Human adenine nucleotide translocases physically and functionally interact with respirasomes

Members of the adenine nucleotide translocase (ANT) family exchange ADP for ATP across the mitochondrial inner membrane, an activity that is essential for oxidative phosphorylation (OXPHOS). Mutations in or dysregulation of ANTs is associated with progressive external ophthalmoplegia, cardiomyopathy, nonsyndromic intellectual disability, apoptosis, and the Warburg effect. Binding partners of human ANTs have not been systematically identified. The absence of such information has prevented a detailed molecular understanding of the assorted ANT-associated diseases, including insight into their disparate phenotypic manifestations. To fill this void, in this study, we define the interactomes of two human ANT isoforms. Analogous to its yeast counterpart, human ANTs associate with heterologous partner proteins, including the respiratory supercomplex (RSC) and other solute carriers. The evolutionarily conserved ANT-RSC association is particularly noteworthy because the composition, and thereby organization, of RSCs in yeast and human is different. Surprisingly, absence of the major ANT isoform only modestly impairs OXPHOS in HEK293 cells, indicating that the low levels of other isoforms provide functional redundancy. In contrast, pharmacological inhibition of OXPHOS expression and function inhibits ANT-dependent ADP/ATP exchange. Thus ANTs and the OXPHOS machinery physically interact and functionally cooperate to enhance ANT transport capacity and mitochondrial respiration.