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An anillin homologue, Mid2p, acts during fission yeast cytokinesis to organize the septin ring and promote cell separation.

Tasto JJ, Morrell JL, Gould KL - J. Cell Biol. (2003)

Bottom Line: Simanis. 1996.Mid2p colocalizes with septins, and mid2 Delta cells display disorganized, diffuse septin rings and a cell separation defect similar to septin deletion strains. mid2 gene expression and protein levels fluctuate during the cell cycle in a sep1- and Skp1/Cdc53/F-box (SCF)-dependent manner, respectively, implying that Mid2p activity must be carefully regulated.Overproduction of Mid2p depolarizes cell growth and affects the organization of both the septin and actin cytoskeletons.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.

ABSTRACT
Anillin is a conserved protein required for cell division (Field, C.M., and B.M. Alberts. 1995. J. Cell Biol. 131:165-178; Oegema, K., M.S. Savoian, T.J. Mitchison, and C.M. Field. 2000. J. Cell Biol. 150:539-552). One fission yeast homologue of anillin, Mid1p, is necessary for the proper placement of the division site within the cell (Chang, F., A. Woollard, and P. Nurse. 1996. J. Cell Sci. 109(Pt 1):131-142; Sohrmann, M., C. Fankhauser, C. Brodbeck, and V. Simanis. 1996. Genes Dev. 10:2707-2719). Here, we identify and characterize a second fission yeast anillin homologue, Mid2p, which is not orthologous with Mid1p. Mid2p localizes as a single ring in the middle of the cell after anaphase in a septin- and actin-dependent manner and splits into two rings during septation. Mid2p colocalizes with septins, and mid2 Delta cells display disorganized, diffuse septin rings and a cell separation defect similar to septin deletion strains. mid2 gene expression and protein levels fluctuate during the cell cycle in a sep1- and Skp1/Cdc53/F-box (SCF)-dependent manner, respectively, implying that Mid2p activity must be carefully regulated. Overproduction of Mid2p depolarizes cell growth and affects the organization of both the septin and actin cytoskeletons. In the presence of a nondegradable Mid2p fragment, the septin ring is stabilized and cell cycle progression is delayed. These results suggest that Mid2p influences septin ring organization at the site of cell division and its turnover might normally be required to permit septin ring disassembly.

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mid2 expression depends on Sep1p. (A) Differential interference contrast images of mid2Δ (KGY3135) and sep1Δ (KGY3419) cells grown in YE medium at 32°C. Bar, 5 μm. (B–E) cdc25-22 mid2–HA (KGY3306) (B and D) and cdc25-22 mid2–HA sep1Δ (KGY3457) (C and E) cells were arrested in G2 by shift to 36°C for 4 h. Cultures were then released to 25°C and cell pellets collected at the indicated times to analyze Mid2p–HA and mid2+ transcript levels. (B and C) Mid2p–HA and Cdc2p levels were determined by immunoblotting with 12CA5 and anti-PSTAIR, respectively. Mitotic progression was determined by assaying Cdc13p fluctuations by immunoblotting. (D and E) Northern blot analysis of mid2 transcript levels from the same time points analyzed in B and C. The his3 transcript served as a loading control.
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fig4: mid2 expression depends on Sep1p. (A) Differential interference contrast images of mid2Δ (KGY3135) and sep1Δ (KGY3419) cells grown in YE medium at 32°C. Bar, 5 μm. (B–E) cdc25-22 mid2–HA (KGY3306) (B and D) and cdc25-22 mid2–HA sep1Δ (KGY3457) (C and E) cells were arrested in G2 by shift to 36°C for 4 h. Cultures were then released to 25°C and cell pellets collected at the indicated times to analyze Mid2p–HA and mid2+ transcript levels. (B and C) Mid2p–HA and Cdc2p levels were determined by immunoblotting with 12CA5 and anti-PSTAIR, respectively. Mitotic progression was determined by assaying Cdc13p fluctuations by immunoblotting. (D and E) Northern blot analysis of mid2 transcript levels from the same time points analyzed in B and C. The his3 transcript served as a loading control.

Mentions: The fact that mid2 mRNA levels oscillate during vegetative growth (Fig. 2 E), coupled with the observation that the mid2Δ cell separation defect resembles the loss of a putative transcription factor, sep1 (Fig. 4 A; Ribar et al., 1999), prompted us to examine whether mid2 transcript abundance depended on Sep1p function. A cdc25-22 block and release synchronization protocol (Moreno et al., 1990) was used to address this question because the sep1Δ strain cannot be synchronized by centrifugal elutriation due to its chained cell phenotype (Ribar et al., 1999). In these cells, progression through mitosis was monitored by examining fluctuations in Cdc13p–cyclin B levels that occur during this period (Alfa et al., 1989). Although readily detectable in cdc25-22 cells proceeding through mitosis (Fig. 4 B), Mid2p–HA levels were clearly diminished in sep1Δ cells during the same time course (Fig. 4 C). The drop in Mid2p levels correlated with a significant decrease in mid2 mRNA abundance in the absence of sep1, although delayed and reduced amounts of the mid2 transcript were detected (Fig. 4, D and E). These results indicate that Sep1p plays an important role in setting Mid2p levels.


An anillin homologue, Mid2p, acts during fission yeast cytokinesis to organize the septin ring and promote cell separation.

Tasto JJ, Morrell JL, Gould KL - J. Cell Biol. (2003)

mid2 expression depends on Sep1p. (A) Differential interference contrast images of mid2Δ (KGY3135) and sep1Δ (KGY3419) cells grown in YE medium at 32°C. Bar, 5 μm. (B–E) cdc25-22 mid2–HA (KGY3306) (B and D) and cdc25-22 mid2–HA sep1Δ (KGY3457) (C and E) cells were arrested in G2 by shift to 36°C for 4 h. Cultures were then released to 25°C and cell pellets collected at the indicated times to analyze Mid2p–HA and mid2+ transcript levels. (B and C) Mid2p–HA and Cdc2p levels were determined by immunoblotting with 12CA5 and anti-PSTAIR, respectively. Mitotic progression was determined by assaying Cdc13p fluctuations by immunoblotting. (D and E) Northern blot analysis of mid2 transcript levels from the same time points analyzed in B and C. The his3 transcript served as a loading control.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172762&req=5

fig4: mid2 expression depends on Sep1p. (A) Differential interference contrast images of mid2Δ (KGY3135) and sep1Δ (KGY3419) cells grown in YE medium at 32°C. Bar, 5 μm. (B–E) cdc25-22 mid2–HA (KGY3306) (B and D) and cdc25-22 mid2–HA sep1Δ (KGY3457) (C and E) cells were arrested in G2 by shift to 36°C for 4 h. Cultures were then released to 25°C and cell pellets collected at the indicated times to analyze Mid2p–HA and mid2+ transcript levels. (B and C) Mid2p–HA and Cdc2p levels were determined by immunoblotting with 12CA5 and anti-PSTAIR, respectively. Mitotic progression was determined by assaying Cdc13p fluctuations by immunoblotting. (D and E) Northern blot analysis of mid2 transcript levels from the same time points analyzed in B and C. The his3 transcript served as a loading control.
Mentions: The fact that mid2 mRNA levels oscillate during vegetative growth (Fig. 2 E), coupled with the observation that the mid2Δ cell separation defect resembles the loss of a putative transcription factor, sep1 (Fig. 4 A; Ribar et al., 1999), prompted us to examine whether mid2 transcript abundance depended on Sep1p function. A cdc25-22 block and release synchronization protocol (Moreno et al., 1990) was used to address this question because the sep1Δ strain cannot be synchronized by centrifugal elutriation due to its chained cell phenotype (Ribar et al., 1999). In these cells, progression through mitosis was monitored by examining fluctuations in Cdc13p–cyclin B levels that occur during this period (Alfa et al., 1989). Although readily detectable in cdc25-22 cells proceeding through mitosis (Fig. 4 B), Mid2p–HA levels were clearly diminished in sep1Δ cells during the same time course (Fig. 4 C). The drop in Mid2p levels correlated with a significant decrease in mid2 mRNA abundance in the absence of sep1, although delayed and reduced amounts of the mid2 transcript were detected (Fig. 4, D and E). These results indicate that Sep1p plays an important role in setting Mid2p levels.

Bottom Line: Simanis. 1996.Mid2p colocalizes with septins, and mid2 Delta cells display disorganized, diffuse septin rings and a cell separation defect similar to septin deletion strains. mid2 gene expression and protein levels fluctuate during the cell cycle in a sep1- and Skp1/Cdc53/F-box (SCF)-dependent manner, respectively, implying that Mid2p activity must be carefully regulated.Overproduction of Mid2p depolarizes cell growth and affects the organization of both the septin and actin cytoskeletons.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.

ABSTRACT
Anillin is a conserved protein required for cell division (Field, C.M., and B.M. Alberts. 1995. J. Cell Biol. 131:165-178; Oegema, K., M.S. Savoian, T.J. Mitchison, and C.M. Field. 2000. J. Cell Biol. 150:539-552). One fission yeast homologue of anillin, Mid1p, is necessary for the proper placement of the division site within the cell (Chang, F., A. Woollard, and P. Nurse. 1996. J. Cell Sci. 109(Pt 1):131-142; Sohrmann, M., C. Fankhauser, C. Brodbeck, and V. Simanis. 1996. Genes Dev. 10:2707-2719). Here, we identify and characterize a second fission yeast anillin homologue, Mid2p, which is not orthologous with Mid1p. Mid2p localizes as a single ring in the middle of the cell after anaphase in a septin- and actin-dependent manner and splits into two rings during septation. Mid2p colocalizes with septins, and mid2 Delta cells display disorganized, diffuse septin rings and a cell separation defect similar to septin deletion strains. mid2 gene expression and protein levels fluctuate during the cell cycle in a sep1- and Skp1/Cdc53/F-box (SCF)-dependent manner, respectively, implying that Mid2p activity must be carefully regulated. Overproduction of Mid2p depolarizes cell growth and affects the organization of both the septin and actin cytoskeletons. In the presence of a nondegradable Mid2p fragment, the septin ring is stabilized and cell cycle progression is delayed. These results suggest that Mid2p influences septin ring organization at the site of cell division and its turnover might normally be required to permit septin ring disassembly.

Show MeSH
Related in: MedlinePlus