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A septin-based hierarchy of proteins required for localized deposition of chitin in the Saccharomyces cerevisiae cell wall.

DeMarini DJ, Adams AE, Fares H, De Virgilio C, Valle G, Chuang JS, Pringle JR - J. Cell Biol. (1997)

Bottom Line: In addition, a synthetic-lethal interaction was found between cdc12-5, a temperature-sensitive septin mutation, and a mutant allele of CHS4, which encodes an activator of chitin synthase III.Chs3p, whose normal localization is similar to that of Chs4p, does not localize properly in bni4, chs4, or septin mutant strains or in strains that accumulate excess Bni4p.Taken together, these results suggest that the normal localization of chitin synthase III activity is achieved by assembly of a complex in which Chs3p is linked to the septins via Chs4p and Bni4p.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of North Carolina, Chapel Hill 27599-3280, USA.

ABSTRACT
Just before bud emergence, a Saccharomyces cerevisiae cell forms a ring of chitin in its cell wall; this ring remains at the base of the bud as the bud grows and ultimately forms part of the bud scar marking the division site on the mother cell. The chitin ring seems to be formed largely or entirely by chitin synthase III, one of the three known chitin synthases in S. cerevisiae. The chitin ring does not form normally in temperature-sensitive mutants defective in any of four septins, a family of proteins that are constituents of the "neck filaments" that lie immediately subjacent to the plasma membrane in the mother-bud neck. In addition, a synthetic-lethal interaction was found between cdc12-5, a temperature-sensitive septin mutation, and a mutant allele of CHS4, which encodes an activator of chitin synthase III. Two-hybrid analysis revealed no direct interaction between the septins and Chs4p but identified a novel gene, BNI4, whose product interacts both with Chs4p and Cdc10p and with one of the septins, Cdc10p; this analysis also revealed an interaction between Chs4p and Chs3p, the catalytic subunit of chitin synthase III. Bni4p has no known homologues; it contains a predicted coiled-coil domain, but no other recognizable motifs. Deletion of BNI4 is not lethal, but causes delocalization of chitin deposition and aberrant cellular morphology. Overexpression of Bni4p also causes delocalization of chitin deposition and produces a cellular morphology similar to that of septin mutants. Immunolocalization experiments show that Bni4p localizes to a ring at the mother-bud neck that lies predominantly on the mother-cell side (corresponding to the predominant site of chitin deposition). This localization depends on the septins but not on Chs4p or Chs3p. A GFP-Chs4p fusion protein also localizes to a ring at the mother-bud neck on the mother-cell side. This localization is dependent on the septins, Bni4p, and Chs3p. Chs3p, whose normal localization is similar to that of Chs4p, does not localize properly in bni4, chs4, or septin mutant strains or in strains that accumulate excess Bni4p. In contrast, localization of the septins is essentially normal in bni4, chs4, and chs3 mutant strains and in strains that accumulate excess Bni4p. Taken together, these results suggest that the normal localization of chitin synthase III activity is achieved by assembly of a complex in which Chs3p is linked to the septins via Chs4p and Bni4p.

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Model for the interactions among the chitin synthase  III complex, Bni4p, and the septins. (A) The transmembrane protein Chs3p synthesizes chitin, which is deposited in the cell wall  (solid arrows). Chs4p, anchored to the cytoplasmic face of the  plasma membrane by its (presumably) prenylated COOH terminus, interacts both with Chs3p and with Bni4p, which is tethered  to the septin complex by its interaction with Cdc10p. The box  with the question mark indicates a postulated protein that is sufficient to anchor the septin complex to the cell membrane in the  absence of Bni4p, Chs4p, and/or Chs3p. (B) In the absence of  Bni4p or the septin complex, the chitin synthase III complex diffuses through the lipid bilayer (dashed arrows), resulting in delocalized chitin delocalization.
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Figure 10: Model for the interactions among the chitin synthase III complex, Bni4p, and the septins. (A) The transmembrane protein Chs3p synthesizes chitin, which is deposited in the cell wall (solid arrows). Chs4p, anchored to the cytoplasmic face of the plasma membrane by its (presumably) prenylated COOH terminus, interacts both with Chs3p and with Bni4p, which is tethered to the septin complex by its interaction with Cdc10p. The box with the question mark indicates a postulated protein that is sufficient to anchor the septin complex to the cell membrane in the absence of Bni4p, Chs4p, and/or Chs3p. (B) In the absence of Bni4p or the septin complex, the chitin synthase III complex diffuses through the lipid bilayer (dashed arrows), resulting in delocalized chitin delocalization.

Mentions: The results described above suggest a model in which a hierarchic assembly of proteins based on the septins is responsible for the spatial localization of cell-wall chitin deposition by chitin synthase III. In this model (Fig. 10 A), the septins are recruited to and anchored at the presumptive bud site and mother-bud neck by mechanisms independent of the other proteins considered here. The septin complex then localizes Bni4p through the interaction of this protein with Cdc10p, and Bni4p, in turn, localizes the chitin synthase III complex (including Chs3p and Chs4p) through its interaction with Chs4p.


A septin-based hierarchy of proteins required for localized deposition of chitin in the Saccharomyces cerevisiae cell wall.

DeMarini DJ, Adams AE, Fares H, De Virgilio C, Valle G, Chuang JS, Pringle JR - J. Cell Biol. (1997)

Model for the interactions among the chitin synthase  III complex, Bni4p, and the septins. (A) The transmembrane protein Chs3p synthesizes chitin, which is deposited in the cell wall  (solid arrows). Chs4p, anchored to the cytoplasmic face of the  plasma membrane by its (presumably) prenylated COOH terminus, interacts both with Chs3p and with Bni4p, which is tethered  to the septin complex by its interaction with Cdc10p. The box  with the question mark indicates a postulated protein that is sufficient to anchor the septin complex to the cell membrane in the  absence of Bni4p, Chs4p, and/or Chs3p. (B) In the absence of  Bni4p or the septin complex, the chitin synthase III complex diffuses through the lipid bilayer (dashed arrows), resulting in delocalized chitin delocalization.
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Related In: Results  -  Collection

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Figure 10: Model for the interactions among the chitin synthase III complex, Bni4p, and the septins. (A) The transmembrane protein Chs3p synthesizes chitin, which is deposited in the cell wall (solid arrows). Chs4p, anchored to the cytoplasmic face of the plasma membrane by its (presumably) prenylated COOH terminus, interacts both with Chs3p and with Bni4p, which is tethered to the septin complex by its interaction with Cdc10p. The box with the question mark indicates a postulated protein that is sufficient to anchor the septin complex to the cell membrane in the absence of Bni4p, Chs4p, and/or Chs3p. (B) In the absence of Bni4p or the septin complex, the chitin synthase III complex diffuses through the lipid bilayer (dashed arrows), resulting in delocalized chitin delocalization.
Mentions: The results described above suggest a model in which a hierarchic assembly of proteins based on the septins is responsible for the spatial localization of cell-wall chitin deposition by chitin synthase III. In this model (Fig. 10 A), the septins are recruited to and anchored at the presumptive bud site and mother-bud neck by mechanisms independent of the other proteins considered here. The septin complex then localizes Bni4p through the interaction of this protein with Cdc10p, and Bni4p, in turn, localizes the chitin synthase III complex (including Chs3p and Chs4p) through its interaction with Chs4p.

Bottom Line: In addition, a synthetic-lethal interaction was found between cdc12-5, a temperature-sensitive septin mutation, and a mutant allele of CHS4, which encodes an activator of chitin synthase III.Chs3p, whose normal localization is similar to that of Chs4p, does not localize properly in bni4, chs4, or septin mutant strains or in strains that accumulate excess Bni4p.Taken together, these results suggest that the normal localization of chitin synthase III activity is achieved by assembly of a complex in which Chs3p is linked to the septins via Chs4p and Bni4p.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of North Carolina, Chapel Hill 27599-3280, USA.

ABSTRACT
Just before bud emergence, a Saccharomyces cerevisiae cell forms a ring of chitin in its cell wall; this ring remains at the base of the bud as the bud grows and ultimately forms part of the bud scar marking the division site on the mother cell. The chitin ring seems to be formed largely or entirely by chitin synthase III, one of the three known chitin synthases in S. cerevisiae. The chitin ring does not form normally in temperature-sensitive mutants defective in any of four septins, a family of proteins that are constituents of the "neck filaments" that lie immediately subjacent to the plasma membrane in the mother-bud neck. In addition, a synthetic-lethal interaction was found between cdc12-5, a temperature-sensitive septin mutation, and a mutant allele of CHS4, which encodes an activator of chitin synthase III. Two-hybrid analysis revealed no direct interaction between the septins and Chs4p but identified a novel gene, BNI4, whose product interacts both with Chs4p and Cdc10p and with one of the septins, Cdc10p; this analysis also revealed an interaction between Chs4p and Chs3p, the catalytic subunit of chitin synthase III. Bni4p has no known homologues; it contains a predicted coiled-coil domain, but no other recognizable motifs. Deletion of BNI4 is not lethal, but causes delocalization of chitin deposition and aberrant cellular morphology. Overexpression of Bni4p also causes delocalization of chitin deposition and produces a cellular morphology similar to that of septin mutants. Immunolocalization experiments show that Bni4p localizes to a ring at the mother-bud neck that lies predominantly on the mother-cell side (corresponding to the predominant site of chitin deposition). This localization depends on the septins but not on Chs4p or Chs3p. A GFP-Chs4p fusion protein also localizes to a ring at the mother-bud neck on the mother-cell side. This localization is dependent on the septins, Bni4p, and Chs3p. Chs3p, whose normal localization is similar to that of Chs4p, does not localize properly in bni4, chs4, or septin mutant strains or in strains that accumulate excess Bni4p. In contrast, localization of the septins is essentially normal in bni4, chs4, and chs3 mutant strains and in strains that accumulate excess Bni4p. Taken together, these results suggest that the normal localization of chitin synthase III activity is achieved by assembly of a complex in which Chs3p is linked to the septins via Chs4p and Bni4p.

Show MeSH
Related in: MedlinePlus