Mechanisms Through Which Lithocholic Acid and Caloric Restriction Delay Yeast Chronological Aging

The objective of studies described in this thesis was to investigate mechanisms through which lithocholic acid (LCA) and caloric restriction (CR) delay chronological aging of the budding yeast Saccharomyces cerevisiae. My overall hypothesis was that each of these aging-delaying interventions slows down yeast chronological aging by targeting different cell biological processes. Findings presented here support this hypothesis. Indeed, I found that LCA delays the chronological mode of S. cerevisiae aging by altering the lipid composition of mitochondrial membranes. I demonstrated that these LCA-driven alterations in mitochondrial membrane lipidome elicit distinct changes in the abundance of many mitochondrial proteins, thereby enabling the establishment of a geroprotective pattern of mitochondrial functionality. These findings provided the first comprehensive evidence that mitochondrial lipidome plays an essential role in defining longevity of chronologically aging yeast. I found that CR slows yeast chronological aging through a different mechanism. This previously unknown mechanism links the chronological mode of S. cerevisiae aging to cell cycle regulation and controlled cell differentiation. The key aspects of my hypothesis-driven investigation of these two different mechanisms of aging delay are outlined below. The Titorenko laboratory has previously found that exogenously added LCA delays yeast chronological aging. The laboratory has demonstrated that LCA enters the yeast cell, is sorted to iv mitochondria, resides in both mitochondrial membranes, changes the relative concentrations of different membrane phospholipids, triggers changes in the concentrations of many mitochondrial proteins, and alters some key aspects of mitochondrial functionality. Based on these observations, I hypothesized that the LCA-driven changes in mitochondrial lipidome may have a causal role in the remodeling of mitochondrial proteome, which may in turn alter the functional state of mitochondria to create a mitochondrial pattern that delays yeast chronological aging. To test this hypothesis, I investigated how the ups1Δ, ups2Δ and psd1Δ mutations that eliminate enzymes involved in mitochondrial phospholipid metabolism influence the mitochondrial lipidome. I also assessed how these mutations affect the mitochondrial proteome, influence mitochondrial functionality and impinge on the efficiency of aging delay by LCA. My findings provide evidence that 1) LCA initially creates a distinct pro-longevity pattern of mitochondrial lipidome by proportionally decreasing phosphatidylethanolamine and cardiolipin concentrations to maintain equimolar concentrations of these phospholipids, and by increasing phosphatidic acid concentration; 2) this pattern of mitochondrial lipidome allows to establish a specific, aging-delaying pattern of mitochondrial proteome; and 3) this pattern of mitochondrial proteome plays an essential role in creating a distinctive, geroprotective pattern of mitochondrial functionality. A yeast culture grown in a nutrient-rich medium initially containing 2% glucose is not limited in calorie supply. When yeast cells cultured in this medium consume glucose, they undergo cell cycle arrest at a checkpoint in late G1 and differentiate into quiescent and non-quiescent cell populations. Studies of such differentiation have provided insights into mechanisms of yeast chronological aging under conditions of excessive calorie intake. CR is an aging-delaying dietary intervention. I hypothesized that CR may slow down yeast chronological aging by eliciting specific changes in cell cycle regulation and controlled cell differentiation. To test this hypothesis, I v investigated how CR influences the differentiation of chronologically aging yeast cultures into quiescent and non-quiescent cells, and how it affects their properties. I found that CR extends yeast chronological lifespan via a mechanism linking cellular aging to cell cycle regulation, maintenance of quiescence, entry into a non-quiescent state and survival in this state. My findings suggest that CR delays yeast chronological aging by causing specific changes in the following: 1) a checkpoint in G1 for cell cycle arrest and entry into a quiescent state; 2) a growth phase in which high-density quiescent cells are committed to become low-density quiescent cells; 3) the differentiation of low-density quiescent cells into low-density non-quiescent cells; and 4) the conversion of high-density quiescent cells into high-density non-quiescent cells.