UC San Diego

Drosophila is an emerging model for studying genetic influences on heart function, and has also been found to be highly tolerant to hypoxia. Strategies for controlling metabolism in the hypoxic adult fly heart may give clues to new therapies for myocardial ischemia in humans, however, the mechanisms of their hypoxic metabolic regulation are not well known. We adopt a systems biology approach to discover important hypoxia-tolerance strategies in ATP-generating metabolism in Drosophila heart. First, we built automation technology for rapidly screening the in vivo cardiac hypoxia response in adult flies, and proved its speed by characterizing the wild type over a range of conditions. The assay detected loss- of-function phenotypes in known hypoxia-sensitive mutants. Next we used ¹H NMR metabolomics to discover the major anaerobic end products (lactate, alanine, and acetate), which we built into a genome-wide reconstruction of central metabolism. We fit metabolomic data to the model and used it to examine the benefits of these pathways under hypoxia. The model was then used to predict the effects of a lactate dehydrogenase (LDH) mutant, which were supported by metabolomic, heart phenotype, and whole- body assays on an LDH mutant strain. The model was further refined with gene expression data and used with metabolomic profiling to study the effects of age on the hypoxia response. Recovery of heart rate, whole-body activity, and ATP concentration was delayed in older flies. After fitting the model to metabolomic data for young and old flies, flux-balance analysis pointed to impaired mitochondrial recovery, with excess pyruvate converted to acetate, as the major source of differences between the age groups. Gene expression and the literature on Drosophila aging supported these conclusions. This approach was repeated for a strain of flies that had been experimentally selected to survive chronic hypoxia. Flux- balance modeling suggested that adapted flies may better divert pyruvate flux through pyruvate dehydrogenase rather than pyruvate carboxylase in order to better tolerate acute hypoxia. Gene expression data from microarrays helped support this finding. The dissertation offers clues to hypoxia tolerance in flies, generating hypotheses for further research, and also provides a technology platform for a systematic perturbation analysis