Based on studies performed with Dnm1, the yeast ortholog of Drp1, it was originally proposed that when Drp1 translocates to mitochondria, it docks onto the Fis1 protein. Although Fis1 is highly conserved throughout the eukaryotic kingdom including humans, mammalian homologues of Mdv1 and Caf4 have yet to be found. Moreover, recent evidence indicates that Fis1 regulates mitochondrial morphology in mammals by recruiting not Drp1, but a GTPase regulator LY2835219 CDK inhibitor domain-containing protein termed TBC1D15. In addition, in mammalian cells other mitochondrial proteins have been proposed to be essential for mitochondrial recruitment and functional regulation of Drp1. Two prominent examples are the mitochondrial fission factor and the mitochondrial dynamics proteins of 51 and 49 kDa, which have been implicated both in Drp1 mitochondrial recruitment and in stimulation of mitochondrial fission. Importantly, neither Mff nor MiD49 can effectively increase the GTP hydrolysis activity of Drp1 as classical GTPase effectors do, therefore whether this interaction is in itself sufficient to trigger functional activation of Drp1 remains unproven. In addition to proteins, specific lipids can also act as regulatory factors of classical dynamin and dynamin-related proteins. Classical dynamins contain a Pleckstrin Homology domain that interacts with polyanionic phosphatidylinositol bisphosphate to regulate protein assembly and function. On the other hand, dynamin-like proteins such as yeast Mgm1 and human Opa1 do not contain a recognizable PH domain, even though they have been shown to interact with the mitochondrionspecific polyanionic phospholipid cardiolipin. Moreover the interaction of CL with Mgm1 and Opa1 stimulates GTPase activity. CL thus appears to play a key role in mitochondrial function. CL is primarily localized at the inner mitochondrial membrane, although significant amounts are also found at the MOM, particularly at the so-called mitochondrial membrane contact sites. Interestingly, during apoptosis Drp1 translocates from cytosol to MOM and co-localizes with Bax, which in turn is known to target to mitochondrial foci enriched in CL. Moreover, we have found that Drp1 interacts directly with liposomes containing CL. Prompted by these observations, we have attempted to gain further insight into the molecular specificity and functional consequences of the Drp1:CL interaction. In this report, we provide evidence that Drp1 interacts specifically with CL through its B insert, and also show that this interaction promotes Drp1 oligomerization and stimulation of its GTPase activity. In addition, it has been found that some physiological events link with an increase in mitochondrial fission, such as apoptosis or mitophagy, trigger CL externalization to MOM. We previously reported that Drp1 binds to CL-containing liposomes. Interestingly, different lines of evidence indicate that two other mitochondrial dynamin-like proteins, Mgm1 and Opa1 also interact directly with CL, and that this interaction is important for the functional regulation of these proteins.