The small (16,569 base pair) human mitochondrial genome plays a significant role in cell metabolism and homeostasis. Mitochondrial DNA (mtDNA) contributes to the generation of complexes which are essential to oxidative phosphorylation (OXPHOS). As such, mtDNA is directly integrated into mitochondrial biogenesis and signaling and regulates mitochondrial metabolism in concert with nuclear-encoded mitochondrial factors. Mitochondria are a highly dynamic, pleiomorphic network that undergoes fission and fusion events. Within this network, mtDNAs are packaged into structures called nucleoids which are actively distributed in discrete foci within the network. This sensitive organelle is frequently disrupted by insults such as oxidants and inflammatory cytokines, and undergoes genomic damage with double-and single-strand breaks that impair its function. Collectively, mtDNA is emerging as a highly sensitive indicator of cellular stress, which is directly integrated into the mitochondrial network as a contributor of a wide range of critical signaling pathways.