To elucidate the functional roles of mitochondrial dynamics in vivo, we identified genes that become essential in cells lacking the dynamin-related proteins Fzo1 and Dnm1, which are required for mitochondrial fusion and division, respectively. The screen identified Num1, a cortical protein implicated in mitochondrial division and distribution that also functions in nuclear migration. Our data indicate that Num1, together with Mdm36, forms a physical tether that robustly anchors mitochondria to the cell cortex but plays no direct role in mitochondrial division. Our analysis indicates that Num1-dependent anchoring is essential for distribution of the static mitochondrial network in fzo1 dnm1 cells. Consistently, expression of a synthetic mitochondria-cortex tether rescues the viability of fzo1 dnm1 num1 cells. We find that the cortical endoplasmic reticulum (ER) also is a constituent of the Num1 mitochondria-cortex tether, suggesting an active role for the ER in mitochondrial positioning in cells. Thus, taken together, our findings identify Num1 as a key component of a mitochondria-ER-cortex anchor, which we termed "MECA," that functions in parallel with mitochondrial dynamics to distribute and position the essential mitochondrial network.T he shape and cellular distribution of mitochondria depend on the integrated and regulated activities of mitochondrial division and fusion, motility, and tethering (1). A key question is how these mitochondrial behaviors are coordinated to shape and position mitochondria properly in response to the changing needs of the cell. To begin to address this question, an understanding of the molecular basis of mitochondrial behaviors is essential.The molecular mechanisms underlying mitochondrial division and fusion are best understood in terms of mitochondrial behaviors (1, 2). At the heart of the molecular machines that mediate mitochondrial division and fusion are dynamin-related proteins (DRPs) that function via GTP-dependent self-assembly and GTP hydrolysismediated conformational changes to remodel membranes. The DRP Dnm1/DRP1 (in yeast and mammals, respectively) drives the scission of mitochondrial membranes, and the DRPs Fzo1/ MFN1/2 and Mgm1/OPA1 mediate fusion of the outer and inner mitochondrial membranes, respectively. The relative rates of mitochondrial division and fusion are major determinants of the steadystate structure of the organelle and greatly influence its distribution. Attenuation of mitochondrial division leads to a more interconnected, collapsed, and less distributable mitochondrial network, and attenuation of mitochondrial fusion results in mitochondrial fragmentation and pronounced defects in the transmission and distribution of mtDNA.Although mitochondrial division and fusion are important, additional parallel pathways also are likely to be important for mitochondrial distribution. For example, the stable positioning of mitochondria at specific cellular locations, an indication of active tethering mechanisms, has been observed in many different cell types. In ...

Ping et al. demonstrate that mitochondria-to-plasma membrane anchoring is mediated by Num1 directly interacting with both organelles through two distinct and spatially separated lipid-specific binding domains. These findings suggest a general mechanism for interorganelle tethering.

Mitochondrial transport and anchoring mechanisms work in concert to position mitochondria to meet cellular needs. In yeast, Mmr1 functions as a mitochondrial adaptor for Myo2 to facilitate actin-based transport of mitochondria to the bud. Posttransport, Mmr1 is proposed to anchor mitochondria at the bud tip. Although both functions require an interaction between Mmr1 and mitochondria, the molecular basis of the Mmr1–mitochondria interaction is poorly understood. Our in vitro phospholipid binding assays indicate Mmr1 can directly interact with phospholipid membranes. Through structure–function studies we identified an unpredicted membrane-binding domain composed of amino acids 76–195 that is both necessary and sufficient for Mmr1 to interact with mitochondria in vivo and liposomes in vitro. In addition, our structure–function analyses indicate that the coiled-coil domain of Mmr1 is necessary and sufficient for Mmr1 self-interaction and facilitates the polarized localization of the protein. Disrupting either the Mmr1–membrane interaction or Mmr1 self-interaction leads to defects in mitochondrial inheritance. Therefore, direct membrane binding and self-interaction are necessary for Mmr1 function in mitochondrial inheritance and are utilized as a means to spatially and temporally regulate mitochondrial positioning.