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The small GTP-binding protein rho regulates cortical activities in cultured cells during division.

O'Connell CB, Wheatley SP, Ahmed S, Wang YL - J. Cell Biol. (1999)

Bottom Line: All treated, well-adherent cells underwent cleavage-like activities and most of them divided successfully.The effects of C3 appeared to be dependent on cell adhesion; less adherent 3T3 fibroblasts exhibited irregular cortical ingression only when cells started to increase attachment during respreading, but managed to complete cytokinesis.Our results indicate that Rho does not simply activate actin-myosin II interactions during cytokinesis, but regulates the spatial pattern of cortical activities and completion of cytokinesis possibly through modulating the mechanical strength of the cortex.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, University of Massachusetts Medical School, Worcester, Massachusetts 01655, USA.

ABSTRACT
We have investigated the role of the small GTP-binding protein Rho in cytokinesis by microinjecting an inhibitor, C3 ribosyltransferase, into cultured cells. Microinjection of C3 into prometaphase or metaphase normal rat kidney epithelial cells induced immediate and global cortical movement of actin toward the metaphase plate, without an apparent effect on the mitotic spindle. During anaphase, concentrated cortical actin filaments migrated with separating chromosomes, leaving no apparent concentration of actin filaments along the equator. Myosin II in injected epithelial cells showed a diffuse distribution throughout cell division. All treated, well-adherent cells underwent cleavage-like activities and most of them divided successfully. However, cytokinesis became abnormal, generating irregular ingressions and ectopic cleavage sites even when mitosis was blocked with nocodazole. The effects of C3 appeared to be dependent on cell adhesion; less adherent 3T3 fibroblasts exhibited irregular cortical ingression only when cells started to increase attachment during respreading, but managed to complete cytokinesis. Poorly adherent HeLa cells showed neither ectopic cleavage nor completion of cytokinesis. Our results indicate that Rho does not simply activate actin-myosin II interactions during cytokinesis, but regulates the spatial pattern of cortical activities and completion of cytokinesis possibly through modulating the mechanical strength of the cortex.

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Time lapse images  of control (a–d) and C3- injected HeLa cells (e–h).  Approximate time in minutes  relative to anaphase onset is  located in the upper right  corner. Cells injected with  C3 were not able to complete  cytokinesis (h), although some  did attempt a weak furrow  that could not progress (g and  h). Control cells injected with  10 mM Hepes, pH 7.5, had no  difficulty completing division  (c and d). Bar, 20 μm.
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Figure 4: Time lapse images of control (a–d) and C3- injected HeLa cells (e–h). Approximate time in minutes relative to anaphase onset is located in the upper right corner. Cells injected with C3 were not able to complete cytokinesis (h), although some did attempt a weak furrow that could not progress (g and h). Control cells injected with 10 mM Hepes, pH 7.5, had no difficulty completing division (c and d). Bar, 20 μm.

Mentions: C3 was also microinjected into HeLa cells that are very weakly adherent to the substrate and their neighbors (Fig. 4). A large fraction of injected HeLa cells either detached from the dish during microinjection or failed to initiate anaphase. Of the limited number of cells injected with C3 that proceeded into anaphase, which we took as an indication of proper injection, all failed cytokinesis (n = 5; Fig. 4, g and h). Some of them showed clear signs of the initiation of cleavage, but never made further progress (n = 2; Fig. 4, g and h). HeLa cells that were microinjected with buffer alone were able to cleave successfully (Fig. 4 d).


The small GTP-binding protein rho regulates cortical activities in cultured cells during division.

O'Connell CB, Wheatley SP, Ahmed S, Wang YL - J. Cell Biol. (1999)

Time lapse images  of control (a–d) and C3- injected HeLa cells (e–h).  Approximate time in minutes  relative to anaphase onset is  located in the upper right  corner. Cells injected with  C3 were not able to complete  cytokinesis (h), although some  did attempt a weak furrow  that could not progress (g and  h). Control cells injected with  10 mM Hepes, pH 7.5, had no  difficulty completing division  (c and d). Bar, 20 μm.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2132903&req=5

Figure 4: Time lapse images of control (a–d) and C3- injected HeLa cells (e–h). Approximate time in minutes relative to anaphase onset is located in the upper right corner. Cells injected with C3 were not able to complete cytokinesis (h), although some did attempt a weak furrow that could not progress (g and h). Control cells injected with 10 mM Hepes, pH 7.5, had no difficulty completing division (c and d). Bar, 20 μm.
Mentions: C3 was also microinjected into HeLa cells that are very weakly adherent to the substrate and their neighbors (Fig. 4). A large fraction of injected HeLa cells either detached from the dish during microinjection or failed to initiate anaphase. Of the limited number of cells injected with C3 that proceeded into anaphase, which we took as an indication of proper injection, all failed cytokinesis (n = 5; Fig. 4, g and h). Some of them showed clear signs of the initiation of cleavage, but never made further progress (n = 2; Fig. 4, g and h). HeLa cells that were microinjected with buffer alone were able to cleave successfully (Fig. 4 d).

Bottom Line: All treated, well-adherent cells underwent cleavage-like activities and most of them divided successfully.The effects of C3 appeared to be dependent on cell adhesion; less adherent 3T3 fibroblasts exhibited irregular cortical ingression only when cells started to increase attachment during respreading, but managed to complete cytokinesis.Our results indicate that Rho does not simply activate actin-myosin II interactions during cytokinesis, but regulates the spatial pattern of cortical activities and completion of cytokinesis possibly through modulating the mechanical strength of the cortex.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, University of Massachusetts Medical School, Worcester, Massachusetts 01655, USA.

ABSTRACT
We have investigated the role of the small GTP-binding protein Rho in cytokinesis by microinjecting an inhibitor, C3 ribosyltransferase, into cultured cells. Microinjection of C3 into prometaphase or metaphase normal rat kidney epithelial cells induced immediate and global cortical movement of actin toward the metaphase plate, without an apparent effect on the mitotic spindle. During anaphase, concentrated cortical actin filaments migrated with separating chromosomes, leaving no apparent concentration of actin filaments along the equator. Myosin II in injected epithelial cells showed a diffuse distribution throughout cell division. All treated, well-adherent cells underwent cleavage-like activities and most of them divided successfully. However, cytokinesis became abnormal, generating irregular ingressions and ectopic cleavage sites even when mitosis was blocked with nocodazole. The effects of C3 appeared to be dependent on cell adhesion; less adherent 3T3 fibroblasts exhibited irregular cortical ingression only when cells started to increase attachment during respreading, but managed to complete cytokinesis. Poorly adherent HeLa cells showed neither ectopic cleavage nor completion of cytokinesis. Our results indicate that Rho does not simply activate actin-myosin II interactions during cytokinesis, but regulates the spatial pattern of cortical activities and completion of cytokinesis possibly through modulating the mechanical strength of the cortex.

Show MeSH
Related in: MedlinePlus