UC Davis

Mitochondria are derived from bacteria and have evolved to become the metabolic hubs of eukaryotic cells. They have retained a reduced genome, which primarily encodes genes essential for oxidative phosphorylation. In humans, mitochondrial DNA (mtDNA) copy number varies between 100-1000 copies per cell, depending on the cell type. This variation in mtDNA number across tissue types is likely tuned to a tissue’s metabolic needs. Depletion of mtDNA in cells causes mitochondrial diseases and is also associated with numerous types of cancers, metabolic disorders, and cardiovascular disease. Although a key feature of human homeostasis, we still do not understand how cells regulate mtDNA copy number. Critical to mtDNA copy number control, is an understanding of the unit of mtDNA inheritance, which is a higher order packaged mtDNA structure, termed a nucleoid with each nucleoid containing 1 or 2 mtDNA. The mitochondrial genome is compacted into nucleoids, primarily by TFAM, an HMG box DNA binding protein. In addition to TFAM, a subset of nucleoids contains proteins that direct the replication and transcription of mtDNA. Increased or decreased dosage of—as well as mutations in—key nucleoid-associated proteins alters mtDNA levels. mtDNA copy number can also be modulated by small molecules that target proteins directly involved in mtDNA gene expression and/or replication, such as the anti-viral 2'3'-dideoxycytidine (ddC), which targets the mtDNA polymerase PolG. Using ddC, we have created a dynamic system for studying the mtDNA copy number in human cells. In this system, we depleted mtDNA copy number to various levels and measured the cellular response to loss of mtDNA and identified potential pathways involved in recovering mtDNA copy number. Our results suggest that the integrated stress response pathway plays a key role in re-initiating mtDNA synthesis during the recovery phase from mtDNA depletion. These results used to further understand how homeostatic levels of mtDNA are maintained in cells.