Molecular Mechanisms Underlying Aerobic Respiration and Heme Regulation in Yeast
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Heme is an important signaling molecule with diverse functions in living organisms ranging from regulating gene transcription to circadian rhythm. In yeast, heme serves as a secondary messenger of oxygen, as heme synthesis depends on the intracellular levels of oxygen. Yeast and higher eukaryotes have adapted to adverse conditions including low oxygen or hypoxia. To understand the molecular mechanisms underlying hypoxia tolerance on a genome-wide scale, protein localization was studied using green fluorescence protein (GFP) tagged library of yeast genes under normoxia and hypoxia by Henke and colleagues. This study identified more than 200 proteins that change their localization under hypoxia. Among them were proteins of the chromatin remodeling SWI/SNF complex. Six proteins of the SWI/SNF complex- Swi3, Snf5, Snf6, Snf11, Snf12 and Swp82 relocalized under hypoxia. One of the objectives of this dissertation was to study the mechanism of regulation of cellular bioenergetics and respiration by the SWI/SNF proteins. Measurement of oxygen consumption rate and promoter activity show that Swi3, not Swi2, regulates aerobic gene expression and oxygen consumption. The levels of mitochondrial respiratory chain complex proteins were found to be increased in Δswi3 cells as compared to the parent cells. Deletion of SWI3 also induced the expression of aerobic respiration genes under high heme conditions. Computational analysis of the promoters of the genes bound by the human homologs of Swi3, BAF155 and BAF170 also show that they modulate aerobic respiration genes. In the next part of the dissertation, we have studied the regulation of a transcription factor Gis1 which was initially identified as a stress response regulator. In the previous genome-wide study of protein sub-cellular localization under hypoxia, Gis1 was identified as one of the fast responders to hypoxia and reoxygenation. Here we have studied the regulation of the transcriptional activity of Gis1 by heme. Gis1 belongs to the JmjC family of histone demethylases. Gis1 protein has two heme regulatory motifs (HRM). We show that the DNA binding zinc finger domain of Gis1 promotes the heme activation of its transcriptional activity, although heme does not affect Gis1 binding to DNA. These results have identified a new class of heme signaling proteins.