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Yeast Miro GTPase, Gem1p, regulates mitochondrial morphology via a novel pathway.

Frederick RL, McCaffery JM, Cunningham KW, Okamoto K, Shaw JM - J. Cell Biol. (2004)

Bottom Line: Biol.Chem. 278:6495-6502), Gem1p is not required for pheromone-induced yeast cell death.Thus, Gem1p defines a novel mitochondrial morphology pathway which may integrate cell signaling events with mitochondrial dynamics.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Utah, Salt Lake City, UT 84112, USA.

ABSTRACT
Cell signaling events elicit changes in mitochondrial shape and activity. However, few mitochondrial proteins that interact with signaling pathways have been identified. Candidates include the conserved mitochondrial Rho (Miro) family of proteins, which contain two GTPase domains flanking a pair of calcium-binding EF-hand motifs. We show that Gem1p (yeast Miro; encoded by YAL048C) is a tail-anchored outer mitochondrial membrane protein. Cells lacking Gem1p contain collapsed, globular, or grape-like mitochondria. We demonstrate that Gem1p is not an essential component of characterized pathways that regulate mitochondrial dynamics. Genetic studies indicate both GTPase domains and EF-hand motifs, which are exposed to the cytoplasm, are required for Gem1p function. Although overexpression of a mutant human Miro protein caused increased apoptotic activity in cultured cells (Fransson et al., 2003. J. Biol. Chem. 278:6495-6502), Gem1p is not required for pheromone-induced yeast cell death. Thus, Gem1p defines a novel mitochondrial morphology pathway which may integrate cell signaling events with mitochondrial dynamics.

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Cells lacking Gem1p grow slowly on synthetic glycerol medium. Strains (JSY7000 GEM1 or JSY7002 gem1Δ) with the indicated plasmid were spotted on SDextrose or SGlycerol lacking the appropriate amino acid and grown for 3 d at the indicated temperature. (A) Gem1p expressed from its own promoter (pRS416-GEM1) complements the gem1Δ glycerol growth defect. (B) GFP-Gem1p expressed from the TPI promoter (pYX142-GFP-GEM1) partially complements the gem1Δ glycerol growth defect.
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fig2: Cells lacking Gem1p grow slowly on synthetic glycerol medium. Strains (JSY7000 GEM1 or JSY7002 gem1Δ) with the indicated plasmid were spotted on SDextrose or SGlycerol lacking the appropriate amino acid and grown for 3 d at the indicated temperature. (A) Gem1p expressed from its own promoter (pRS416-GEM1) complements the gem1Δ glycerol growth defect. (B) GFP-Gem1p expressed from the TPI promoter (pYX142-GFP-GEM1) partially complements the gem1Δ glycerol growth defect.

Mentions: To determine whether Gem1p is required during mitotic growth, we replaced the entire GEM1 ORF with a HIS3 cassette in the W303 strain background. This gem1Δ strain was viable, but grew at a slightly reduced rate relative to wild type on solid medium containing dextrose (Fig. 2 A, SDextrose 30°C). The gem1Δ strain grew significantly slower than wild type on minimal media containing the nonfermentable carbon source glycerol (Fig. 2 A, SGlycerol 30°C). This defect was enhanced when the strain was grown at 37°C (Fig. 2 A). Introduction of a plasmid encoding wild-type Gem1p restored wild-type growth in gem1Δ cells (Fig. 2 A). Interestingly, DAPI staining indicated that mtDNA nucleoids were retained in ∼75% of gem1Δ cells cultured in dextrose-containing medium overnight. Even after extended log-phase growth in dextrose-containing medium, 43% of gem1Δ cells retain detectable mtDNA (Fig. S1, available at http://www.jcb.org/cgi/content/full/jcb.200405100/DC1). These findings suggest that loss of Gem1p may cause secondary defects in mtDNA expression or integrity.


Yeast Miro GTPase, Gem1p, regulates mitochondrial morphology via a novel pathway.

Frederick RL, McCaffery JM, Cunningham KW, Okamoto K, Shaw JM - J. Cell Biol. (2004)

Cells lacking Gem1p grow slowly on synthetic glycerol medium. Strains (JSY7000 GEM1 or JSY7002 gem1Δ) with the indicated plasmid were spotted on SDextrose or SGlycerol lacking the appropriate amino acid and grown for 3 d at the indicated temperature. (A) Gem1p expressed from its own promoter (pRS416-GEM1) complements the gem1Δ glycerol growth defect. (B) GFP-Gem1p expressed from the TPI promoter (pYX142-GFP-GEM1) partially complements the gem1Δ glycerol growth defect.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172521&req=5

fig2: Cells lacking Gem1p grow slowly on synthetic glycerol medium. Strains (JSY7000 GEM1 or JSY7002 gem1Δ) with the indicated plasmid were spotted on SDextrose or SGlycerol lacking the appropriate amino acid and grown for 3 d at the indicated temperature. (A) Gem1p expressed from its own promoter (pRS416-GEM1) complements the gem1Δ glycerol growth defect. (B) GFP-Gem1p expressed from the TPI promoter (pYX142-GFP-GEM1) partially complements the gem1Δ glycerol growth defect.
Mentions: To determine whether Gem1p is required during mitotic growth, we replaced the entire GEM1 ORF with a HIS3 cassette in the W303 strain background. This gem1Δ strain was viable, but grew at a slightly reduced rate relative to wild type on solid medium containing dextrose (Fig. 2 A, SDextrose 30°C). The gem1Δ strain grew significantly slower than wild type on minimal media containing the nonfermentable carbon source glycerol (Fig. 2 A, SGlycerol 30°C). This defect was enhanced when the strain was grown at 37°C (Fig. 2 A). Introduction of a plasmid encoding wild-type Gem1p restored wild-type growth in gem1Δ cells (Fig. 2 A). Interestingly, DAPI staining indicated that mtDNA nucleoids were retained in ∼75% of gem1Δ cells cultured in dextrose-containing medium overnight. Even after extended log-phase growth in dextrose-containing medium, 43% of gem1Δ cells retain detectable mtDNA (Fig. S1, available at http://www.jcb.org/cgi/content/full/jcb.200405100/DC1). These findings suggest that loss of Gem1p may cause secondary defects in mtDNA expression or integrity.

Bottom Line: Biol.Chem. 278:6495-6502), Gem1p is not required for pheromone-induced yeast cell death.Thus, Gem1p defines a novel mitochondrial morphology pathway which may integrate cell signaling events with mitochondrial dynamics.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Utah, Salt Lake City, UT 84112, USA.

ABSTRACT
Cell signaling events elicit changes in mitochondrial shape and activity. However, few mitochondrial proteins that interact with signaling pathways have been identified. Candidates include the conserved mitochondrial Rho (Miro) family of proteins, which contain two GTPase domains flanking a pair of calcium-binding EF-hand motifs. We show that Gem1p (yeast Miro; encoded by YAL048C) is a tail-anchored outer mitochondrial membrane protein. Cells lacking Gem1p contain collapsed, globular, or grape-like mitochondria. We demonstrate that Gem1p is not an essential component of characterized pathways that regulate mitochondrial dynamics. Genetic studies indicate both GTPase domains and EF-hand motifs, which are exposed to the cytoplasm, are required for Gem1p function. Although overexpression of a mutant human Miro protein caused increased apoptotic activity in cultured cells (Fransson et al., 2003. J. Biol. Chem. 278:6495-6502), Gem1p is not required for pheromone-induced yeast cell death. Thus, Gem1p defines a novel mitochondrial morphology pathway which may integrate cell signaling events with mitochondrial dynamics.

Show MeSH