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The role of Fis1p-Mdv1p interactions in mitochondrial fission complex assembly.

Karren MA, Coonrod EM, Anderson TK, Shaw JM - J. Cell Biol. (2005)

Bottom Line: Furthermore, we show that conditional mutations in the Fis1p TPR-like domain cause fission complex assembly defects that are suppressed by mutations in the Mdv1p-predicted coiled coil.We also define separable functions for the Fis1p NH(2)-terminal arm and TPR-like fold.These studies suggest that the concave binding surface of the Fis1p TPR-like fold interacts with Mdv1p during mitochondrial fission and that Mdv1p facilitates Dnm1p recruitment into functional fission complexes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, UT 84132, USA.

ABSTRACT
Mitochondrial division requires coordinated interactions among Fis1p, Mdv1p, and the Dnm1p GTPase, which assemble into fission complexes on the outer mitochondrial membrane. The integral outer membrane protein Fis1p contains a cytoplasmic domain consisting of a tetratricopeptide repeat (TPR)-like fold and a short NH(2)-terminal helix. Although it is known that the cytoplasmic domain is necessary for assembly of Mdv1p and Dnm1p into fission complexes, the molecular details of this assembly are not clear. In this study, we provide new evidence that the Fis1p-Mdv1p interaction is direct. Furthermore, we show that conditional mutations in the Fis1p TPR-like domain cause fission complex assembly defects that are suppressed by mutations in the Mdv1p-predicted coiled coil. We also define separable functions for the Fis1p NH(2)-terminal arm and TPR-like fold. These studies suggest that the concave binding surface of the Fis1p TPR-like fold interacts with Mdv1p during mitochondrial fission and that Mdv1p facilitates Dnm1p recruitment into functional fission complexes.

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Mutations in the TPR-like domain of Fis1-3p cause a temperature-sensitive fission defect. (A) Schematic representations of the yeast Fis1-3p cytoplasmic domain with mutated residues indicated in red and the NH2-terminal 15 residues in blue. Ribbon diagram shows mutated residues in α-helix 4 of the TPR-like domain (left). Space-fill representations of Fis1-3p showing the position of mutated residues in the concave TPR-like binding pocket are shown with (middle) or without (right) the NH2-terminal 15 residues. (B) Growth phenotypes of wild-type and fission mutant strains (4 × 103 cells per spot) grown on S-glycerol selective medium at 25 or 37°C. (C) Quantification of mitochondrial morphology in strains grown at 25 or 37°C (n ≥ 300; SDs are indicated).
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fig1: Mutations in the TPR-like domain of Fis1-3p cause a temperature-sensitive fission defect. (A) Schematic representations of the yeast Fis1-3p cytoplasmic domain with mutated residues indicated in red and the NH2-terminal 15 residues in blue. Ribbon diagram shows mutated residues in α-helix 4 of the TPR-like domain (left). Space-fill representations of Fis1-3p showing the position of mutated residues in the concave TPR-like binding pocket are shown with (middle) or without (right) the NH2-terminal 15 residues. (B) Growth phenotypes of wild-type and fission mutant strains (4 × 103 cells per spot) grown on S-glycerol selective medium at 25 or 37°C. (C) Quantification of mitochondrial morphology in strains grown at 25 or 37°C (n ≥ 300; SDs are indicated).

Mentions: We hypothesized that the concave TPR-like binding pocket of Fis1p is important for interaction with either Mdv1p or Dnm1p. To test this idea, we isolated fis1 temperature-sensitive mutations (fis1-ts; see Materials and methods) and chose one allele with mutations in the concave surface of the TPR-like domain for further study. This allele (fis1-3) encodes three amino acid substitutions (E78D, I85T, and Y88H; Fig. 1 A) that alter semiconserved or conserved residues in helix 4 of the TPR-like domain. Fis1-3p expression and targeting to mitochondria is similar to wild type at both permissive (25°C) and nonpermissive (37°C) temperatures (unpublished data). Individual and pair-wise analyses of each mutation indicate that all three contribute to the temperature-sensitive phenotype of the fis1-3 allele (unpublished data).


The role of Fis1p-Mdv1p interactions in mitochondrial fission complex assembly.

Karren MA, Coonrod EM, Anderson TK, Shaw JM - J. Cell Biol. (2005)

Mutations in the TPR-like domain of Fis1-3p cause a temperature-sensitive fission defect. (A) Schematic representations of the yeast Fis1-3p cytoplasmic domain with mutated residues indicated in red and the NH2-terminal 15 residues in blue. Ribbon diagram shows mutated residues in α-helix 4 of the TPR-like domain (left). Space-fill representations of Fis1-3p showing the position of mutated residues in the concave TPR-like binding pocket are shown with (middle) or without (right) the NH2-terminal 15 residues. (B) Growth phenotypes of wild-type and fission mutant strains (4 × 103 cells per spot) grown on S-glycerol selective medium at 25 or 37°C. (C) Quantification of mitochondrial morphology in strains grown at 25 or 37°C (n ≥ 300; SDs are indicated).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171191&req=5

fig1: Mutations in the TPR-like domain of Fis1-3p cause a temperature-sensitive fission defect. (A) Schematic representations of the yeast Fis1-3p cytoplasmic domain with mutated residues indicated in red and the NH2-terminal 15 residues in blue. Ribbon diagram shows mutated residues in α-helix 4 of the TPR-like domain (left). Space-fill representations of Fis1-3p showing the position of mutated residues in the concave TPR-like binding pocket are shown with (middle) or without (right) the NH2-terminal 15 residues. (B) Growth phenotypes of wild-type and fission mutant strains (4 × 103 cells per spot) grown on S-glycerol selective medium at 25 or 37°C. (C) Quantification of mitochondrial morphology in strains grown at 25 or 37°C (n ≥ 300; SDs are indicated).
Mentions: We hypothesized that the concave TPR-like binding pocket of Fis1p is important for interaction with either Mdv1p or Dnm1p. To test this idea, we isolated fis1 temperature-sensitive mutations (fis1-ts; see Materials and methods) and chose one allele with mutations in the concave surface of the TPR-like domain for further study. This allele (fis1-3) encodes three amino acid substitutions (E78D, I85T, and Y88H; Fig. 1 A) that alter semiconserved or conserved residues in helix 4 of the TPR-like domain. Fis1-3p expression and targeting to mitochondria is similar to wild type at both permissive (25°C) and nonpermissive (37°C) temperatures (unpublished data). Individual and pair-wise analyses of each mutation indicate that all three contribute to the temperature-sensitive phenotype of the fis1-3 allele (unpublished data).

Bottom Line: Furthermore, we show that conditional mutations in the Fis1p TPR-like domain cause fission complex assembly defects that are suppressed by mutations in the Mdv1p-predicted coiled coil.We also define separable functions for the Fis1p NH(2)-terminal arm and TPR-like fold.These studies suggest that the concave binding surface of the Fis1p TPR-like fold interacts with Mdv1p during mitochondrial fission and that Mdv1p facilitates Dnm1p recruitment into functional fission complexes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, UT 84132, USA.

ABSTRACT
Mitochondrial division requires coordinated interactions among Fis1p, Mdv1p, and the Dnm1p GTPase, which assemble into fission complexes on the outer mitochondrial membrane. The integral outer membrane protein Fis1p contains a cytoplasmic domain consisting of a tetratricopeptide repeat (TPR)-like fold and a short NH(2)-terminal helix. Although it is known that the cytoplasmic domain is necessary for assembly of Mdv1p and Dnm1p into fission complexes, the molecular details of this assembly are not clear. In this study, we provide new evidence that the Fis1p-Mdv1p interaction is direct. Furthermore, we show that conditional mutations in the Fis1p TPR-like domain cause fission complex assembly defects that are suppressed by mutations in the Mdv1p-predicted coiled coil. We also define separable functions for the Fis1p NH(2)-terminal arm and TPR-like fold. These studies suggest that the concave binding surface of the Fis1p TPR-like fold interacts with Mdv1p during mitochondrial fission and that Mdv1p facilitates Dnm1p recruitment into functional fission complexes.

Show MeSH
Related in: MedlinePlus