Mitochondrial function is critically dependent on the folding of the mitochondrial inner membrane into cristae; indeed, numerous human diseases are associated with aberrant crista morphologies. With the MICOS complex, OPA1 and the F 1 F o ‐ATP synthase, key players of cristae biogenesis have been identified, yet their interplay is poorly understood. Harnessing super‐resolution light and 3D electron microscopy, we dissect the roles of these proteins in the formation of cristae in human mitochondria. We individually disrupted the genes of all seven MICOS subunits in human cells and re‐expressed Mic10 or Mic60 in the respective knockout cell line. We demonstrate that assembly of the MICOS complex triggers remodeling of pre‐existing unstructured cristae and de novo formation of crista junctions (CJs) on existing cristae. We show that the Mic60‐subcomplex is sufficient for CJ formation, whereas the Mic10‐subcomplex controls lamellar cristae biogenesis. OPA1 stabilizes tubular CJs and, along with the F 1 F o ‐ATP synthase, fine‐tunes the positioning of the MICOS complex and CJs. We propose a new model of cristae formation, involving the coordinated remodeling of an unstructured crista precursor into multiple lamellar cristae.

The mitochondrial inner membrane is highly folded and displays a complex molecular architecture. Cristae junctions are highly curved tubular openings that separate cristae membrane invaginations from the surrounding boundary membrane. Despite their central role in many vital cellular processes like apoptosis, the details of cristae junction formation remain elusive. Here we identify Mic10, a core subunit of the recently discovered MICOS complex, as an inner mitochondrial membrane protein with the ability to change membrane morphology in vitro and in vivo. We show that Mic10 spans the inner membrane in a hairpin topology and that its ability to sculpt membranes depends on oligomerization through a glycine-rich motif. Oligomerization mutants fail to induce curvature in model membranes, and when expressed in yeast, mitochondria display an altered inner membrane architecture characterized by drastically decreased numbers of cristae junctions. Thus, we demonstrate that membrane sculpting by Mic10 is essential for cristae junction formation.

The multisubunit mitochondrial contact site and cristae organizing system (MICOS) plays an important role in cristae junction formation. Tarasenko et al. show that the MICOS component Mic60 actively bends membranes and that this activity is evolutionarily conserved and necessary for cristae structure.