The dynamin-related GTPase Dnm1 regulates mitochondrial fission in yeast

The dynamin-related GTPase Dnm1 controls mitochondrial morphology in yeast. Here we show that dnm1 mutations convert the mitochondrial compartment into a planar 'net' of interconnected tubules. We propose that this net morphology results from a defect in mitochondrial fission. Immunogold labelling localizes Dnm1 to the cytoplasmic face of constricted mitochondrial tubules that appear to be dividing and to the ends of mitochondrial tubules that appear to have recently completed division. The activity of Dnm1 is epistatic to that of Fzo1, a GTPase in the outer mitochondrial membrane that regulates mitochondrial fusion. dnm1 mutations prevent mitochondrial fragmentation in fzo1 mutant strains. These findings indicate that Dnm1 regulates mitochondrial fission, assembling on the cytoplasmic face of mitochondrial tubules at sites at which division will occur.     

Mgm1p,a dynamin-related GTPase,is essential for fusion of the mitochondrial outer membrane

In Saccharomyces cerevisiae, mitochondrial fusion requires at least two outer membrane proteins, Fzo1p and Ugo1p. We provide direct evidence that the dynamin-related Mgm1 protein is also required for mitochondrial fusion. Like fzo1 and ugo1 mutants, cells disrupted for the MGM1 gene contain numerous mitochondrial fragments instead of the few long, tubular organelles seen in wild-type cells. Fragmentation of mitochondria in mgm1 mutants is rescued by disrupting DNM1, a gene required for mitochondrial division. In zygotes formed by mating mgm1 mutants, mitochondria do not fuse and mix their contents. Introducing mutations in the GTPase domain of Mgm1p completely block mitochondrial fusion. Furthermore, we show that mgm1 mutants fail to fuse both their mitochondrial outer and inner membranes. Electron microscopy demonstrates that although mgm1 mutants display aberrant mitochondrial inner membrane cristae, mgm1 dnm1 double mutants restore normal inner membrane structures. However, mgm1 dnm1 mutants remain defective in mitochondrial fusion, indicating that mitochondrial fusion requires Mgm1p regardless of the morphology of mitochondria. Finally, we find that Mgm1p, Fzo1p, and Ugo1p physically interact in the mitochondrial outer membrane. Our results raise the possibility that Mgm1p regulates fusion of the mitochondrial outer membrane through its interactions with Fzo1p and Ugo1p.     

Yeast mitochondrial division and distribution require the cortical num1 protein

Yeast mitochondrial division requires the dynamin-related Dnm1 protein. By isolating high-copy suppressors of a dominant-negative Dnm1p mutant, we uncovered an unexpected role in mitochondrial division and inheritance for Num1p, a protein previously shown to facilitate nuclear migration. num1 mutants contain an interconnected network of mitochondrial tubules, remarkably similar to cells lacking Dnm1p, and time-lapse microscopy confirms that mitochondrial fission is greatly reduced in num1Delta cells. We also find that Num1p assembles into punctate structures, which often colocalize with mitochondrial-bound Dnm1p particles. Suggesting a role for both Num1p and Dnm1p in mitochondrial inheritance, we find that num1 dnm1 double mutants accumulate mitochondria in daughter buds and that mother cells are frequently devoid of all mitochondria. Thus, our studies have revealed an additional role for Dnm1p in mitochondrial transmission through its interaction with Num1p, thereby providing a link between mitochondrial division and inheritance.     

Cells lacking Pcp1p/Ugo2p,a rhomboid-like protease required for Mgm1p processing,lose mtDNA and mitochondrial structure in a Dnm1p-dependent manner,but remain competent for mitochondrial fusion

The dynamin-related GTPase, Mgm1p, is critical for the fusion of the mitochondrial outer membrane, maintenance of mitochondrial DNA (mtDNA), formation of normal inner membrane structures, and inheritance of mitochondria. Although there are two forms of Mgm1p, 100 and 90 kDa, their respective functions and the mechanism by which these two forms are produced are not clear. We previously isolated ugo2 mutants in a genetic screen to identify components involved in mitochondrial fusion [J. Cell Biol. 152 (2001) 1123]. In this paper, we show that ugo2 mutants are defective in PCP1, a gene encoding a rhomboid-related serine protease. Cells lacking Pcp1p are defective in the processing of Mgm1p and produce only the larger (100 kDa) form of Mgm1p. Similar to mgm1delta cells, pcp1delta cells contain partially fragmented mitochondria, instead of the long tubular branched mitochondria of wild-type cells. In addition, pcp1delta cells, like mgm1delta cells, lack mtDNA and therefore are unable to grow on nonfermentable medium. Mutations in the catalytic domain lead to complete loss of Pcp1p function. Similar to mgm1delta cells, the fragmentation of mitochondria and loss of mtDNA of pcp1delta cells were rescued when mitochondrial division was blocked by inactivating Dnm1p, a dynamin-related GTPase. Surprisingly, in contrast to mgm1delta cells, which are completely defective in mitochondrial fusion, pcp1delta cells can fuse their mitochondria after yeast cell mating. Our study demonstrates that Pcp1p is required for the processing of Mgm1p and controls normal mitochondrial shape and mtDNA maintenance by producing the 90 kDa form of Mgm1p. However, the processing of Mgm1p is not strictly required for mitochondrial fusion, indicating that the 100 kDa form is sufficient to promote fusion.     

UGO1 encodes an outer membrane protein required for mitochondrial fusion

Membrane fusion plays an important role in controlling the shape, number, and distribution of mitochondria. In the yeast Saccharomyces cerevisiae, the outer membrane protein Fzo1p has been shown to mediate mitochondrial fusion. Using a novel genetic screen, we have isolated new mutants defective in the fusion of their mitochondria. One of these mutants, ugo1, shows several similarities to fzo1 mutants. ugo1 cells contain numerous mitochondrial fragments instead of the few long, tubular organelles seen in wild-type cells. ugo1 mutants lose mitochondrial DNA (mtDNA). In zygotes formed by mating two ugo1 cells, mitochondria do not fuse and mix their matrix contents. Fragmentation of mitochondria and loss of mtDNA in ugo1 mutants are rescued by disrupting DNM1, a gene required for mitochondrial division. We find that UGO1 encodes a 58-kD protein located in the mitochondrial outer membrane. Ugo1p appears to contain a single transmembrane segment, with its NH(2) terminus facing the cytosol and its COOH terminus in the intermembrane space. Our results suggest that Ugo1p is a new outer membrane component of the mitochondrial fusion machinery.     

Division of mitochondria requires a novel DMN1-interacting protein, Net2p

Mitochondria are dynamic organelles that undergo frequent division and fusion, but the molecular mechanisms of these two events are not well understood. Dnm1p, a mitochondria-associated, dynamin-related GTPase was previously shown to mediate mitochondrial fission. Recently, a genome-wide yeast two-hybrid screen identified an uncharacterized protein that interacts with Dnm1p. Cells disrupted in this new gene, which we call NET2, contain a single mitochondrion that consists of a network formed by interconnected tubules, similar to the phenotype of dnm1 Delta cells. NET2 encodes a mitochondria-associated protein with a predicted coiled-coil region and six WD-40 repeats. Immunofluorescence microscopy indicates that Net2p is located in distinct, dot-like structures along the mitochondrial surface, many of which colocalize with the Dnm1 protein. Fluorescence and immunoelectron microscopy shows that Dnm1p and Net2p preferentially colocalize at constriction sites along mitochondrial tubules. Our results suggest that Net2p is a new component of the mitochondrial division machinery.     

The intramitochondrial dynamin-related GTPase,Mgm1p,is a component of a protein complex that mediates mitochondrial fusion

A balance between fission and fusion events determines the morphology of mitochondria. In yeast, mitochondrial fission is regulated by the outer membrane-associated dynamin-related GTPase, Dnm1p. Mitochondrial fusion requires two integral outer membrane components, Fzo1p and Ugo1p. Interestingly, mutations in a second mitochondrial-associated dynamin-related GTPase, Mgm1p, produce similar phenotypes to fzo1 and ugo cells. Specifically, mutations in MGM1 cause mitochondrial fragmentation and a loss of mitochondrial DNA that are suppressed by abolishing DNM1-dependent fission. In contrast to fzo1ts mutants, blocking DNM1-dependent fission restores mitochondrial fusion in mgm1ts cells during mating. Here we show that blocking DNM1-dependent fission in Deltamgm1 cells fails to restore mitochondrial fusion during mating. To examine the role of Mgm1p in mitochondrial fusion, we looked for molecular interactions with known fusion components. Immunoprecipitation experiments revealed that Mgm1p is associated with both Ugo1p and Fzo1p in mitochondria, and that Ugo1p and Fzo1p also are associated with each other. In addition, genetic analysis of specific mgm1 alleles indicates that Mgm1p's GTPase and GTPase effector domains are required for its ability to promote mitochondrial fusion and that Mgm1p self-interacts, suggesting that it functions in fusion as a self-assembling GTPase. Mgm1p's localization within mitochondria has been controversial. Using protease protection and immuno-EM, we have shown previously that Mgm1p localizes to the intermembrane space, associated with the inner membrane. To further test our conclusions, we have used a novel method using the tobacco etch virus protease and confirm that Mgm1p is present in the intermembrane space compartment in vivo. Taken together, these data suggest a model where Mgm1p functions in fusion to remodel the inner membrane and to connect the inner membrane to the outer membrane via its interactions with Ugo1p and Fzo1p, thereby helping to coordinate the behavior of the four mitochondrial membranes during fusion.     

The WD-repeats of Net2p interact with Dnm1p and Fis1p to regulate division of mitochondria

The Net2, Fis1, and Dnm1 proteins are required for the division of mitochondria in the yeast Saccharomyces cerevisiae. Net2p has an amino-terminal region that contains predicted coiled-coil motifs and a carboxyl-terminal domain composed of WD-40 repeats. We found that the amino-terminal part of Net2p interacts with Fis1p, whereas the carboxyl-terminal region interacts with both Dnm1p and Fis1p. Overproduction of either domain of Net2p in yeast cells poisons mitochondrial fission, and the dominant-negative effect caused by the WD-repeats of Net2p is suppressed by increased levels of Dnm1p. Point mutations in the WD-region of Net2p or in the GTPase region of Dnm1p disrupt the normal Net2p-Dnm1p interaction, causing Net2p to lose its normal punctate distribution. Our results suggest that Dnm1p interacts with the WD-repeats of Net2p and in a GTP-dependent manner recruits Net2p to sites of mitochondrial division. Furthermore, our results indicate that Net2p is required for proper assembly of the mitochondrial fission components to regulate organelle division.     

The WD40 protein Caf4p is a component of the mitochondrial fission machinery and recruits Dnm1p to mitochondria

The mitochondrial division machinery regulates mitochondrial dynamics and consists of Fis1p, Mdv1p, and Dnm1p. Mitochondrial division relies on the recruitment of the dynamin-related protein Dnm1p to mitochondria. Dnm1p recruitment depends on the mitochondrial outer membrane protein Fis1p. Mdv1p interacts with Fis1p and Dnm1p, but is thought to act at a late step during fission because Mdv1p is dispensable for Dnm1p localization. We identify the WD40 repeat protein Caf4p as a Fis1p-associated protein that localizes to mitochondria in a Fis1p-dependent manner. Caf4p interacts with each component of the fission apparatus: with Fis1p and Mdv1p through its NH2-terminal half and with Dnm1p through its COOH-terminal WD40 domain. We demonstrate that mdv1delta yeast contain residual mitochondrial fission due to the redundant activity of Caf4p. Moreover, recruitment of Dnm1p to mitochondria is disrupted in mdv1delta caf4delta yeast, demonstrating that Mdv1p and Caf4p are molecular adaptors that recruit Dnm1p to mitochondrial fission sites. Our studies support a revised model for assembly of the mitochondrial fission apparatus.     

Mdv1p is a WD repeat protein that interacts with the dynamin-related GTPase, Dnm1p, to trigger mitochondrial division

Mitochondrial fission is mediated by the dynamin-related GTPase, Dnm1p, which assembles on the mitochondrial outer membrane into punctate structures associated with sites of membrane constriction and fission. We have identified additional nuclear genes required for mitochondrial fission, termed MDV (for mitochondrial division). MDV1 encodes a predicted soluble protein, containing a coiled-coil motif and seven COOH-terminal WD repeats. Genetic and two-hybrid analyses indicate that Mdv1p interacts with Dnm1p to mediate mitochondrial fission. In addition, Mdv1p colocalizes with Dnm1p in fission-mediating punctate structures on the mitochondrial outer membrane. Whereas localization of Mdv1p to these structures requires Dnm1p, localization of Mdv1p to mitochondrial membranes does not. This indicates that Mdv1p possesses a Dnm1p-independent mitochondrial targeting signal. Dnm1p-independent targeting of Mdv1p to mitochondria requires MDV2. Our data indicate that MDV2 also functions separately to regulate the assembly of Dnm1p into punctate structures. In contrast, Mdv1p is not required for the assembly of Dnm1p, but Dnm1p-containing punctate structures lacking Mdv1p are not able to complete division. Our studies suggest that mitochondrial fission is a multi-step process in which Mdv2p regulates the assembly of Dnm1p into punctate structures and together with Mdv1p functions later during fission to facilitate Dnm1p-dependent mitochondrial membrane constriction and/or division.     

The Dynamin-related GTPase,Dnm1p,Controls Mitochondrial Morphology in Yeast

The Saccharomyces cerevisiae Dnm1 protein is structurally related to dynamin, a GTPase required for membrane scission during endocytosis. Here we show that Dnm1p is essential for the maintenance of mitochondrial morphology. Disruption of the DNM1 gene causes the wild-type network of tubular mitochondrial membranes to collapse to one side of the cell but does not affect the morphology or distribution of other cytoplasmic organelles. Dnm1 proteins containing point mutations in the predicted GTP-binding domain or completely lacking the GTP-binding domain fail to rescue mitochondrial morphology defects in a dnm1 mutant and induce dominant mitochondrial morphology defects in wild-type cells. Indirect immunofluorescence reveals that Dnm1p is distributed in punctate structures at the cell cortex that colocalize with the mitochondrial compartment. These Dnm1p-containing structures remain associated with the spherical mitochondria found in an mdm10 mutant strain. In addition, a portion of Dnm1p cofractionates with mitochondrial membranes during differential sedimentation and sucrose gradient fractionation of wild-type cells. Our results demonstrate that Dnm1p is required for the cortical distribution of the mitochondrial network in yeast, a novel function for a dynamin-related protein.     

Peroxisome proliferation in Hansenula polymorpha requires Dnm1p which mediates fission but not de novo formation

We show that the dynamin-like proteins Dnm1p and Vps1p are not required for re-introduction of peroxisomes in Hansenula polymorpha pex3 cells upon complementation with PEX3-GFP. Instead, Dnm1p, but not Vps1p, plays a crucial role in organelle proliferation via fission. In H. polymorpha DNM1 deletion cells (dnm1) a single peroxisome is present that forms long extensions, which protrude into developing buds and divide during cytokinesis. Budding pex11.dnm1 double deletion cells lack these peroxisomal extensions, suggesting that the peroxisomal membrane protein Pex11p is required for their formation. Life cell imaging revealed that fluorescent Dnm1p-GFP spots fluctuate between peroxisomes and mitochondria. On the other hand Pex11p is present over the entire organelle surface, but concentrates during fission at the basis of the organelle extension in dnm1 cells.Our data indicate that peroxisome fission is the major pathway for peroxisome multiplication in H. polymorpha.     

DNM1, a dynamin-related gene, participates in endosomal trafficking in yeast

We identified DNM1, a novel dynamin-related gene in Saccharomyces cerevisiae. Molecular and genetic mapping showed that DNM1 is the most proximal gene to the right of centromere 12, and is predicted to encode a protein of 85 kD, designated Dnm1p. The protein exhibits 41% overall identity with full-length dynamin I and 55% identity with the most highly conserved 400-amino acid GTPase region. Our findings show that like mammalian dynamin, Dnm1p participates in endocytosis; however, it is unlikely to be a cognate homologue. Cells with a disruption in the DNM1 gene showed mating response defects consistent with a delay in receptor-mediated endocytosis. The half-life of the Ste3p pheromone receptor was increased two- to threefold in the dnm1 mutant, demonstrating that Dnm1p participates in the constitutive turnover of the receptor. To define the step in the endocytic pathway at which Dnm1p acts, we analyzed mutant strains at both early and late steps of the process. Initial internalization of epitope-tagged pheromone receptor or of labeled pheromone proceeded with wild-type kinetics. However, delivery of the internalized receptor to the vacuole was greatly impeded during ligand-induced endocytosis. These data suggest that during receptor-mediated endocytosis, Dnm1p acts after internalization, but before fusion with the vacuole. The dnm1 mutant was not defective for sorting of vacuolar proteins, indicating that Dnm1p is not required for transport from the late endosome to the vacuole. Therefore, we suggest that Dnm1p participates at a novel step before fusion with the late endosome.     

Mmm1p, a mitochondrial outer membrane protein, is connected to mitochondrial DNA (mtDNA) nucleoids and required for mtDNA stability

In the yeast Saccharomyces cerevisiae, mitochondria form a branched, tubular reticulum in the periphery of the cell. Mmm1p is required to maintain normal mitochondrial shape and in mmm1 mutants mitochondria form large, spherical organelles. To further explore Mmm1p function, we examined the localization of a Mmm1p-green fluorescent protein (GFP) fusion in living cells. We found that Mmm1p-GFP is located in small, punctate structures on the mitochondrial outer membrane, adjacent to a subset of matrix-localized mitochondrial DNA nucleoids. We also found that the temperature-sensitive mmm1-1 mutant was defective in transmission of mitochondrial DNA to daughter cells immediately after the shift to restrictive temperature. Normal mitochondrial nucleoid structure also collapsed at the nonpermissive temperature with similar kinetics. Moreover, we found that mitochondrial inner membrane structure is dramatically disorganized in mmm1 disruption strains. We propose that Mmm1p is part of a connection between the mitochondrial outer and inner membranes, anchoring mitochondrial DNA nucleoids in the matrix.     

Dnm1p GTPase-mediated mitochondrial fission is a multi-step process requiring the novel integral membrane component Fis1p

Yeast Dnm1p is a soluble, dynamin-related GTPase that assembles on the outer mitochondrial membrane at sites where organelle division occurs. Although these Dnm1p-containing complexes are thought to trigger constriction and fission, little is known about their composition and assembly, and molecules required for their membrane recruitment have not been isolated. Using a genetic approach, we identified two new genes in the fission pathway, FIS1 and FIS2. FIS1 encodes a novel, outer mitochondrial membrane protein with its amino terminus exposed to the cytoplasm. Fis1p is the first integral membrane protein shown to participate in a eukaryotic membrane fission event. In a related study (Tieu, Q., and J. Nunnari. 2000. J. Cell Biol. 151:353-365), it was shown that the FIS2 gene product (called Mdv1p) colocalizes with Dnm1p on mitochondria. Genetic and morphological evidence indicate that Fis1p, but not Mdv1p, function is required for the proper assembly and distribution of Dnm1p-containing fission complexes on mitochondrial tubules. We propose that mitochondrial fission in yeast is a multi-step process, and that membrane-bound Fis1p is required for the proper assembly, membrane distribution, and function of Dnm1p-containing complexes during fission.     

The dynamin-related GTPase, Mgm1p, is an intermembrane space protein required for maintenance of fusion competent mitochondria

Mutations in the dynamin-related GTPase, Mgm1p, have been shown to cause mitochondrial aggregation and mitochondrial DNA loss in Saccharomyces cerevisiae cells, but Mgm1p's exact role in mitochondrial maintenance is unclear. To study the primary function of MGM1, we characterized new temperature sensitive MGM1 alleles. Examination of mitochondrial morphology in mgm1 cells indicates that fragmentation of mitochondrial reticuli is the primary phenotype associated with loss of MGM1 function, with secondary aggregation of mitochondrial fragments. This mgm1 phenotype is identical to that observed in cells with a conditional mutation in FZO1, which encodes a transmembrane GTPase required for mitochondrial fusion, raising the possibility that Mgm1p is also required for fusion. Consistent with this idea, mitochondrial fusion is blocked in mgm1 cells during mating, and deletion of DNM1, which encodes a dynamin-related GTPase required for mitochondrial fission, blocks mitochondrial fragmentation in mgm1 cells. However, in contrast to fzo1 cells, deletion of DNM1 in mgm1 cells restores mitochondrial fusion during mating. This last observation indicates that despite the phenotypic similarities observed between mgm1 and fzo1 cells, MGM1 does not play a direct role in mitochondrial fusion. Although Mgm1p was recently reported to localize to the mitochondrial outer membrane, our studies indicate that Mgm1p is localized to the mitochondrial intermembrane space. Based on our localization data and Mgm1p's structural homology to dynamin, we postulate that it functions in inner membrane remodeling events. In this context, the observed mgm1 phenotypes suggest that inner and outer membrane fission is coupled and that loss of MGM1 function may stimulate Dnm1p-dependent outer membrane fission, resulting in the formation of mitochondrial fragments that are structurally incompetent for fusion.     

Dimeric Dnm1-G385D interacts with Mdv1 on mitochondria and can be stimulated to assemble into fission complexes containing Mdv1 and Fis1

Interactions between yeast Dnm1p, Mdv1p, and Fis1p are required to form fission complexes that catalyze division of the mitochondrial compartment. During the formation of mitochondrial fission complexes, the Dnm1p GTPase self-assembles into large multimeric complexes on the outer mitochondrial membrane that are visualized as punctate structures by fluorescent labeling. Although it is clear that Fis1p.Mdv1p complexes on mitochondria are required for the initial recruitment of Dnm1p, it is not clear whether Dnm1p puncta assemble before or after this recruitment step. Here we show that the minimum oligomeric form of cytoplasmic Dnm1p is a dimer. The middle domain mutant protein Dnm1G385Dp forms dimers in vivo but fails to assemble into punctate structures. However, this dimeric mutant stably interacts with Mdv1p on the outer mitochondrial membrane, demonstrating that assembly of stable Dnm1p multimers is not required for Dnm1p-Mdv1p association or for mitochondrial recruitment of Dnm1p. Dnm1G385Dp is reported to be a terminal dimer in vitro. We describe conditions that allow assembly of Dnm1G385Dp into functional fission complexes on mitochondria in vivo. Using these conditions, we demonstrate that multimerization of Dnm1p is required to promote reorganization of Mdv1p from a uniform mitochondrial localization into punctate fission complexes. Our studies also reveal that Fis1p is present in these assembled fission complexes. Based on our results, we propose that Dnm1p dimers are initially recruited to the membrane via interaction with Mdv1p.Fis1p complexes. These dimers then assemble into multimers that subsequently promote the reorganization of Mdv1p into punctate fission complexes.     

Inhibition of peroxisome fission,but not mitochondrial fission,increases yeast chronological lifespan

Mitochondria are key players in aging and cell death. It has been suggested that mitochondrial fragmentation, mediated by the Dnm1/Fis1 organelle fission machinery, stimulates aging and cell death. This was based on the observation that Saccharomyces cerevisiae Δdnm1 and Δfis1 mutants show an enhanced lifespan and increased resistance to cell death inducers. However, the Dnm1/Fis1 fission machinery is also required for peroxisome division. Here we analyzed the significance of peroxisome fission in yeast chronological lifespan, using yeast strains in which fission of mitochondria was selectively blocked. Our data indicate that the lifespan extension caused by deletion of FIS1 is mainly due to a defect in peroxisome fission and not caused by a block in mitochondrial fragmentation. These observations are underlined by our observation that deletion of FIS1 does not lead to lifespan extension in yeast peroxisome deficient mutant cells.     

Peroxisome fission in Hansenula polymorpha requires Mdv1 and Fis1, two proteins also involved in mitochondrial fission

We show that Mdv1 and Caf4, two components of the mitochondrial fission machinery in Saccharomyces cerevisiae , also function in peroxisome proliferation. Deletion of MDV1 , CAF4 or both, however, had only a minor effect on peroxisome numbers at peroxisome-inducing growth conditions, most likely related to the fact that Vps1 – and not Dnm1 – is the key player in peroxisome fission in this organism. In contrast, in Hansenula polymorpha , which has only a Dnm1-dependent peroxisome fission machinery, deletion of MDV1 led to a drastic reduction of peroxisome numbers. This phenotype was accompanied by a strong defect in mitochondrial fission. The MDV1 paralog CAF4 is absent in H. polymorpha . In wild-type H. polymorpha , cells Dnm1–mCherry and green fluorescent protein (GFP)–Mdv1 colocalize in spots that associate with both peroxisomes and mitochondria. Furthermore, Fis1 is essential to recruit Mdv1 to the peroxisomal and mitochondrial membrane. However, formation of GFP–Mdv1 spots – and related to this normal organelle fission – is strictly dependent on the presence of Dnm1. In dnm1 cells, GFP–Mdv1 is dispersed over the surface of peroxisomes and mitochondria. Also, in H. polymorpha mdv1 or fis1 cells, the number of Dnm1–GFP spots is strongly reduced. These spots still associate to organelles but are functionally inactive.     

Mmm2p, a mitochondrial outer membrane protein required for yeast mitochondrial shape and maintenance of mtDNA nucleoids

The mitochondrial outer membrane protein, Mmm1p, is required for normal mitochondrial shape in yeast. To identify new morphology proteins, we isolated mutations incompatible with the mmm1-1 mutant. One of these mutants, mmm2-1, is defective in a novel outer membrane protein. Lack of Mmm2p causes a defect in mitochondrial shape and loss of mitochondrial DNA (mtDNA) nucleoids. Like the Mmm1 protein (Aiken Hobbs, A.E., M. Srinivasan, J.M. McCaffery, and R.E. Jensen. 2001. J. Cell Biol. 152:401-410.), Mmm2p is located in dot-like particles on the mitochondrial surface, many of which are adjacent to mtDNA nucleoids. While some of the Mmm2p-containing spots colocalize with those containing Mmm1p, at least some of Mmm2p is separate from Mmm1p. Moreover, while Mmm2p and Mmm1p both appear to be part of large complexes, we find that Mmm2p and Mmm1p do not stably interact and appear to be members of two different structures. We speculate that Mmm2p and Mmm1p are components of independent machinery, whose dynamic interactions are required to maintain mitochondrial shape and mtDNA structure.