The mTOR pathway is a major signaling hub that coordinates growth and metabolism in response to the nutritional state of the organism. Its dysregulation has been implicated in a broad spectrum of common disease states, including cancer, diabetes, and aging. Interestingly, two interventions that suppress mTOR signaling - rapamycin and caloric restriction (a reduction in caloric intake to a fraction of ad libitum levels - have both been robustly linked to diminished tumor growth and augmented lifespan in mice. Little is known, however, about the molecular mechanisms responsible for these physiological and pathophysiological phenomena. Organismal aging is thought to be due, in part, to the progressive functional decline in stem cell compartments. We have chosen to focus on the intestine, and, in particular, on understanding the effects of caloric restriction (CR) and mTOR inhibition on intestinal stem cells (ISCs) and their niche. We find that CR strongly augments the self-renewal capacity of ISCs through a non-cell autonomous mechanism requiring the inhibition of mTORC1 and the upregulation of Bst1 in Paneth cells. Surprisingly, Paneth cells transitioned from CR conditions to nutrient-rich conditions maintained the CR phenotypes for at least 3 days, suggesting that the CR state itself is not simply the consequence of an acute signaling event. The goal of this project is to understand how CR, via mTORC1 inhibition, upregulates Bst1 expression, a necessary and sufficient event for the CR phenotype. One hypothesis we are entertaining is that mTORC1 inhibition stably modulates Bst1 expression by remodeling the epigenome. We are using computational and experimental methods in tandem to identify candidate genes that may mediate the signaling cascade between mTORC1 and the Bst1 gene. We will then systematically test the necessity and sufficiency of each candidate gene, first in test tube-based organoid co-culture and then in mice engineered to afford genetic manipulations specifically in the Paneth cell population. A clearer understanding of this pathway may shed light on new ways of mimicking the beneficial CR state. To determine the mechanisms by which long-term CR regulates gene expression, I propose the following specific aims: I: Elucidate the molecular mechanisms that modulate Bst1 mRNA levels in response to CR or mTORC1 inhibition by rapamycin. II: Determine the role(s) of the regulatory factor(s) identified in Specifc Aim I on the in vivo effects of CR and rapamycin on ISC self-renewal.
This project will shed light on how the chronic inhibition of mTORC1, a master regulator of growth and viability, alters the gene expression profile of intestinal Paneth niche cells. Paneth niche cells transduce signals from the nutrient environment to neighboring intestinal stem cells, thereby augmenting the latter's capacity for self- renewal. Elucidation of these mechanisms may lend insights on how mTOR controls the aging process and may further reveal novel targets to combat aging.