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

Wong ED, Wagner JA, Scott SV, Okreglak V, Holewinske TJ, Cassidy-Stone A, Nunnari J - J. Cell Biol. (2003)

Bottom Line: In yeast, mitochondrial fission is regulated by the outer membrane-associated dynamin-related GTPase, Dnm1p.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.

View Article: PubMed Central - PubMed

Affiliation: Section of Molecular and Cellular Biology, University of California, Davis, Davis, California, 95616, USA.

ABSTRACT
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.

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Intragenic complementation is observed between mgm1 temperature-sensitive mutants. (A) Schematic of MGM1 domain structure designating the position of the temperature-sensitive mutations analyzed for intragenic complementation is shown. (B) Analysis of intragenic complementation of mgm1ts alleles. Growth on glycerol is indicated by +.
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fig4: Intragenic complementation is observed between mgm1 temperature-sensitive mutants. (A) Schematic of MGM1 domain structure designating the position of the temperature-sensitive mutations analyzed for intragenic complementation is shown. (B) Analysis of intragenic complementation of mgm1ts alleles. Growth on glycerol is indicated by +.

Mentions: To further test whether Mgm1p self-assembles in vivo, we analyzed diploids from crosses of several mgm1ts mutants, previously isolated in a forward genetic screen for mutants unable to maintain mtDNA at elevated temperatures (Fig. 4; Meeusen et al., 1999). We reasoned that if Mgm1p self-interacts, two different mutations in MGM1 may be able to compensate for one another within an assembled Mgm1p complex and restore mitochondrial fusion activity, resulting in maintenance of mtDNA and growth on a nonfermentable carbon source at the nonpermissive temperature. Interestingly, the entire collection of mgm1ts mutants isolated in the screen contained mutations only in either the GTPase domain or the GED, underscoring the importance of these domains in MGM1 function (Fig. 4 A; unpublished data). A combination of a subset of these mgm1ts alleles in diploids grew at the nonpermissive temperature on media containing glycerol, indicating that they displayed intragenic complementation (Fig. 4 B). In total, data from our genetic analysis of mgm1 mutants support a model where Mgm1p functions in fusion as a self-assembling GTPase.


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

Wong ED, Wagner JA, Scott SV, Okreglak V, Holewinske TJ, Cassidy-Stone A, Nunnari J - J. Cell Biol. (2003)

Intragenic complementation is observed between mgm1 temperature-sensitive mutants. (A) Schematic of MGM1 domain structure designating the position of the temperature-sensitive mutations analyzed for intragenic complementation is shown. (B) Analysis of intragenic complementation of mgm1ts alleles. Growth on glycerol is indicated by +.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172654&req=5

fig4: Intragenic complementation is observed between mgm1 temperature-sensitive mutants. (A) Schematic of MGM1 domain structure designating the position of the temperature-sensitive mutations analyzed for intragenic complementation is shown. (B) Analysis of intragenic complementation of mgm1ts alleles. Growth on glycerol is indicated by +.
Mentions: To further test whether Mgm1p self-assembles in vivo, we analyzed diploids from crosses of several mgm1ts mutants, previously isolated in a forward genetic screen for mutants unable to maintain mtDNA at elevated temperatures (Fig. 4; Meeusen et al., 1999). We reasoned that if Mgm1p self-interacts, two different mutations in MGM1 may be able to compensate for one another within an assembled Mgm1p complex and restore mitochondrial fusion activity, resulting in maintenance of mtDNA and growth on a nonfermentable carbon source at the nonpermissive temperature. Interestingly, the entire collection of mgm1ts mutants isolated in the screen contained mutations only in either the GTPase domain or the GED, underscoring the importance of these domains in MGM1 function (Fig. 4 A; unpublished data). A combination of a subset of these mgm1ts alleles in diploids grew at the nonpermissive temperature on media containing glycerol, indicating that they displayed intragenic complementation (Fig. 4 B). In total, data from our genetic analysis of mgm1 mutants support a model where Mgm1p functions in fusion as a self-assembling GTPase.

Bottom Line: In yeast, mitochondrial fission is regulated by the outer membrane-associated dynamin-related GTPase, Dnm1p.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.

View Article: PubMed Central - PubMed

Affiliation: Section of Molecular and Cellular Biology, University of California, Davis, Davis, California, 95616, USA.

ABSTRACT
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.

Show MeSH
Related in: MedlinePlus