C-4 photosynthesis involves a series of biochemical and anatomical traits that significantly improve plant productivity under conditions that reduce the efficiency of C-3 photosynthesis. We explore how evolution of the three classical biochemical types of C-4 photosynthesis (NADP-ME, NAD-ME and PCK types) has affected the functions and properties of mitochondria. Mitochondria in C-4 NAD-ME and PCK types play a direct role in decarboxylation of metabolites for C-4 photosynthesis. Mitochondria in C-4 PCK type also provide ATP for C-4 metabolism, although this role for ATP provision is not seen in NAD-ME type. Such involvement has increased mitochondrial abundance/size and associated enzymatic capacity, led to changes in mitochondrial location and ultrastructure, and altered the role of mitochondria in cellular carbon metabolism in the NAD-ME and PCK types. By contrast, these changes in mitochondrial properties are absent in the C-4 NADP-ME type and C-3 leaves, where mitochondria play no direct role in photosynthesis. From an eco-physiological perspective, rates of leaf respiration in darkness vary considerably among C-4 species but does not differ systematically among the three C-4 types. This review outlines further mitochondrial research in key areas central to the engineering of the C-4 pathway into C-3 plants and to the understanding of variation in rates of C-4 dark respiration.