The third generation human mitochondria-focused cDNA microarray (hMitChip3) and its software for analysis of gene expression in cells derived from healthy and diseased individuals

Abstract

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Mitochondria play a central role in the life cycle of the cell in health and disease. These intracellular organelles are in essence “energy factories” providing the ATP for the cell and are involved in calcium signaling, steroid hormone and heme biosynthesis, generation of radical oxygen species, apoptosis and a significant number of other metabolic activities. Human mitochondrial DNA (mtDNA) codes for 37 genes; yet, most proteins needed for mitochondrial structure and function are encoded by nuclear DNA (nDNA). While defects in mtDNA or nDNA result in rare genetic syndromes such as childhood mitochondrial encephalomyopathies, primary or secondary mitochondrial dysfunction has been associated with the pathogenesis of a wide range of common abnormalities, such as obesity, diabetes, cancer, neurodegeneration, cardiomyopathy, coronary artery disease, depression and aging. To facilitate the systematic study of the role of mitochondria in health and disease, we developed a third generation human mitochondria-focused cDNA microarray (hMitChip3) and software for analysis of the gene expression profile. hMitChip3 consists of mtDNA-encoded 37 genes, nDNA-encoded 1,053 genes and hundreds of control-elements for filtering data and calculating expression ratios, all in triplicate. Functional categories of the hMitChip3 genes include mitochondrial structure and biogenesis, metabolism, energy production, signal transduction, survival, apoptosis, and stress response. Analysis of Cy3- and Cy5-labeled melanocyte and melanoma cDNA samples hybridized to hMitChip3 to study the Warburg effect (i.e. tumor cells produce ATP by metabolizing glucose at a significantly higher rate than normal cells) demonstrated consistency (R² = 1.0) of informative gene expression among triplicate signals within hMitChip3 and reproducibility (R² > 0.80) of hMitChip3 in triplicate experiments. We shall present both statistical data showing the high quality of hMitChip3, and the accompanying software, as a novel integral tool for mitochondria-oriented research.