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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 3T3 fibroblasts and cells microinjected with C3 at metaphase. Approximate time in minutes relative to anaphase onset is located in the upper right corner. Control cells (a–e) maintained a very short cleavage furrow. C3-injected cells  (f–j) developed abnormalities very similar to those in NRK cells when they started to respread (compare h–j with Fig. 1, h–j), forming  both elongated furrows and ectopic ingressions. The latter resulted in the formation of anuclear fragments (i and j, arrows). Bar, 20 μm.
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Figure 3: Time lapse images of control 3T3 fibroblasts and cells microinjected with C3 at metaphase. Approximate time in minutes relative to anaphase onset is located in the upper right corner. Control cells (a–e) maintained a very short cleavage furrow. C3-injected cells (f–j) developed abnormalities very similar to those in NRK cells when they started to respread (compare h–j with Fig. 1, h–j), forming both elongated furrows and ectopic ingressions. The latter resulted in the formation of anuclear fragments (i and j, arrows). Bar, 20 μm.

Mentions: To further investigate the role of cell adhesion, C3 was injected into 3T3 fibroblasts that round up during mitosis (Fig. 3). Unlike NRK cells, no spreading was observed immediately after the microinjection of C3. The initiation of cytokinesis in C3-injected cells appeared indistinguishable from that of control 3T3 cells (Fig. 3 g). However, the effects of C3 became clear as the cell started to spread out. The furrow elongated as in NRK cells, while other regions of the cell developed additional sites of ingression or ectopic furrows (Fig. 3, i and j). Control cells never developed such broad or ectopic furrows. All C3-injected 3T3 cells completed cytokinesis (n = 5; Fig. 3 j).


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 3T3 fibroblasts and cells microinjected with C3 at metaphase. Approximate time in minutes relative to anaphase onset is located in the upper right corner. Control cells (a–e) maintained a very short cleavage furrow. C3-injected cells  (f–j) developed abnormalities very similar to those in NRK cells when they started to respread (compare h–j with Fig. 1, h–j), forming  both elongated furrows and ectopic ingressions. The latter resulted in the formation of anuclear fragments (i and j, arrows). Bar, 20 μm.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 3: Time lapse images of control 3T3 fibroblasts and cells microinjected with C3 at metaphase. Approximate time in minutes relative to anaphase onset is located in the upper right corner. Control cells (a–e) maintained a very short cleavage furrow. C3-injected cells (f–j) developed abnormalities very similar to those in NRK cells when they started to respread (compare h–j with Fig. 1, h–j), forming both elongated furrows and ectopic ingressions. The latter resulted in the formation of anuclear fragments (i and j, arrows). Bar, 20 μm.
Mentions: To further investigate the role of cell adhesion, C3 was injected into 3T3 fibroblasts that round up during mitosis (Fig. 3). Unlike NRK cells, no spreading was observed immediately after the microinjection of C3. The initiation of cytokinesis in C3-injected cells appeared indistinguishable from that of control 3T3 cells (Fig. 3 g). However, the effects of C3 became clear as the cell started to spread out. The furrow elongated as in NRK cells, while other regions of the cell developed additional sites of ingression or ectopic furrows (Fig. 3, i and j). Control cells never developed such broad or ectopic furrows. All C3-injected 3T3 cells completed cytokinesis (n = 5; Fig. 3 j).

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