Mitochondrial retrograde signaling (MRS) pathways relay the functional status of mitochondria to elicit homeostatic or adaptive changes in nuclear gene expression. Budding yeast have "intergenomic signaling" pathways that sense the amount of mitochondrial DNA (mtDNA) independently of oxidative phosphorylation (OXPHOS), the primary function of genes encoded by mtDNA. However, MRS pathways that sense the amount of mtDNA in mammalian cells remain poorly understood. We found that mtDNA-depleted IMR90 cells can sustain OXPHOS for a significant amount of time, providing a robust model system to interrogate human intergenomic signaling. We identified FAM43A, a largely uncharacterized protein, as a CHK2-dependent early responder to mtDNA depletion. Depletion of FAM43A activates a mitochondrial biogenesis program, resulting in an increase in mitochondrial mass and mtDNA copy number via CHK2-mediated upregulation of the p53R2 form of ribonucleotide reductase. We propose that FAM43A performs a checkpoint-like function to limit mitochondrial biogenesis and turnover under conditions of mtDNA depletion or replication stress.