| The formation and storage of neutral lipids such as triglyceride (TG) and steryl ester (SE) is a ubiquitous process essential to cellular homeostasis. In addition to being a concentrated energy reservoir, TG and SE provide cells with the safe sequestration of lipid intermediates pertinent to the formation of membranes, hormones, and bile acids. The evolutionarily conserved process of neutral lipid biosynthesis occurs in the ER through either an acyl-CoA dependent or independent fashion and is mediated by acyltransferase enzymes belonging chiefly to Acyl-CoA Cholesterol Acyltransferase (ACAT), Lecithin Cholesterol Acyltransferase (LCAT), or Diacylglycerol Acyltransferase 2 (DGAT2) gene families. The ER is limited in its capacity to store NL and, therefore, these molecules are packaged within the core cytoplasmic lipid droplets (CLDs).;Increased neutral lipid levels and the associated accumulation of CLDs, is highly correlated with obesity and insulin resistance and therefore contributes to the pathogenesis of the metabolic syndrome. The mechanisms involved in the development of these lipid-related disorders coalesce around lipotoxicity, or the cellular dysfunction resulting from lipid overload. Using the yeast Saccharomyces cerevisiae, we were able to establish both a genetic and environmental model to study lipotoxic events at the cellular level. Specifically, we examined strains that were deficient in triglyceride biosynthesis ( dga1Delta Iro1Delta), steryl ester production (are1Delta are2Delta), or completely devoid of neutral lipid (dga1Delta Iro1Delta are1Delta are2Delta). The complete absence of neutral lipid biosynthesis leads to defective CLD formation and confers lipoapoptosis in the presence of exogenous unsaturated fatty acid (UFA). This phenomenon was associated with the activation of stress response pathways as well as the induction ER safeguards such as the unfolded protein response (UPR). Conversely, saturated fatty acids (SFA) were tolerated in the absence of neutral lipid storage and this reflects an alternate salvage point satisfied by altered phospholipid metabolism. Epistatic interactions identified additional components, such as proteins involved in golgi-to-ER transport (GET proteins), acting as potential modifiers of neutral lipid homeostasis. Furthermore, the genome-wide assessment of growth on PO has correlated enzymes of DNA metabolism and vesicular transport proteins as mediators of CLD biogenesis. The experimental results described in this thesis reveal previously undiscovered aspects of neutral lipid biology and has identified potential avenues for the prevention and treatment of their associated diseases. |
