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Lack of GTP-bound Rho1p in secretory vesicles of Saccharomyces cerevisiae.

Abe M, Qadota H, Hirata A, Ohya Y - J. Cell Biol. (2003)

Bottom Line: Rho1p, an essential Rho-type GTPase in Saccharomyces cerevisiae, activates its effectors in the GTP-bound form.Rom2p, the GDP/GTP exchange factor of Rho1p, is preferentially localized on the plasma membrane even when vesicular transport is blocked.Based on these results, we propose that Rho1p is kept inactive in intracellular secretory organelles, resulting in repression of the activity of the cell wall-synthesizing enzyme within cells.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Integrated Biosciences, Graduate School of Frontier Sciences, University of Tokyo, FSB-101, 5-1-5 Kashiwanoha, Kashiwa, Chiba 277-8562, Japan.

ABSTRACT
Rho1p, an essential Rho-type GTPase in Saccharomyces cerevisiae, activates its effectors in the GTP-bound form. Here, we show that Rho1p in secretory vesicles cannot activate 1,3-beta-glucan synthase, a cell wall synthesizing enzyme, during vesicular transport to the plasma membrane. Analyses with an antibody preferentially reacting with the GTP-bound form of Rho1p revealed that Rho1p remains in the inactive form in secretory vesicles. Rom2p, the GDP/GTP exchange factor of Rho1p, is preferentially localized on the plasma membrane even when vesicular transport is blocked. Overexpression of Rom2p results in delocalization of Rom2p and accumulation of 1,3-beta-glucan in secretory vesicles. Based on these results, we propose that Rho1p is kept inactive in intracellular secretory organelles, resulting in repression of the activity of the cell wall-synthesizing enzyme within cells.

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Effect of activatedRHO1overexpression on the growth of wild-type and sec mutant cells. (A) Inhibition of cell proliferation in sec1 and sec6 mutant cells. Wild-type (YPH500), sec1, and sec6 cells were transformed with a control vector, or YEpU-RHO1 (G19V), and incubated for 3 d at 31°C. [RHO1 (G19V)] indicates the transformants with YEpU-RHO1 (G19V). (B) Suppression of inhibition by disturbing 1,3-β-glucan synthesis. sec1 fks1–1154 fks2Δ cells were transformed with a control vector, YEpT-RHO1 (G19V), or YCpU-FKS1, and incubated for 3 d at 31°C. [RHO1 (G19V)] and [FKS1] indicate the transformants with YEpU-RHO1 (G19V) and YCpU-FKS1, respectively.
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fig9: Effect of activatedRHO1overexpression on the growth of wild-type and sec mutant cells. (A) Inhibition of cell proliferation in sec1 and sec6 mutant cells. Wild-type (YPH500), sec1, and sec6 cells were transformed with a control vector, or YEpU-RHO1 (G19V), and incubated for 3 d at 31°C. [RHO1 (G19V)] indicates the transformants with YEpU-RHO1 (G19V). (B) Suppression of inhibition by disturbing 1,3-β-glucan synthesis. sec1 fks1–1154 fks2Δ cells were transformed with a control vector, YEpT-RHO1 (G19V), or YCpU-FKS1, and incubated for 3 d at 31°C. [RHO1 (G19V)] and [FKS1] indicate the transformants with YEpU-RHO1 (G19V) and YCpU-FKS1, respectively.

Mentions: To test the possibility that the activation of Rho1p in secretory vesicles affects cell proliferation, we examined the growth phenotypes of sec1 and sec6 cells overexpressing the activated form of RHO1. Overexpression of the activated form of RHO1 led to growth inhibition of sec1 and sec6 cells at 31°C, a semipermissive temperature at which the temperature-sensitive sec mutants can grow (Fig. 9Figure 9.


Lack of GTP-bound Rho1p in secretory vesicles of Saccharomyces cerevisiae.

Abe M, Qadota H, Hirata A, Ohya Y - J. Cell Biol. (2003)

Effect of activatedRHO1overexpression on the growth of wild-type and sec mutant cells. (A) Inhibition of cell proliferation in sec1 and sec6 mutant cells. Wild-type (YPH500), sec1, and sec6 cells were transformed with a control vector, or YEpU-RHO1 (G19V), and incubated for 3 d at 31°C. [RHO1 (G19V)] indicates the transformants with YEpU-RHO1 (G19V). (B) Suppression of inhibition by disturbing 1,3-β-glucan synthesis. sec1 fks1–1154 fks2Δ cells were transformed with a control vector, YEpT-RHO1 (G19V), or YCpU-FKS1, and incubated for 3 d at 31°C. [RHO1 (G19V)] and [FKS1] indicate the transformants with YEpU-RHO1 (G19V) and YCpU-FKS1, respectively.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172714&req=5

fig9: Effect of activatedRHO1overexpression on the growth of wild-type and sec mutant cells. (A) Inhibition of cell proliferation in sec1 and sec6 mutant cells. Wild-type (YPH500), sec1, and sec6 cells were transformed with a control vector, or YEpU-RHO1 (G19V), and incubated for 3 d at 31°C. [RHO1 (G19V)] indicates the transformants with YEpU-RHO1 (G19V). (B) Suppression of inhibition by disturbing 1,3-β-glucan synthesis. sec1 fks1–1154 fks2Δ cells were transformed with a control vector, YEpT-RHO1 (G19V), or YCpU-FKS1, and incubated for 3 d at 31°C. [RHO1 (G19V)] and [FKS1] indicate the transformants with YEpU-RHO1 (G19V) and YCpU-FKS1, respectively.
Mentions: To test the possibility that the activation of Rho1p in secretory vesicles affects cell proliferation, we examined the growth phenotypes of sec1 and sec6 cells overexpressing the activated form of RHO1. Overexpression of the activated form of RHO1 led to growth inhibition of sec1 and sec6 cells at 31°C, a semipermissive temperature at which the temperature-sensitive sec mutants can grow (Fig. 9Figure 9.

Bottom Line: Rho1p, an essential Rho-type GTPase in Saccharomyces cerevisiae, activates its effectors in the GTP-bound form.Rom2p, the GDP/GTP exchange factor of Rho1p, is preferentially localized on the plasma membrane even when vesicular transport is blocked.Based on these results, we propose that Rho1p is kept inactive in intracellular secretory organelles, resulting in repression of the activity of the cell wall-synthesizing enzyme within cells.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Integrated Biosciences, Graduate School of Frontier Sciences, University of Tokyo, FSB-101, 5-1-5 Kashiwanoha, Kashiwa, Chiba 277-8562, Japan.

ABSTRACT
Rho1p, an essential Rho-type GTPase in Saccharomyces cerevisiae, activates its effectors in the GTP-bound form. Here, we show that Rho1p in secretory vesicles cannot activate 1,3-beta-glucan synthase, a cell wall synthesizing enzyme, during vesicular transport to the plasma membrane. Analyses with an antibody preferentially reacting with the GTP-bound form of Rho1p revealed that Rho1p remains in the inactive form in secretory vesicles. Rom2p, the GDP/GTP exchange factor of Rho1p, is preferentially localized on the plasma membrane even when vesicular transport is blocked. Overexpression of Rom2p results in delocalization of Rom2p and accumulation of 1,3-beta-glucan in secretory vesicles. Based on these results, we propose that Rho1p is kept inactive in intracellular secretory organelles, resulting in repression of the activity of the cell wall-synthesizing enzyme within cells.

Show MeSH
Related in: MedlinePlus