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cdc12p, a protein required for cytokinesis in fission yeast, is a component of the cell division ring and interacts with profilin.

Chang F, Drubin D, Nurse P - J. Cell Biol. (1997)

Bottom Line: Here we show that cdc12p is a member of a family of proteins including Drosophila diaphanous, Saccharomyces cerevisiae BNI1, and S. pombe fus1, which are involved in cytokinesis or other actin-mediated processes.Based on these patterns, we present a model in which ring assembly originates from a single point on the cortex and in which a molecular pathway for the functions of cytokinesis proteins is suggested.Finally, we found that cdc12 and cdc3 mutants show a synthetic-lethal genetic interaction, and a proline-rich domain of cdc12p binds directly to profilin cdc3p in vitro, suggesting that one function of cdc12p in ring assembly is to bind profilin.

View Article: PubMed Central - PubMed

Affiliation: Imperial Cancer Research Fund, London, United Kingdom. fc99@columbia.edu

ABSTRACT
As in many other eukaryotic cells, cell division in fission yeast depends on the assembly of an actin ring that circumscribes the middle of the cell. Schizosaccharomyces pombe cdc12 is an essential gene necessary for actin ring assembly and septum formation. Here we show that cdc12p is a member of a family of proteins including Drosophila diaphanous, Saccharomyces cerevisiae BNI1, and S. pombe fus1, which are involved in cytokinesis or other actin-mediated processes. Using indirect immunofluorescence, we show that cdc12p is located in the cell division ring and not in other actin structures. When overexpressed, cdc12p is located at a medial spot in interphase that anticipates the future ring site. cdc12p localization is altered in actin ring mutants. cdc8 (tropomyosin homologue), cdc3 (profilin homologue), and cdc15 mutants exhibit no specific cdc12p staining during mitosis. cdc4 mutant cells exhibit a medial cortical cdc12p spot in place of a ring. mid1 mutant cells generally exhibit a cdc12p spot with a single cdc12p strand extending in a random direction. Based on these patterns, we present a model in which ring assembly originates from a single point on the cortex and in which a molecular pathway for the functions of cytokinesis proteins is suggested. Finally, we found that cdc12 and cdc3 mutants show a synthetic-lethal genetic interaction, and a proline-rich domain of cdc12p binds directly to profilin cdc3p in vitro, suggesting that one function of cdc12p in ring assembly is to bind profilin.

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The positions of the cdc12p spot and the ring coincide. FC565 (pnmt-cdc12) cells were processed as described in Fig. 4. Shown  are four representative cells in early mitosis, around the period of ring formation. Top three panels show three different focal planes of  the same cell, and the bottom two panels show tubulin and DAPI staining as labeled. (A) Cell with a cdc12p spot. (B) Cell with a cdc12p  associated with a complete ring. (C) Cell with a cdc12p spot with an incomplete ring. Note the discontinuity of the ring adjacent to the  spot in the top panel. (D) Cell with a cdc12p ring but no spot. Bar, 10 μm.
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Figure 5: The positions of the cdc12p spot and the ring coincide. FC565 (pnmt-cdc12) cells were processed as described in Fig. 4. Shown are four representative cells in early mitosis, around the period of ring formation. Top three panels show three different focal planes of the same cell, and the bottom two panels show tubulin and DAPI staining as labeled. (A) Cell with a cdc12p spot. (B) Cell with a cdc12p associated with a complete ring. (C) Cell with a cdc12p spot with an incomplete ring. Note the discontinuity of the ring adjacent to the spot in the top panel. (D) Cell with a cdc12p ring but no spot. Bar, 10 μm.

Mentions: Immunofluorescence using this anti-cdc12p antibody revealed staining of the fission yeast cell division ring (Fig. 3). cdc12p was present in early mitotic cells as a thin, faint ring (Fig. 3 D). This ring was more intense during anaphase (Fig. 3 H) and, after completion of anaphase, appeared to contract to a single dot between the daughter cells during cytokinesis (Fig. 3 J). cdc12p staining was very similar to staining patterns of other components of the ring such as actin and cdc4p (Marks and Hyams, 1985; McCollum et al., 1995). cdc12p did not localize to other actin structures, such as interphase actin patches at the ends of cells or actin dots adjacent to the septum during septation (Marks and Hyams, 1985). The cdc12p staining was specific because there was no ring staining in cdc12-112 mutant cells (see Fig. 5 A), and, in addition, a green fluorescent protein–cdc12 fusion is located at similar structures (Chang, F., unpublished observations).


cdc12p, a protein required for cytokinesis in fission yeast, is a component of the cell division ring and interacts with profilin.

Chang F, Drubin D, Nurse P - J. Cell Biol. (1997)

The positions of the cdc12p spot and the ring coincide. FC565 (pnmt-cdc12) cells were processed as described in Fig. 4. Shown  are four representative cells in early mitosis, around the period of ring formation. Top three panels show three different focal planes of  the same cell, and the bottom two panels show tubulin and DAPI staining as labeled. (A) Cell with a cdc12p spot. (B) Cell with a cdc12p  associated with a complete ring. (C) Cell with a cdc12p spot with an incomplete ring. Note the discontinuity of the ring adjacent to the  spot in the top panel. (D) Cell with a cdc12p ring but no spot. Bar, 10 μm.
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Related In: Results  -  Collection

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Figure 5: The positions of the cdc12p spot and the ring coincide. FC565 (pnmt-cdc12) cells were processed as described in Fig. 4. Shown are four representative cells in early mitosis, around the period of ring formation. Top three panels show three different focal planes of the same cell, and the bottom two panels show tubulin and DAPI staining as labeled. (A) Cell with a cdc12p spot. (B) Cell with a cdc12p associated with a complete ring. (C) Cell with a cdc12p spot with an incomplete ring. Note the discontinuity of the ring adjacent to the spot in the top panel. (D) Cell with a cdc12p ring but no spot. Bar, 10 μm.
Mentions: Immunofluorescence using this anti-cdc12p antibody revealed staining of the fission yeast cell division ring (Fig. 3). cdc12p was present in early mitotic cells as a thin, faint ring (Fig. 3 D). This ring was more intense during anaphase (Fig. 3 H) and, after completion of anaphase, appeared to contract to a single dot between the daughter cells during cytokinesis (Fig. 3 J). cdc12p staining was very similar to staining patterns of other components of the ring such as actin and cdc4p (Marks and Hyams, 1985; McCollum et al., 1995). cdc12p did not localize to other actin structures, such as interphase actin patches at the ends of cells or actin dots adjacent to the septum during septation (Marks and Hyams, 1985). The cdc12p staining was specific because there was no ring staining in cdc12-112 mutant cells (see Fig. 5 A), and, in addition, a green fluorescent protein–cdc12 fusion is located at similar structures (Chang, F., unpublished observations).

Bottom Line: Here we show that cdc12p is a member of a family of proteins including Drosophila diaphanous, Saccharomyces cerevisiae BNI1, and S. pombe fus1, which are involved in cytokinesis or other actin-mediated processes.Based on these patterns, we present a model in which ring assembly originates from a single point on the cortex and in which a molecular pathway for the functions of cytokinesis proteins is suggested.Finally, we found that cdc12 and cdc3 mutants show a synthetic-lethal genetic interaction, and a proline-rich domain of cdc12p binds directly to profilin cdc3p in vitro, suggesting that one function of cdc12p in ring assembly is to bind profilin.

View Article: PubMed Central - PubMed

Affiliation: Imperial Cancer Research Fund, London, United Kingdom. fc99@columbia.edu

ABSTRACT
As in many other eukaryotic cells, cell division in fission yeast depends on the assembly of an actin ring that circumscribes the middle of the cell. Schizosaccharomyces pombe cdc12 is an essential gene necessary for actin ring assembly and septum formation. Here we show that cdc12p is a member of a family of proteins including Drosophila diaphanous, Saccharomyces cerevisiae BNI1, and S. pombe fus1, which are involved in cytokinesis or other actin-mediated processes. Using indirect immunofluorescence, we show that cdc12p is located in the cell division ring and not in other actin structures. When overexpressed, cdc12p is located at a medial spot in interphase that anticipates the future ring site. cdc12p localization is altered in actin ring mutants. cdc8 (tropomyosin homologue), cdc3 (profilin homologue), and cdc15 mutants exhibit no specific cdc12p staining during mitosis. cdc4 mutant cells exhibit a medial cortical cdc12p spot in place of a ring. mid1 mutant cells generally exhibit a cdc12p spot with a single cdc12p strand extending in a random direction. Based on these patterns, we present a model in which ring assembly originates from a single point on the cortex and in which a molecular pathway for the functions of cytokinesis proteins is suggested. Finally, we found that cdc12 and cdc3 mutants show a synthetic-lethal genetic interaction, and a proline-rich domain of cdc12p binds directly to profilin cdc3p in vitro, suggesting that one function of cdc12p in ring assembly is to bind profilin.

Show MeSH
Related in: MedlinePlus