The nuclear genome drives differences in immune cell populations and differentiation potentials, in part regulated by changes in metabolism. Despite this connection, the role of mitochondrial DNA (mtDNA) polymorphisms (SNP) in this process has not been examined. Using mitochondrial nuclear exchange (MNX) mice, we and others have shown that mtDNA strongly influences varying aspects of cell biology and disease. Based upon an established connection between mitochondria and immune cell polarization, we hypothesized that mtDNA SNP alter immune cell development, trafficking, and/or differentiation. Innate and adaptive immune cell populations were isolated and characterizated from the peritoneum and spleen. While most differences between mouse strains are regulated by nuclear DNA (nDNA), there are selective changes that are mediated by mtDNA differences (e.g., macrophage (CD11c) differentiation), These findings highlight how nuclear-mitochondrial crosstalk may alter pathology and physiology via regulation of specific components of the immune system. •mtDNA SNP are quantitative trait loci for the immune system.•Major baseline strain differences in immune cell populations appear to be driven by nuclear encoded factors.•mtDNA regulates selective changes in immune cell populations.•Primary mtDNA-mediated effects drive immune cell differentiation/polarization.•CD11c+ expression in large peritoneal macrophages appears to exhibit strong regulation by mtDNA.