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A microtubule-dependent zone of active RhoA during cleavage plane specification.

Bement WM, Benink HA, von Dassow G - J. Cell Biol. (2005)

Bottom Line: Cytokinetic RhoA activity zones are common to four echinoderm species, the vertebrate Xenopus laevis, and the highly asymmetric cytokinesis accompanying meiosis.Microtubules direct the formation and placement of the RhoA activity zone, and the zone is repositioned after physical spindle displacement.We conclude that microtubules specify the cytokinetic apparatus via a dynamic zone of local RhoA activity.

View Article: PubMed Central - PubMed

Affiliation: Center for Cell Dynamics, Friday Harbor Laboratories, University of Washington, Friday Harbor, WA 98250, USA. wmbement@wisc.edu

ABSTRACT
Cytokinesis in animal cells results from the assembly and constriction of a circumferential array of actin filaments and myosin-2. Microtubules of the mitotic apparatus determine the position at which the cytokinetic actomyosin array forms, but the molecular mechanisms by which they do so remain unknown. The small GTPase RhoA has previously been implicated in cytokinesis. Using four-dimensional microscopy and a probe for active RhoA, we show that active RhoA concentrates in a precisely bounded zone before cytokinesis and is independent of actin assembly. Cytokinetic RhoA activity zones are common to four echinoderm species, the vertebrate Xenopus laevis, and the highly asymmetric cytokinesis accompanying meiosis. Microtubules direct the formation and placement of the RhoA activity zone, and the zone is repositioned after physical spindle displacement. We conclude that microtubules specify the cytokinetic apparatus via a dynamic zone of local RhoA activity.

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A zone of RhoA activity in echinoderm and amphibian cytokinesis. (A) Surface view of four-cell purple urchin embryo; projection of 18 1-μm sections. eGFP-rGBD begins to accumulate immediately before furrowing (arrowheads), brightens, and disappears once furrows are complete. (B) Surface view of eight-cell purple urchin embryo; projection of 18 1-μm sections. eGFP-rGBD accumulates before the furrow appears (arrowheads) at the site of the future furrow. (C) Sectional view through eight-cell green urchin embryo; projection of 16 1-μm sections. Before cleavage (00:00), eGFP-rGBD reveals uniform, cortical RhoA activity, which disappears (02:40) before localized activation of RhoA in the equator (arrowhead, 04:40), where the furrow will develop (06:40). (D) Surface view of X. laevis embryo; projection of 12 sections. eGFP-rGBD appears in narrow stripes (arrowheads) that presage furrow formation. See online supplemental material for Videos corresponding to B–D (Videos 1–3, respectively, available at http://www.jcb.org/cgi/content/full/jcb.200501131/DC1). Times are given in minutes:seconds after filming began. Bars, 25 μm.
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fig1: A zone of RhoA activity in echinoderm and amphibian cytokinesis. (A) Surface view of four-cell purple urchin embryo; projection of 18 1-μm sections. eGFP-rGBD begins to accumulate immediately before furrowing (arrowheads), brightens, and disappears once furrows are complete. (B) Surface view of eight-cell purple urchin embryo; projection of 18 1-μm sections. eGFP-rGBD accumulates before the furrow appears (arrowheads) at the site of the future furrow. (C) Sectional view through eight-cell green urchin embryo; projection of 16 1-μm sections. Before cleavage (00:00), eGFP-rGBD reveals uniform, cortical RhoA activity, which disappears (02:40) before localized activation of RhoA in the equator (arrowhead, 04:40), where the furrow will develop (06:40). (D) Surface view of X. laevis embryo; projection of 12 sections. eGFP-rGBD appears in narrow stripes (arrowheads) that presage furrow formation. See online supplemental material for Videos corresponding to B–D (Videos 1–3, respectively, available at http://www.jcb.org/cgi/content/full/jcb.200501131/DC1). Times are given in minutes:seconds after filming began. Bars, 25 μm.

Mentions: Urchin embryos injected with mRNA encoding eGFP-rGBD showed sufficient expression to allow four-dimensional imaging by the two-to-four-cell stage. Four-dimensional image series showed that eGFP-rGBD consistently concentrated at the equator of purple (Fig. 1, A and B, and Video 1) and green (Fig. 1 C and Video 2, available at http://www.jcb.org/cgi/content/full/jcb.200501131/DC1) urchin blastomeres in narrow zones, coinciding precisely with the sites of furrow formation (Fig. 1, A–C). The zones moved inward in concert with ingressing cleavage furrows and disappeared after the completion of cleavage. Before anaphase, RhoA activity was apparent at a lower level throughout the cortex. This ubiquitous activity, which was more prominent in green than in purple urchins (Fig. 1 C), disappeared abruptly at the onset of anaphase before the emergence of the equatorial RhoA zone.


A microtubule-dependent zone of active RhoA during cleavage plane specification.

Bement WM, Benink HA, von Dassow G - J. Cell Biol. (2005)

A zone of RhoA activity in echinoderm and amphibian cytokinesis. (A) Surface view of four-cell purple urchin embryo; projection of 18 1-μm sections. eGFP-rGBD begins to accumulate immediately before furrowing (arrowheads), brightens, and disappears once furrows are complete. (B) Surface view of eight-cell purple urchin embryo; projection of 18 1-μm sections. eGFP-rGBD accumulates before the furrow appears (arrowheads) at the site of the future furrow. (C) Sectional view through eight-cell green urchin embryo; projection of 16 1-μm sections. Before cleavage (00:00), eGFP-rGBD reveals uniform, cortical RhoA activity, which disappears (02:40) before localized activation of RhoA in the equator (arrowhead, 04:40), where the furrow will develop (06:40). (D) Surface view of X. laevis embryo; projection of 12 sections. eGFP-rGBD appears in narrow stripes (arrowheads) that presage furrow formation. See online supplemental material for Videos corresponding to B–D (Videos 1–3, respectively, available at http://www.jcb.org/cgi/content/full/jcb.200501131/DC1). Times are given in minutes:seconds after filming began. Bars, 25 μm.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2171391&req=5

fig1: A zone of RhoA activity in echinoderm and amphibian cytokinesis. (A) Surface view of four-cell purple urchin embryo; projection of 18 1-μm sections. eGFP-rGBD begins to accumulate immediately before furrowing (arrowheads), brightens, and disappears once furrows are complete. (B) Surface view of eight-cell purple urchin embryo; projection of 18 1-μm sections. eGFP-rGBD accumulates before the furrow appears (arrowheads) at the site of the future furrow. (C) Sectional view through eight-cell green urchin embryo; projection of 16 1-μm sections. Before cleavage (00:00), eGFP-rGBD reveals uniform, cortical RhoA activity, which disappears (02:40) before localized activation of RhoA in the equator (arrowhead, 04:40), where the furrow will develop (06:40). (D) Surface view of X. laevis embryo; projection of 12 sections. eGFP-rGBD appears in narrow stripes (arrowheads) that presage furrow formation. See online supplemental material for Videos corresponding to B–D (Videos 1–3, respectively, available at http://www.jcb.org/cgi/content/full/jcb.200501131/DC1). Times are given in minutes:seconds after filming began. Bars, 25 μm.
Mentions: Urchin embryos injected with mRNA encoding eGFP-rGBD showed sufficient expression to allow four-dimensional imaging by the two-to-four-cell stage. Four-dimensional image series showed that eGFP-rGBD consistently concentrated at the equator of purple (Fig. 1, A and B, and Video 1) and green (Fig. 1 C and Video 2, available at http://www.jcb.org/cgi/content/full/jcb.200501131/DC1) urchin blastomeres in narrow zones, coinciding precisely with the sites of furrow formation (Fig. 1, A–C). The zones moved inward in concert with ingressing cleavage furrows and disappeared after the completion of cleavage. Before anaphase, RhoA activity was apparent at a lower level throughout the cortex. This ubiquitous activity, which was more prominent in green than in purple urchins (Fig. 1 C), disappeared abruptly at the onset of anaphase before the emergence of the equatorial RhoA zone.

Bottom Line: Cytokinetic RhoA activity zones are common to four echinoderm species, the vertebrate Xenopus laevis, and the highly asymmetric cytokinesis accompanying meiosis.Microtubules direct the formation and placement of the RhoA activity zone, and the zone is repositioned after physical spindle displacement.We conclude that microtubules specify the cytokinetic apparatus via a dynamic zone of local RhoA activity.

View Article: PubMed Central - PubMed

Affiliation: Center for Cell Dynamics, Friday Harbor Laboratories, University of Washington, Friday Harbor, WA 98250, USA. wmbement@wisc.edu

ABSTRACT
Cytokinesis in animal cells results from the assembly and constriction of a circumferential array of actin filaments and myosin-2. Microtubules of the mitotic apparatus determine the position at which the cytokinetic actomyosin array forms, but the molecular mechanisms by which they do so remain unknown. The small GTPase RhoA has previously been implicated in cytokinesis. Using four-dimensional microscopy and a probe for active RhoA, we show that active RhoA concentrates in a precisely bounded zone before cytokinesis and is independent of actin assembly. Cytokinetic RhoA activity zones are common to four echinoderm species, the vertebrate Xenopus laevis, and the highly asymmetric cytokinesis accompanying meiosis. Microtubules direct the formation and placement of the RhoA activity zone, and the zone is repositioned after physical spindle displacement. We conclude that microtubules specify the cytokinetic apparatus via a dynamic zone of local RhoA activity.

Show MeSH
Related in: MedlinePlus