Limits...
UGO1 encodes an outer membrane protein required for mitochondrial fusion.

Sesaki H, Jensen RE - J. Cell Biol. (2001)

Bottom Line: In zygotes formed by mating two ugo1 cells, mitochondria do not fuse and mix their matrix contents.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Anatomy, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA. hsesaki@jhmi.edu

ABSTRACT
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.

Show MeSH

Related in: MedlinePlus

Mitochondrial fusion is defective in ugo1Δ and ugo1Δ dnm1Δ cells. MATa cells containing matrix-targeted RFP under the control of the GAL1 promoter (pHS51) were grown to log phase in SRaf medium and then transferred to SGalSuc medium for 3–5 h to induce the expression of the COX4-RFP fusion protein. MATα cells containing matrix-targeted CFP under the control of the GAL1 promoter (pHS52) were grown to log phase in SGalSuc medium overnight to induce COX-CFP. MATa and α cells were mated for 3.5 h on YEPD medium. The distribution of COX4-RFP and COX4-CFP in representative zygotes containing a medial bud (asterisks) is shown. Zygotes formed by mating between wild-type cells (FY833 and FY834, WT), ugo1Δ mutants (YHS72 and YHS73), dnm1Δ mutants (YHS83 and YHS84), and ugo1Δ dnm1Δ mutants (YHS85 and YHS86) were examined. Bar, 3 μm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2199209&req=5

Figure 6: Mitochondrial fusion is defective in ugo1Δ and ugo1Δ dnm1Δ cells. MATa cells containing matrix-targeted RFP under the control of the GAL1 promoter (pHS51) were grown to log phase in SRaf medium and then transferred to SGalSuc medium for 3–5 h to induce the expression of the COX4-RFP fusion protein. MATα cells containing matrix-targeted CFP under the control of the GAL1 promoter (pHS52) were grown to log phase in SGalSuc medium overnight to induce COX-CFP. MATa and α cells were mated for 3.5 h on YEPD medium. The distribution of COX4-RFP and COX4-CFP in representative zygotes containing a medial bud (asterisks) is shown. Zygotes formed by mating between wild-type cells (FY833 and FY834, WT), ugo1Δ mutants (YHS72 and YHS73), dnm1Δ mutants (YHS83 and YHS84), and ugo1Δ dnm1Δ mutants (YHS85 and YHS86) were examined. Bar, 3 μm.

Mentions: We found that ugo1Δ and ugo1Δ dnm1Δ mutants are defective in fusion. In Fig. 6, representative examples of zygotes containing a medial diploid bud from each mating mixture are shown, but >50 zygotes for each mating mixture were actually examined. When two wild-type cells were mated, mitochondria in the zygote efficiently fused and a complete overlap of the RFP and CFP fluorescence was seen. Consistent with previous studies (Bleazard et al. 1999; Sesaki and Jensen 1999), mitochondrial fusion also occurred in dnm1Δ/dnm1Δ zygotes. Interestingly, dnm1Δ/dnm1Δ zygotes often contained a single tubule emerging from the mitochondrial network of each parent. Fusion appeared to occur at a discrete point near the middle of the zygote. In contrast to wild-type and dnm1Δ cells, fusion was defective in ugo1Δ mutants. ugo1Δ/ugo1Δ zygotes contained many mitochondrial fragments, but these organelles contained only RFP or CFP fluorescence. No organelles containing both fluorophores were seen. Although disruption of DNM1 suppresses the fragmentation of mitochondria and the loss of mtDNA of ugo1Δ mutants, ugo1Δ dnm1Δ double mutants still failed to fuse their mitochondria. Although ugo1Δ dnm1Δ cells displayed tubular mitochondrial shape, each mitochondrial tubule contained only RFP or CFP. We found that mitochondria in ugo1Δ/ugo1Δ or ugo1Δ dnm1Δ/ugo1Δ dnm1Δ zygotes were often closely positioned to each other near the middle of zygotes, but nonetheless did not fuse. Our results thus indicate that ugo1Δ and ugo1Δ dnm1Δ cells are defective in mitochondrial fusion and argue that Ugo1p plays a direct role in the fusion pathway.


UGO1 encodes an outer membrane protein required for mitochondrial fusion.

Sesaki H, Jensen RE - J. Cell Biol. (2001)

Mitochondrial fusion is defective in ugo1Δ and ugo1Δ dnm1Δ cells. MATa cells containing matrix-targeted RFP under the control of the GAL1 promoter (pHS51) were grown to log phase in SRaf medium and then transferred to SGalSuc medium for 3–5 h to induce the expression of the COX4-RFP fusion protein. MATα cells containing matrix-targeted CFP under the control of the GAL1 promoter (pHS52) were grown to log phase in SGalSuc medium overnight to induce COX-CFP. MATa and α cells were mated for 3.5 h on YEPD medium. The distribution of COX4-RFP and COX4-CFP in representative zygotes containing a medial bud (asterisks) is shown. Zygotes formed by mating between wild-type cells (FY833 and FY834, WT), ugo1Δ mutants (YHS72 and YHS73), dnm1Δ mutants (YHS83 and YHS84), and ugo1Δ dnm1Δ mutants (YHS85 and YHS86) were examined. Bar, 3 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199209&req=5

Figure 6: Mitochondrial fusion is defective in ugo1Δ and ugo1Δ dnm1Δ cells. MATa cells containing matrix-targeted RFP under the control of the GAL1 promoter (pHS51) were grown to log phase in SRaf medium and then transferred to SGalSuc medium for 3–5 h to induce the expression of the COX4-RFP fusion protein. MATα cells containing matrix-targeted CFP under the control of the GAL1 promoter (pHS52) were grown to log phase in SGalSuc medium overnight to induce COX-CFP. MATa and α cells were mated for 3.5 h on YEPD medium. The distribution of COX4-RFP and COX4-CFP in representative zygotes containing a medial bud (asterisks) is shown. Zygotes formed by mating between wild-type cells (FY833 and FY834, WT), ugo1Δ mutants (YHS72 and YHS73), dnm1Δ mutants (YHS83 and YHS84), and ugo1Δ dnm1Δ mutants (YHS85 and YHS86) were examined. Bar, 3 μm.
Mentions: We found that ugo1Δ and ugo1Δ dnm1Δ mutants are defective in fusion. In Fig. 6, representative examples of zygotes containing a medial diploid bud from each mating mixture are shown, but >50 zygotes for each mating mixture were actually examined. When two wild-type cells were mated, mitochondria in the zygote efficiently fused and a complete overlap of the RFP and CFP fluorescence was seen. Consistent with previous studies (Bleazard et al. 1999; Sesaki and Jensen 1999), mitochondrial fusion also occurred in dnm1Δ/dnm1Δ zygotes. Interestingly, dnm1Δ/dnm1Δ zygotes often contained a single tubule emerging from the mitochondrial network of each parent. Fusion appeared to occur at a discrete point near the middle of the zygote. In contrast to wild-type and dnm1Δ cells, fusion was defective in ugo1Δ mutants. ugo1Δ/ugo1Δ zygotes contained many mitochondrial fragments, but these organelles contained only RFP or CFP fluorescence. No organelles containing both fluorophores were seen. Although disruption of DNM1 suppresses the fragmentation of mitochondria and the loss of mtDNA of ugo1Δ mutants, ugo1Δ dnm1Δ double mutants still failed to fuse their mitochondria. Although ugo1Δ dnm1Δ cells displayed tubular mitochondrial shape, each mitochondrial tubule contained only RFP or CFP. We found that mitochondria in ugo1Δ/ugo1Δ or ugo1Δ dnm1Δ/ugo1Δ dnm1Δ zygotes were often closely positioned to each other near the middle of zygotes, but nonetheless did not fuse. Our results thus indicate that ugo1Δ and ugo1Δ dnm1Δ cells are defective in mitochondrial fusion and argue that Ugo1p plays a direct role in the fusion pathway.

Bottom Line: In zygotes formed by mating two ugo1 cells, mitochondria do not fuse and mix their matrix contents.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Anatomy, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA. hsesaki@jhmi.edu

ABSTRACT
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.

Show MeSH
Related in: MedlinePlus