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Septin ring assembly involves cycles of GTP loading and hydrolysis by Cdc42p.

Gladfelter AS, Bose I, Zyla TR, Bardes ES, Lew DJ - J. Cell Biol. (2002)

Bottom Line: In performing its roles in actin polarization and transcriptional activation, GTP-Cdc42p is thought to function by activating and/or recruiting effectors to the site of polarization.Excess accumulation of GTP-Cdc42p due to a defect in GTP hydrolysis by the septin-specific alleles might cause unphysiological activation of effectors, interfering with septin assembly.These results suggest that a single GTPase, Cdc42p, can act either as a ras-like GTP-dependent "switch" to turn on effectors or as an EF-Tu-like "assembly factor" using the GTPase cycle to assemble a macromolecular structure.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, NC 27710, USA.

ABSTRACT
At the beginning of the budding yeast cell cycle, the GTPase Cdc42p promotes the assembly of a ring of septins at the site of future bud emergence. Here, we present an analysis of cdc42 mutants that display specific defects in septin organization, which identifies an important role for GTP hydrolysis by Cdc42p in the assembly of the septin ring. The mutants show defects in basal or stimulated GTP hydrolysis, and the septin misorganization is suppressed by overexpression of a Cdc42p GTPase-activating protein (GAP). Other mutants known to affect GTP hydrolysis by Cdc42p also caused septin misorganization, as did deletion of Cdc42p GAPs. In performing its roles in actin polarization and transcriptional activation, GTP-Cdc42p is thought to function by activating and/or recruiting effectors to the site of polarization. Excess accumulation of GTP-Cdc42p due to a defect in GTP hydrolysis by the septin-specific alleles might cause unphysiological activation of effectors, interfering with septin assembly. However, the recessive and dose-sensitive genetic behavior of the septin-specific cdc42 mutants is inconsistent with the septin defect stemming from a dominant interference of this type. Instead, we suggest that assembly of the septin ring involves repeated cycles of GTP loading and GTP hydrolysis by Cdc42p. These results suggest that a single GTPase, Cdc42p, can act either as a ras-like GTP-dependent "switch" to turn on effectors or as an EF-Tu-like "assembly factor" using the GTPase cycle to assemble a macromolecular structure.

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Model for the role of Cdc42p GTP hydrolysis in septin ring assembly. GTP-bound Cdc42p (red) is proposed to bind to septin filaments (green) in a complex together with other assembly factors (blue), which may include the Cdc42p GAPs Rga1p, Rga2p, or Bem3p. Interaction of the assembly complex with the ring is reversible, and we imagine that improper interactions would lead to dissociation of the complex from the ring before GTP hydrolysis. Upon proper docking of the septin assembly complex to the growing septin ring, the GAPs promote GTP hydrolysis by Cdc42p (green arrow), triggering disassembly of the complex and departure of GDP-Cdc42p (pink) and the assembly factors, leaving the septin filament incorporated into the ring. Exchange of bound GDP for GTP catalyzed by the GEF Cdc24p allows repeated cycles of Cdc42p-mediated septin recruitment, resulting in a fully assembled septin ring. The organization of septin filaments within the ring is purely speculative, but the model does not depend on the details of this organization as long as the ring is composed of repeated units (which could equally well be septin complexes rather than polymerized filaments). GAP action is hypothesized to occur upon docking of the assembly complex to the ring, but the timing of GAP association could either be early (if the GAPs are themselves assembly factors; blue) or late (if the GAPs only recognize the docked complex; green arrow). We stress that this model only aims to account for the findings reported here regarding the importance of Cdc42p GTP hydrolysis in septin ring assembly. Cdc42p may well have additional roles in septin ring assembly (e.g., directing the location at which the ring assembles) that were not perturbed by the mutants we analyzed and are not addressed by the model.
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fig8: Model for the role of Cdc42p GTP hydrolysis in septin ring assembly. GTP-bound Cdc42p (red) is proposed to bind to septin filaments (green) in a complex together with other assembly factors (blue), which may include the Cdc42p GAPs Rga1p, Rga2p, or Bem3p. Interaction of the assembly complex with the ring is reversible, and we imagine that improper interactions would lead to dissociation of the complex from the ring before GTP hydrolysis. Upon proper docking of the septin assembly complex to the growing septin ring, the GAPs promote GTP hydrolysis by Cdc42p (green arrow), triggering disassembly of the complex and departure of GDP-Cdc42p (pink) and the assembly factors, leaving the septin filament incorporated into the ring. Exchange of bound GDP for GTP catalyzed by the GEF Cdc24p allows repeated cycles of Cdc42p-mediated septin recruitment, resulting in a fully assembled septin ring. The organization of septin filaments within the ring is purely speculative, but the model does not depend on the details of this organization as long as the ring is composed of repeated units (which could equally well be septin complexes rather than polymerized filaments). GAP action is hypothesized to occur upon docking of the assembly complex to the ring, but the timing of GAP association could either be early (if the GAPs are themselves assembly factors; blue) or late (if the GAPs only recognize the docked complex; green arrow). We stress that this model only aims to account for the findings reported here regarding the importance of Cdc42p GTP hydrolysis in septin ring assembly. Cdc42p may well have additional roles in septin ring assembly (e.g., directing the location at which the ring assembles) that were not perturbed by the mutants we analyzed and are not addressed by the model.

Mentions: The detailed organization of septins within the septin ring is unknown, although the documented ability of septins to polymerize makes it likely that the ring is comprised of septin filaments (Byers and Goetsch, 1976; Byers, 1981; Frazier et al., 1998). Fig. 8 presents a speculative model illustrating how cycles of GTP loading and GTP hydrolysis by Cdc42p may contribute to septin ring assembly. This model is based on the paradigm established for the role of another small G protein, the translation elongation factor EF-Tu (EF-1α in eukaryotes), in protein synthesis (Thompson, 1988).


Septin ring assembly involves cycles of GTP loading and hydrolysis by Cdc42p.

Gladfelter AS, Bose I, Zyla TR, Bardes ES, Lew DJ - J. Cell Biol. (2002)

Model for the role of Cdc42p GTP hydrolysis in septin ring assembly. GTP-bound Cdc42p (red) is proposed to bind to septin filaments (green) in a complex together with other assembly factors (blue), which may include the Cdc42p GAPs Rga1p, Rga2p, or Bem3p. Interaction of the assembly complex with the ring is reversible, and we imagine that improper interactions would lead to dissociation of the complex from the ring before GTP hydrolysis. Upon proper docking of the septin assembly complex to the growing septin ring, the GAPs promote GTP hydrolysis by Cdc42p (green arrow), triggering disassembly of the complex and departure of GDP-Cdc42p (pink) and the assembly factors, leaving the septin filament incorporated into the ring. Exchange of bound GDP for GTP catalyzed by the GEF Cdc24p allows repeated cycles of Cdc42p-mediated septin recruitment, resulting in a fully assembled septin ring. The organization of septin filaments within the ring is purely speculative, but the model does not depend on the details of this organization as long as the ring is composed of repeated units (which could equally well be septin complexes rather than polymerized filaments). GAP action is hypothesized to occur upon docking of the assembly complex to the ring, but the timing of GAP association could either be early (if the GAPs are themselves assembly factors; blue) or late (if the GAPs only recognize the docked complex; green arrow). We stress that this model only aims to account for the findings reported here regarding the importance of Cdc42p GTP hydrolysis in septin ring assembly. Cdc42p may well have additional roles in septin ring assembly (e.g., directing the location at which the ring assembles) that were not perturbed by the mutants we analyzed and are not addressed by the model.
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Related In: Results  -  Collection

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fig8: Model for the role of Cdc42p GTP hydrolysis in septin ring assembly. GTP-bound Cdc42p (red) is proposed to bind to septin filaments (green) in a complex together with other assembly factors (blue), which may include the Cdc42p GAPs Rga1p, Rga2p, or Bem3p. Interaction of the assembly complex with the ring is reversible, and we imagine that improper interactions would lead to dissociation of the complex from the ring before GTP hydrolysis. Upon proper docking of the septin assembly complex to the growing septin ring, the GAPs promote GTP hydrolysis by Cdc42p (green arrow), triggering disassembly of the complex and departure of GDP-Cdc42p (pink) and the assembly factors, leaving the septin filament incorporated into the ring. Exchange of bound GDP for GTP catalyzed by the GEF Cdc24p allows repeated cycles of Cdc42p-mediated septin recruitment, resulting in a fully assembled septin ring. The organization of septin filaments within the ring is purely speculative, but the model does not depend on the details of this organization as long as the ring is composed of repeated units (which could equally well be septin complexes rather than polymerized filaments). GAP action is hypothesized to occur upon docking of the assembly complex to the ring, but the timing of GAP association could either be early (if the GAPs are themselves assembly factors; blue) or late (if the GAPs only recognize the docked complex; green arrow). We stress that this model only aims to account for the findings reported here regarding the importance of Cdc42p GTP hydrolysis in septin ring assembly. Cdc42p may well have additional roles in septin ring assembly (e.g., directing the location at which the ring assembles) that were not perturbed by the mutants we analyzed and are not addressed by the model.
Mentions: The detailed organization of septins within the septin ring is unknown, although the documented ability of septins to polymerize makes it likely that the ring is comprised of septin filaments (Byers and Goetsch, 1976; Byers, 1981; Frazier et al., 1998). Fig. 8 presents a speculative model illustrating how cycles of GTP loading and GTP hydrolysis by Cdc42p may contribute to septin ring assembly. This model is based on the paradigm established for the role of another small G protein, the translation elongation factor EF-Tu (EF-1α in eukaryotes), in protein synthesis (Thompson, 1988).

Bottom Line: In performing its roles in actin polarization and transcriptional activation, GTP-Cdc42p is thought to function by activating and/or recruiting effectors to the site of polarization.Excess accumulation of GTP-Cdc42p due to a defect in GTP hydrolysis by the septin-specific alleles might cause unphysiological activation of effectors, interfering with septin assembly.These results suggest that a single GTPase, Cdc42p, can act either as a ras-like GTP-dependent "switch" to turn on effectors or as an EF-Tu-like "assembly factor" using the GTPase cycle to assemble a macromolecular structure.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, NC 27710, USA.

ABSTRACT
At the beginning of the budding yeast cell cycle, the GTPase Cdc42p promotes the assembly of a ring of septins at the site of future bud emergence. Here, we present an analysis of cdc42 mutants that display specific defects in septin organization, which identifies an important role for GTP hydrolysis by Cdc42p in the assembly of the septin ring. The mutants show defects in basal or stimulated GTP hydrolysis, and the septin misorganization is suppressed by overexpression of a Cdc42p GTPase-activating protein (GAP). Other mutants known to affect GTP hydrolysis by Cdc42p also caused septin misorganization, as did deletion of Cdc42p GAPs. In performing its roles in actin polarization and transcriptional activation, GTP-Cdc42p is thought to function by activating and/or recruiting effectors to the site of polarization. Excess accumulation of GTP-Cdc42p due to a defect in GTP hydrolysis by the septin-specific alleles might cause unphysiological activation of effectors, interfering with septin assembly. However, the recessive and dose-sensitive genetic behavior of the septin-specific cdc42 mutants is inconsistent with the septin defect stemming from a dominant interference of this type. Instead, we suggest that assembly of the septin ring involves repeated cycles of GTP loading and GTP hydrolysis by Cdc42p. These results suggest that a single GTPase, Cdc42p, can act either as a ras-like GTP-dependent "switch" to turn on effectors or as an EF-Tu-like "assembly factor" using the GTPase cycle to assemble a macromolecular structure.

Show MeSH
Related in: MedlinePlus