Structural and Mechanistic Bases of Drp1-Cardiolipin Interactions in Mitochondrial Fission

Abstract

Mitochondria are dynamic organelles that undergo regulated cycles of fission and fusion. Emergent data indicate that a direct interaction of the cytosolic, mechanochemical GTPase, dynamin-related protein 1 (Drp1) with mitochondrial outer membrane-localized cardiolipin (CL) catalyzes mitochondrial fission. However, neither the identity of the amino acid residues nor the mechanism by which Drp1 accomplishes specific CL recognition and subsequent membrane remodeling is known. Here using a comprehensive toolkit of structural, biochemical, biophysical, and cell biological approaches, we reveal the structure, identity, and mechanism by which the intrinsically disordered Drp1 variable domain (VD) specifically binds and remodels membrane CL. We demonstrate that lipid-packing defects in CL-containing membranes enable specific Drp1-CL recognition independent of CL acyl chain composition or shape. We further show that the Drp1 VD intercalates into the hydrocarbon core of the membrane to specifically restrict CL motion and induce transient CL non-bilayer topological transitions that facilitate membrane constriction. We reveal a disorder-to-order structural transition of the Drp1 VD upon CL binding, and identify critical VD residues, the substitution of which selectively reduces Drp1-CL binding affinity. Remarkably, the expression of mutant VD in Drp1-null cells induces the formation of hitherto unseen, ring-like ‘donut’ mitochondria subsequent to mitochondrial fission. Our data reveal that a rapid degradation of pro-fission CL to fusogenic phosphatidic acid (PA) at the poles of newly divided mitochondria in mutant VD-expressing cells enable mitochondrial back-fusion and donut formation. These studies firmly establish an indispensable role for Drp1 VD-CL interactions in regulating mitochondrial dynamics and morphology both pre- and post-fission.