Mitochondria play a key role in ageing. The pursuit of genes that regulate variation in life span and ageing have shown that several nuclear-encoded mitochondrial genes are important. However, the role of mitochondrial encoded genes (mtDNA) is more controversial and our appreciation of the role of mtDNA for the evolution of life span is limited. We use replicated lines of seed beetles that have been artificially selected for long or short life for >190 generations, now showing dramatic phenotypic differences, to test for a possible role of mtDNA in the divergent evolution of ageing and life span. We show that these divergent selection regimes led to the evolution of significantly different mtDNA haplotype frequencies. Selection for a long life and late reproduction generated positive selection for one specific haplotype, which was fixed in most such lines. In contrast, selection for reproduction early in life led to both positive selection as well as negative frequency-dependent selection on two different haplotypes, which were both present in all such lines. Our findings suggest that the evolution of life span was in part mediated by mtDNA, providing support for the emerging general tenet that adaptive evolution of life-history syndromes may involve mtDNA.