In cardiac tissue, several physiological changes, most notably hypertrophy, cause rather specific increases in the mitochondrial membrane mass. The mechanism for this stimulated membrane biogenesis is not understood. Additionally, it is well documented that various phospholipids and increased free fatty acid levels during hypoxia or in acute myocardial ischemia markedly alter the integrity and function of the energy transducing mitochrondrial membrane. Thus, a more detailed understanding of the molecular basis for the interaction of phospholipids and proteins during their biogenesis and assembly into this key intracellular membrane may provide insight into these lesions which are defined at the clinical level. The well developed mitochondrial system of yeast provides a unique opportunity to examine the relationship of newly made protein and lipid during membrane assembly. We propose to examine the interaction of newly made phospholipid with the ATPase complex during its assembly in the membrane. This complex contains approximately 10% firmly bound phospholipid on a weight basis when isolated in pure form. These studies will employ pulse labeling the lipd or protein components of yeast S. cerevisiae under conditions which perturb either lipid or protein biosynthesis to probe the dependence of each for the proper biogenesis of the ATPase complex. This will be determined by a combination of enzymatic biochemical and immunological methods. In addition, temperature-sensitive mutants, which lack ATPase at the non-permissive temperature, will be used to examine the association of newly made (32P-phosphate labeled) phospholipid with newly made ATPase upon shift to the permissive temperature. These combined studies should offer insights into lipid-protein interaction during assembly and the way exogenous lipids may perturb this.
