MIEF1/2 function as adaptors to recruit Drp1 to mitochondria and regulate the association of Drp1 with Mff

Abstract

Mitochondrial dynamics is a fundamental cellular process and recruitment of Drp1 to mitochondria is an essential step in mitochondrial fission. Mff and MIEF1/2 (MiD51/49) serve as key receptors for recruitment of Drp1 to mitochondria in mammals. However, if and how these receptors work together in mitochondrial fission is poorly understood. Here we show that MIEFs interact with both Drp1 and Mff on the mitochondrial surface and serve as adaptors linking Drp1 and Mff together in a trimeric Drp1-MIEF-Mff complex. Thus, MIEFs can regulate the interaction between Drp1 and Mff, and also Mffinduced Drp1 accumulation on mitochondria. It is shown that loss of endogenous MIEFs severely impairs these processes. Additionally, in cells depleted of endogenous MIEF1/2, high levels of exogenous MIEFs sequester Drp1 on the mitochondrial surface, resulting in mitochondrial elongation, whereas low-tomoderate levels of MIEFs promote mitochondrial fission, leading to mitochondrial fragmentation. In sum, the data suggest that MIEFs and Mff work coordinately in Drp1-mediated mitochondrial fission and that the level of MIEF1/2 relative to Mff sets the balance between mitochondrial fission and fusion.Cells need to regulate the morphology of mitochondria in response to various physiological challenges and the dynamin-related GTPase Drp1 has emerged as a central regulator in mitochondrial fission. Drp1 is primarily distributed in the cytoplasm, but shuttles between the cytoplasm and mitochondria 1, 2 . Drp1 recruitment from the cytoplasm to the mitochondrial outer membrane (MOM) is an essential step in mitochondrial fission [3][4][5] . At the MOM, Drp1 is assembled into helical structures that wrap around the mitochondria to induce mitochondrial fission via its GTPase activity 1,5,6 . Several proteins located at the MOM, including Fis1, Mff and MIEFs (MIEF1 and MIEF2, also known as MiD51/MiD49) have been identified as receptors for the recruitment of Drp1 to mitochondria in mammals. While Fis1 was the first proposed Drp1 receptor at the MOM 7,8 , several recent studies suggest that Fis1 plays only a minor role in Drp1 recruitment [9][10][11] . Mff and MIEFs have been identified as alternative receptors for Drp1 9, 12, 13 . Despite they both function independently as receptors to bind and recruit cytosolic Drp1 to the mitochondrial surface, Mff and MIEFs have opposing effects on mitochondrial morphology following exogenous expression: overexpression of Mff results in excessive mitochondrial fragmentation 9, 14 , whereas overexpression of MIEF1 or MIEF2 leads to mitochondrial elongation most likely by inhibiting fission [11][12][13] . Thus, it is believed that Mff is the primary receptor for Drp1 to facilitate mitochondrial fission 9,11,14,15 , whereas MIEFs recruit but presumably suppress Drp1's function by sequestering the protein in an inactive state on the mitochondrial surface 11,13,16 . Although Mff, MIEF1 and MIEF2 as well as hFis1 are known to be simultaneously expressed in cells 17,18 , it is unclear whether and how these receptor...

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“…50 Phosphorylation of CatSper1 is seen to be lower in Ppp3r2 KO compared to WT sperm. Phosphorylation of a mitochondrial elongation factor, Mief1 known to be involved in mitochondrial fission 47,53 was lower in the sperm lacking calcineurin ( Figure 9B-IV). Figure 9B-II shows that phosphorylation of Ccnyl1, a protein involved in sperm Wnt 51 was significantly reduced in sperm lacking calcineurin.…”

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“…Drp1 originates from nuclear DNA and following translation, remains cytosolic , undergoing post‐translational modification, including phosphorylation prior to recruitment to the mitochondria . Although contact with the endoplasmic reticulum is believed to designate the fission point on the mitochondrion , Drp1 is essential for the fission process, inducing fission via its GTPase activity following self‐assembly .…”

“…Drp1 is maintained in its cytosolic location through scaffolding in a complex with AKAP which facilitates its phosphorylation at Ser 637 by protein kinase A (PKA) and inhibits its GTPase activity . Upon dephosphorylation by calcineurin or PP2A and rephosphorylation at Ser 616 by cdk1, Drp1 translocates to the mitochondrion and binds to one of a number of OMM‐localised receptors including Fis1, Mff, MiD49 and MiD51 . The interaction between one of these membrane‐anchored proteins and Drp1 is essential for the fission process, however, the specific mechanism of recruitment is still unclear .…”

“…The interaction between one of these membrane‐anchored proteins and Drp1 is essential for the fission process, however, the specific mechanism of recruitment is still unclear . Although Fis1 was the first mitochondrial receptor to be identified for Drp1 , recent evidence supports Mff as mediating the majority of Drp1 interactions ; overexpression of Mff leads to excessive mitochondrial fragmentation . At the mitochondrial membrane, Drp1 spirals around mitochondria forming a helical structure and causing GTP‐driven constriction which results in severing of both the outer and inner mitochondrial membranes .…”

“…Additional regulation of the mitochondrial recruitment of Drp1 was recently identified through its trimolecular interaction with Mff and MiD49/51. High expression levels of MiD49/51 results in suppression of fission, whereas lower levels of MiD49/51 promote fission .…”