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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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Effects of microtubule depolymerization on C3-induced cleavage. NRK cells were microinjected with C3 during metaphase  followed by treatment with 2.5 μM nocodazole. Time in minutes after the addition (t = 0) of nocodazole is located in the upper right corner. When cells were treated with nocodazole after anaphase onset or at the initiation of cytokinesis (a–e) the furrow continued to ingress. Those locked in metaphase with nocodazole treatment (f–j) underwent cortical ingression without entering anaphase (g and h). This  cell eventually divided into a daughter cell containing all of the chromosomes and an anuclear fragment (i and j, arrows). Bar, 20 μm.
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Figure 7: Effects of microtubule depolymerization on C3-induced cleavage. NRK cells were microinjected with C3 during metaphase followed by treatment with 2.5 μM nocodazole. Time in minutes after the addition (t = 0) of nocodazole is located in the upper right corner. When cells were treated with nocodazole after anaphase onset or at the initiation of cytokinesis (a–e) the furrow continued to ingress. Those locked in metaphase with nocodazole treatment (f–j) underwent cortical ingression without entering anaphase (g and h). This cell eventually divided into a daughter cell containing all of the chromosomes and an anuclear fragment (i and j, arrows). Bar, 20 μm.

Mentions: The above observations suggest that C3 may disrupt the coordination between mitosis and cytokinesis. In control NRK cells, depolymerization of microtubules before anaphase onset causes arrest of mitosis and inhibition of cytokinesis. Depolymerization after the initiation of the cleavage furrow causes it to regress or to follow an irregular path (Wheatley and Wang, 1996; Wheatley et al., 1998). However, all C3-injected NRK cells proceeded with cleavage when treated with the microtubule depolymerizing drug nocodazole after anaphase onset (Fig. 7, c–e). Even when nocodazole was added at metaphase, broad ingressions developed despite the inhibition of mitosis (Fig. 7, f–j). Unlike NRK cells, 3T3 cells showed no C3-induced cleavage activity in the presence of nocodazole.


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)

Effects of microtubule depolymerization on C3-induced cleavage. NRK cells were microinjected with C3 during metaphase  followed by treatment with 2.5 μM nocodazole. Time in minutes after the addition (t = 0) of nocodazole is located in the upper right corner. When cells were treated with nocodazole after anaphase onset or at the initiation of cytokinesis (a–e) the furrow continued to ingress. Those locked in metaphase with nocodazole treatment (f–j) underwent cortical ingression without entering anaphase (g and h). This  cell eventually divided into a daughter cell containing all of the chromosomes and an anuclear fragment (i and j, arrows). Bar, 20 μm.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 7: Effects of microtubule depolymerization on C3-induced cleavage. NRK cells were microinjected with C3 during metaphase followed by treatment with 2.5 μM nocodazole. Time in minutes after the addition (t = 0) of nocodazole is located in the upper right corner. When cells were treated with nocodazole after anaphase onset or at the initiation of cytokinesis (a–e) the furrow continued to ingress. Those locked in metaphase with nocodazole treatment (f–j) underwent cortical ingression without entering anaphase (g and h). This cell eventually divided into a daughter cell containing all of the chromosomes and an anuclear fragment (i and j, arrows). Bar, 20 μm.
Mentions: The above observations suggest that C3 may disrupt the coordination between mitosis and cytokinesis. In control NRK cells, depolymerization of microtubules before anaphase onset causes arrest of mitosis and inhibition of cytokinesis. Depolymerization after the initiation of the cleavage furrow causes it to regress or to follow an irregular path (Wheatley and Wang, 1996; Wheatley et al., 1998). However, all C3-injected NRK cells proceeded with cleavage when treated with the microtubule depolymerizing drug nocodazole after anaphase onset (Fig. 7, c–e). Even when nocodazole was added at metaphase, broad ingressions developed despite the inhibition of mitosis (Fig. 7, f–j). Unlike NRK cells, 3T3 cells showed no C3-induced cleavage activity in the presence of nocodazole.

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