It has been suggested that the beneficial, anti-cancer functions of certain tumor suppressor mechanisms such as senescence may untowardly contribute to aging. In this model, genes that prevent cancer also compromise organismal fitness by limiting tissue regeneration and repair. Several lines of evidence suggest the tumor suppressor p16INK4a, a potent mediator of senescence, is a strong candidate for such a molecular effector of aging. First, mice lacking p16INK4a are tumor prone but otherwise developmentally normal; suggesting p16INK4a is majorin vivo function is to abrogate neoplasia. Additionally, we and others have shown that significant p16INK4a expression is not detected in young rodents and humans, yet is widely expressed in older mammals. As the only known function of p16INK4a is to induce cell cycle arrest, this increased p16 INK4a expression may play a significant role in the impaired tissue regeneration and stem cell function characteristic of aging. In support of this hypothesis, our recent data from the study of p16INK4a-/- animals demonstrates that mice lacking p16 INK4a possess a resistance to certain aging phenotypes. For example, p16INK4a-/- mice demonstrate attenuation of the physiologic age-associated decline of function of T-cells and pancreatic islets. In particular, this latter phenotype produces enhanced glucose tolerance and resistance to an islet cell toxin in adult p16INK4a -/- mice.
In specific aim I, we propose to characterize rigorously the expression of p16INK4a in several tissue compartments in young and aged mice through real-time PCR, immunohistochemical and RNA in situ approaches. This data will inform specific aim II, in which we plan to study further several aging phenotypes (e.g. longevity, glucose tolerance, hematopoetic stem cell function, immune function, bone density, wound healing, graying, etc.) in mice lacking p 16INK4a compared to normal littermate controls. For these experiments, we have backcrossed p16INK4a deficient animals to a homogeneous and more tumor-resistant genetic background (C57Bl/6), allowing for the study of p16INK4a -/- mice at older ages.
In specific aim III, we will seek to provide genetic confirmation that p16INK4a expression contributes to aging through the generation of low copy-number BAC transgenic mice that over-express p16INK4a under the control of its endogenous promoter. The aging phenotypes of these animals will be characterized in a way analogous to specific aim II. Through these approaches, we will examine the in vivo tissue and age-specific effects of p16INK4a expression, and delineate its contribution to a variety of mammalian aging phenotypes.
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