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Regulation of Rho-GEF Rgf3 by the arrestin Art1 in fission yeast cytokinesis.

Davidson R, Laporte D, Wu JQ - Mol. Biol. Cell (2014)

Bottom Line: Rgf3 is an essential GEF for Rho1 GTPase in fission yeast.Biochemical experiments reveal that the Rgf3 C-terminus binds to Art1.Using an Rgf3 conditional mutant and mislocalization experiments, we found that Art1 and Rgf3 are interdependent for localization to the division site.

View Article: PubMed Central - PubMed

Affiliation: Graduate Program of Molecular, Cellular, and Developmental Biology, The Ohio State University, Columbus, OH 43210 Department of Molecular Genetics, The Ohio State University, Columbus, OH 43210.

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art1 mutants have defective division septa. (A–C) Cell lysis phenotype of art1 mutant cells. Arrowheads mark boomerang-shaped lysed cells. (A) Differential interference contrast images and (B) quantification of cell lysis in wt (strain JW81), art1-s34 (JW404), and art1∆ (JW3563) cells. (C) Time-lapse imaging of an art1∆ (JW3563) cell lysed during cell separation. (D and E) Micrographs (D) and quantification (E) of septa under EM in wt (JW81) and art1∆ (JW2543) cells. Insets in D show magnification of the cell wall region in the white box. Double-headed arrows mark cell wall thickness. (E) A thin septum means the cell wall at the division site is continuous but ≤ 50% the thickness of that in wt cells. Thin and uneven septum is the same as thin septum, except that the septum is discontinuous. Scale bars: A and C, 5 μm; D, 0.5 μm.
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Figure 1: art1 mutants have defective division septa. (A–C) Cell lysis phenotype of art1 mutant cells. Arrowheads mark boomerang-shaped lysed cells. (A) Differential interference contrast images and (B) quantification of cell lysis in wt (strain JW81), art1-s34 (JW404), and art1∆ (JW3563) cells. (C) Time-lapse imaging of an art1∆ (JW3563) cell lysed during cell separation. (D and E) Micrographs (D) and quantification (E) of septa under EM in wt (JW81) and art1∆ (JW2543) cells. Insets in D show magnification of the cell wall region in the white box. Double-headed arrows mark cell wall thickness. (E) A thin septum means the cell wall at the division site is continuous but ≤ 50% the thickness of that in wt cells. Thin and uneven septum is the same as thin septum, except that the septum is discontinuous. Scale bars: A and C, 5 μm; D, 0.5 μm.

Mentions: The art1-s34 mutant was isolated in a synthetic lethal/sick screen carried out in a septin-depletion strain along with several genes that are known or predicted components of the cell-integrity pathway (Wu et al., 2010). art1-s34 cells have a mild cell lysis defect (∼5% of asynchronous cells) when grown in minimal medium EMM5S at 30°C (Wu et al., 2010). However, when grown in rich YE5S medium at 25°C, ∼20% of art1-s34 cells lysed (Figure 1, A and B). To further study the role of Art1, we deleted its whole open reading frame and found that cell lysis was similar (∼22%) in art1∆ cells (Figure 1, A and B), which is consistent with the prediction that art1-s34 is a allele (Wu et al., 2010). The lysis phenotype of the art1 mutants resembled that of the Rho-GEF rgf3(lad1-1) mutant (Morrell-Falvey et al., 2005), in which the lysed daughter cells often attach to each other in the shape of “boomerangs” (Figure 1A, arrowheads). Time-lapse imaging of art1∆ cells revealed that lysis occurred during cell separation (Figure 1C and Supplemental Video S1). The lysis phenotype suggests that the plasma membrane and/or the septum at the division site are defective in the art1 mutants.


Regulation of Rho-GEF Rgf3 by the arrestin Art1 in fission yeast cytokinesis.

Davidson R, Laporte D, Wu JQ - Mol. Biol. Cell (2014)

art1 mutants have defective division septa. (A–C) Cell lysis phenotype of art1 mutant cells. Arrowheads mark boomerang-shaped lysed cells. (A) Differential interference contrast images and (B) quantification of cell lysis in wt (strain JW81), art1-s34 (JW404), and art1∆ (JW3563) cells. (C) Time-lapse imaging of an art1∆ (JW3563) cell lysed during cell separation. (D and E) Micrographs (D) and quantification (E) of septa under EM in wt (JW81) and art1∆ (JW2543) cells. Insets in D show magnification of the cell wall region in the white box. Double-headed arrows mark cell wall thickness. (E) A thin septum means the cell wall at the division site is continuous but ≤ 50% the thickness of that in wt cells. Thin and uneven septum is the same as thin septum, except that the septum is discontinuous. Scale bars: A and C, 5 μm; D, 0.5 μm.
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Related In: Results  -  Collection

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Figure 1: art1 mutants have defective division septa. (A–C) Cell lysis phenotype of art1 mutant cells. Arrowheads mark boomerang-shaped lysed cells. (A) Differential interference contrast images and (B) quantification of cell lysis in wt (strain JW81), art1-s34 (JW404), and art1∆ (JW3563) cells. (C) Time-lapse imaging of an art1∆ (JW3563) cell lysed during cell separation. (D and E) Micrographs (D) and quantification (E) of septa under EM in wt (JW81) and art1∆ (JW2543) cells. Insets in D show magnification of the cell wall region in the white box. Double-headed arrows mark cell wall thickness. (E) A thin septum means the cell wall at the division site is continuous but ≤ 50% the thickness of that in wt cells. Thin and uneven septum is the same as thin septum, except that the septum is discontinuous. Scale bars: A and C, 5 μm; D, 0.5 μm.
Mentions: The art1-s34 mutant was isolated in a synthetic lethal/sick screen carried out in a septin-depletion strain along with several genes that are known or predicted components of the cell-integrity pathway (Wu et al., 2010). art1-s34 cells have a mild cell lysis defect (∼5% of asynchronous cells) when grown in minimal medium EMM5S at 30°C (Wu et al., 2010). However, when grown in rich YE5S medium at 25°C, ∼20% of art1-s34 cells lysed (Figure 1, A and B). To further study the role of Art1, we deleted its whole open reading frame and found that cell lysis was similar (∼22%) in art1∆ cells (Figure 1, A and B), which is consistent with the prediction that art1-s34 is a allele (Wu et al., 2010). The lysis phenotype of the art1 mutants resembled that of the Rho-GEF rgf3(lad1-1) mutant (Morrell-Falvey et al., 2005), in which the lysed daughter cells often attach to each other in the shape of “boomerangs” (Figure 1A, arrowheads). Time-lapse imaging of art1∆ cells revealed that lysis occurred during cell separation (Figure 1C and Supplemental Video S1). The lysis phenotype suggests that the plasma membrane and/or the septum at the division site are defective in the art1 mutants.

Bottom Line: Rgf3 is an essential GEF for Rho1 GTPase in fission yeast.Biochemical experiments reveal that the Rgf3 C-terminus binds to Art1.Using an Rgf3 conditional mutant and mislocalization experiments, we found that Art1 and Rgf3 are interdependent for localization to the division site.

View Article: PubMed Central - PubMed

Affiliation: Graduate Program of Molecular, Cellular, and Developmental Biology, The Ohio State University, Columbus, OH 43210 Department of Molecular Genetics, The Ohio State University, Columbus, OH 43210.

Show MeSH
Related in: MedlinePlus