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Assembly of normal actomyosin rings in the absence of Mid1p and cortical nodes in fission yeast.

Huang Y, Yan H, Balasubramanian MK - J. Cell Biol. (2008)

Bottom Line: In this study, we test this model in cells that are unable to assemble cortical nodes.Although the cortical nodes play a role in the timing of ring assembly, we find that they are dispensable for the assembly of orthogonal actomyosin rings.Thus, a mechanism that is independent of cortical nodes is sufficient for the assembly of normal actomyosin rings.

View Article: PubMed Central - PubMed

Affiliation: Temasek Life Sciences Laboratory, National University of Singapore, Singapore 117604.

ABSTRACT
Cytokinesis in many eukaryotes depends on the function of an actomyosin contractile ring. The mechanisms regulating assembly and positioning of this ring are not fully understood. The fission yeast Schizosaccharomyces pombe divides using an actomyosin ring and is an attractive organism for the study of cytokinesis. Recent studies in S. pombe (Wu, J.Q., V. Sirotkin, D.R. Kovar, M. Lord, C.C. Beltzner, J.R. Kuhn, and T.D. Pollard. 2006. J. Cell Biol. 174:391-402; Vavylonis, D., J.Q. Wu, S. Hao, B. O'Shaughnessy, and T.D. Pollard. 2008. Science. 319:97-100) have suggested that the assembly of the actomyosin ring is initiated from a series of cortical nodes containing several components of this ring. These studies have proposed that actomyosin interactions bring together the cortical nodes to form a compacted ring structure. In this study, we test this model in cells that are unable to assemble cortical nodes. Although the cortical nodes play a role in the timing of ring assembly, we find that they are dispensable for the assembly of orthogonal actomyosin rings. Thus, a mechanism that is independent of cortical nodes is sufficient for the assembly of normal actomyosin rings.

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Orthogonal actomyosin rings assemble with high efficiency in mid1 and plo1 mutants. (A and B) Cells of the indicated genotypes were shifted to 36°C, fixed, and stained with Alexa Fluor 488 phalloidin to visualize F-actin rings. (C) Quantitation of cells with orthogonal rings. At least 500 cells were scored for each genotype. (D) Nodes are not restored in cps1-191 mid1Δ mutants. cps1-191 and cps1-191 mid1Δ cells were fixed and stained with Tat1 antibodies (tubulin), Cdc4p antibodies, and DAPI (nuclei). (E) Randomly oriented myosin II bundles eventually organize into orthogonal myosin II rings in cps1-191 mid1-18 cells. cps1-191 mid1-18 cells expressing Rlc1p-GFP were imaged by confocal microscopy at 36°C. The elapsed time is shown in minutes. wt, wild type. Bars, 5 μm.
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fig2: Orthogonal actomyosin rings assemble with high efficiency in mid1 and plo1 mutants. (A and B) Cells of the indicated genotypes were shifted to 36°C, fixed, and stained with Alexa Fluor 488 phalloidin to visualize F-actin rings. (C) Quantitation of cells with orthogonal rings. At least 500 cells were scored for each genotype. (D) Nodes are not restored in cps1-191 mid1Δ mutants. cps1-191 and cps1-191 mid1Δ cells were fixed and stained with Tat1 antibodies (tubulin), Cdc4p antibodies, and DAPI (nuclei). (E) Randomly oriented myosin II bundles eventually organize into orthogonal myosin II rings in cps1-191 mid1-18 cells. cps1-191 mid1-18 cells expressing Rlc1p-GFP were imaged by confocal microscopy at 36°C. The elapsed time is shown in minutes. wt, wild type. Bars, 5 μm.

Mentions: Cells defective for Mid1p or Plo1p have been shown to assemble abnormal actomyosin rings (Chang et al., 1996; Sohrmann et al., 1996; Bahler et al., 1998), which at first glance is consistent with the predictions of recent studies (Wu et al., 2006; Vavylonis et al., 2008). We have found that the abnormal rings in mid1 and plo1 mutant cells can become stabilized as a result of septum assembly before ring compaction. Fig. 2 A shows examples of F-actin distribution in wild type, cells of three different mutant alleles of mid1 (mid1-18, mid1-ΔNES, and mid1Δ), and plo1-1 cells. Unlike in mitotic wild-type cells, F-actin rings in mitotic mid1 and plo1-1 mutants are detected at various angles (Fig. 2 A; Bahler et al., 1998). To eliminate any confusion in scoring phenotypic effects that might arise from the assembly of division septum along improperly organized rings, we inactivated Cps1p, a 1,3-β-glucan synthase involved in septum assembly (Liu et al., 1999), in cells defective in Mid1p or Plo1p function. Surprisingly, >85% of cps1-191 mid1-18, cps1-191 mid1-ΔNES, cps1-191 mid1Δ, and cps1-191 plo1-1 cells assembled normal and orthogonal actomyosin rings (Fig. 2, B and C). Rings in cps1-191 mid1-18, cps1-191 mid1-ΔNES, cps1-191 mid1Δ, and cps1-191 plo1-1 cells were positioned at abnormal sites, which is consistent with a role for Mid1p and Plo1p in actomyosin ring positioning.


Assembly of normal actomyosin rings in the absence of Mid1p and cortical nodes in fission yeast.

Huang Y, Yan H, Balasubramanian MK - J. Cell Biol. (2008)

Orthogonal actomyosin rings assemble with high efficiency in mid1 and plo1 mutants. (A and B) Cells of the indicated genotypes were shifted to 36°C, fixed, and stained with Alexa Fluor 488 phalloidin to visualize F-actin rings. (C) Quantitation of cells with orthogonal rings. At least 500 cells were scored for each genotype. (D) Nodes are not restored in cps1-191 mid1Δ mutants. cps1-191 and cps1-191 mid1Δ cells were fixed and stained with Tat1 antibodies (tubulin), Cdc4p antibodies, and DAPI (nuclei). (E) Randomly oriented myosin II bundles eventually organize into orthogonal myosin II rings in cps1-191 mid1-18 cells. cps1-191 mid1-18 cells expressing Rlc1p-GFP were imaged by confocal microscopy at 36°C. The elapsed time is shown in minutes. wt, wild type. Bars, 5 μm.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2600752&req=5

fig2: Orthogonal actomyosin rings assemble with high efficiency in mid1 and plo1 mutants. (A and B) Cells of the indicated genotypes were shifted to 36°C, fixed, and stained with Alexa Fluor 488 phalloidin to visualize F-actin rings. (C) Quantitation of cells with orthogonal rings. At least 500 cells were scored for each genotype. (D) Nodes are not restored in cps1-191 mid1Δ mutants. cps1-191 and cps1-191 mid1Δ cells were fixed and stained with Tat1 antibodies (tubulin), Cdc4p antibodies, and DAPI (nuclei). (E) Randomly oriented myosin II bundles eventually organize into orthogonal myosin II rings in cps1-191 mid1-18 cells. cps1-191 mid1-18 cells expressing Rlc1p-GFP were imaged by confocal microscopy at 36°C. The elapsed time is shown in minutes. wt, wild type. Bars, 5 μm.
Mentions: Cells defective for Mid1p or Plo1p have been shown to assemble abnormal actomyosin rings (Chang et al., 1996; Sohrmann et al., 1996; Bahler et al., 1998), which at first glance is consistent with the predictions of recent studies (Wu et al., 2006; Vavylonis et al., 2008). We have found that the abnormal rings in mid1 and plo1 mutant cells can become stabilized as a result of septum assembly before ring compaction. Fig. 2 A shows examples of F-actin distribution in wild type, cells of three different mutant alleles of mid1 (mid1-18, mid1-ΔNES, and mid1Δ), and plo1-1 cells. Unlike in mitotic wild-type cells, F-actin rings in mitotic mid1 and plo1-1 mutants are detected at various angles (Fig. 2 A; Bahler et al., 1998). To eliminate any confusion in scoring phenotypic effects that might arise from the assembly of division septum along improperly organized rings, we inactivated Cps1p, a 1,3-β-glucan synthase involved in septum assembly (Liu et al., 1999), in cells defective in Mid1p or Plo1p function. Surprisingly, >85% of cps1-191 mid1-18, cps1-191 mid1-ΔNES, cps1-191 mid1Δ, and cps1-191 plo1-1 cells assembled normal and orthogonal actomyosin rings (Fig. 2, B and C). Rings in cps1-191 mid1-18, cps1-191 mid1-ΔNES, cps1-191 mid1Δ, and cps1-191 plo1-1 cells were positioned at abnormal sites, which is consistent with a role for Mid1p and Plo1p in actomyosin ring positioning.

Bottom Line: In this study, we test this model in cells that are unable to assemble cortical nodes.Although the cortical nodes play a role in the timing of ring assembly, we find that they are dispensable for the assembly of orthogonal actomyosin rings.Thus, a mechanism that is independent of cortical nodes is sufficient for the assembly of normal actomyosin rings.

View Article: PubMed Central - PubMed

Affiliation: Temasek Life Sciences Laboratory, National University of Singapore, Singapore 117604.

ABSTRACT
Cytokinesis in many eukaryotes depends on the function of an actomyosin contractile ring. The mechanisms regulating assembly and positioning of this ring are not fully understood. The fission yeast Schizosaccharomyces pombe divides using an actomyosin ring and is an attractive organism for the study of cytokinesis. Recent studies in S. pombe (Wu, J.Q., V. Sirotkin, D.R. Kovar, M. Lord, C.C. Beltzner, J.R. Kuhn, and T.D. Pollard. 2006. J. Cell Biol. 174:391-402; Vavylonis, D., J.Q. Wu, S. Hao, B. O'Shaughnessy, and T.D. Pollard. 2008. Science. 319:97-100) have suggested that the assembly of the actomyosin ring is initiated from a series of cortical nodes containing several components of this ring. These studies have proposed that actomyosin interactions bring together the cortical nodes to form a compacted ring structure. In this study, we test this model in cells that are unable to assemble cortical nodes. Although the cortical nodes play a role in the timing of ring assembly, we find that they are dispensable for the assembly of orthogonal actomyosin rings. Thus, a mechanism that is independent of cortical nodes is sufficient for the assembly of normal actomyosin rings.

Show MeSH
Related in: MedlinePlus