regulators converge on fat metabolism, autophagy, and quality control mechanisms that are required for longevity, but the underlying mechanisms and the crosstalk between pathways are not known. We are carrying out genetic and metabolite screens to unravel the molecular and cellular pathways that specify this long-lived state.
Gonadal longevity pathway
Suppression of germline stem cell proliferation or removal of the germline precursors results in a 60% increase in life span. This increase in life span depends upon the somatic gonad, which together with other tissues helps synthesize the dafachronic acids (DA). DA activates the DAF-12/FXR nuclear hormone receptor in unknown tissues, and potentiates the nuclear localization and activity of DAF-16/FOXO transcription factor. DAF-16/FOXO activity is also activated from the nervous system by the mir-71 microRNA. In response to repression of TOR kinase, DAF-16/FOXO turns on genes important for lipolysis and other processes important for longevity. PHA-4/FOXA transcription factor activates autophagy, which works in conjunction with lipases to release free fatty acids (FFA). In parallel the nuclear receptor NHR-80 turns on genes for fatty acid (FA) desaturation and oleic acid (OA) production, also important for longevity.
When pre-adult larvae are exposed to starvation conditions, they mature into reproductively arrested adults, termed the adult reproductive diapause, in which the germline ceases proliferation, and metabolic changes geared towards extended survival and longevity ensue. Upon return to favorable conditions, the germline and soma reconstitute, animals reproduce, and can live a normal life span. Little is known about the pathways connecting nutrient sensing to the molecular architecture underlying adult reproductive diapause, and we are currently dissecting the physiologic and metabolic mechanisms involved using genetic and genomic methods.
A related but different type of nutritional regimen, dietary restriction, is a decrease in food intake without malnutrition. It has beneficial effects on health and leads to an increase in life span in a wide variety of species, suggesting an evolutionary conserved process. Despite its importance, only recently are the molecular mechanisms beginning to come to light. We are carrying out genetic screens to identify the molecular pathways that specify longevity in response to reduced food intake and dietary composition.
Long-lived wild type adults in reproductive diapause.
Worms in adult reproductive diapause are able to survive up to 3 month without food. These worms don't lay eggs, and during early adult development worms in adult reproductive diapause are characterized by the presence of one or two embryos (e, day 2 of adulthood). During ageing the embryos are degraded to embryonic corpses (ec, day 10 and day 50 of adulthood). v, vulva
The progression through the life stages and the life span of a species is genetically determined, yet relatively little is known about the temporal programming of these processes. We speculate that there are close relationships between developmental timing, life stage structure, and life span, based on the observation that DAF-12/FXR works at the confluence of developmental timing and longevity pathways. C. elegans develops from embryo through four larval stages to adult. We have found that DAF-12/FXR regulates second to third larval stage transitions through a hormone regulated-microRNA switch. In response to its steroidal ligands, DAF-12/FXR directly upregulates mir-84 and mir-241, two let-7 related microRNAs, which turn off earlier stage programs to allow for later ones. We are currently studying a host of other novel transcriptional, translational, and post-translational regulators, including a highly conserved F-box protein called, DRE-1/FBXO11, which regulates larval to adult transitions through ubiquitin-mediated proteolysis. Importantly, some of the so called "heterochronic factors" have been implicated in stem cell progression from worms to mammals, and thus may give insight into stem cell biology, pluripotency, and regenerative medicine.