Trends in Cell Biology
Volume 15, Issue 12, December 2005, Pages 651-658
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Cytokinesis series
Cytokinesis: welcome to the Rho zone

https://doi.org/10.1016/j.tcb.2005.10.006Get rights and content

Cytokinesis follows nuclear division and generates two distinct daughter cells, each replete with a full complement of the genome and cytoplasmic organelles. Members of the Rho family of GTPases are crucial regulators of this process in a wide variety of species. In many cell types, cytokinesis is mediated by a discretely localized contractile ring that is rich in actin and myosin. In this article (which is part of the Cytokinesis series), we review recent studies in animal cells that have shown that local assembly of the contractile ring is mediated by a discrete pool of GTP-bound, active RhoA. Advances in detecting the active pool of RhoA have allowed insights into the mechanisms and the molecules that promote the accumulation of active RhoA at the correct time and place in the cell.

Introduction

Members of the Rho family of GTPases regulate a wide variety of biological processes. Many, although not all, of these processes involve regulation of the actin cytoskeleton and the contractility of myosin II. Cytokinesis – the generation of two separated daughter cells from a single progenitor – requires the spatial and temporal control of actin and myosin, and various members of the Rho GTPase family control this process in essentially all eukaryotic organisms. This review discusses the current literature that reveals the molecular mechanisms through which Rho GTPases orchestrate cytokinesis. We focus on recent insights into the mechanism of RhoA localization and activation as well as the function of RhoA effector proteins.

Rho family proteins are ∼20-kDa GTPases that simply consist of a characteristic GTPase domain followed by ∼30 amino acids, the very last four of which are subjected to a series of posttranslational modifications resulting in loss of the three terminal residues and prenylation of the C-terminal cysteine residue. This lipid group contributes to interactions with regulatory components and membrane compartments such as the plasma membrane, endosomes and Golgi. The Rho family is distinguished from other classes of small GTPases by a helix that is inserted between the fifth β-strand and the fourth α-helix [1]. Although members of the Rho family are clearly distinct from other members of the Ras-related GTPase superfamily, there is significant divergence within the subfamily (22 members in humans). Many members of the Rho family can be further subdivided on the basis of sequence into three major subsets, known as Rho, Rac and Cdc42 (Figure 1).

Rho family proteins bind to both GTP or GDP with high affinity and have a slow intrinsic GTPase activity (t1/2 ∼5–10 min [2]). The balance between the GTP- and GDP-bound states is regulated by specific guanine nucleotide dissociation inhibitors (GDIs), GDP–GTP exchange factors (GEFs) that promote GTP binding, and GTPase-activating proteins (GAPs) that promote GTP hydrolysis. GEF or GAP domains are commonly found in the eukaryotic genome (102 and 105 proteins in humans, respectively; smart.embl-heidelberg.de), and many proteins that contain GEF or GAP domains contain additional protein- or lipid-interaction domains that might enable them to respond to changes in cell physiology. GEFs and GAPs are not generally selective for a single GTPase in vitro, allowing for immense regulatory complexity. However, in vivo some processes, including cytokinesis, rely on a single GEF, indicating that there is not a lot of functional redundancy among the many proteins containing GEF domains. When Rho proteins bind to GTP, a conformational change occurs that increases their affinity for a specific set of proteins called effectors. Active Rho binding regulates the activity of the effectors and allows Rho proteins to regulate cellular physiology.

Multiple Rho GTPases are implicated in cytokinesis in various eukaryotes (Box 1), but, in animal cells, RhoA is the central player, and its role will be the focus of this review. Cells depleted of RhoA by RNAi, or cells injected with a Clostridial enzyme, C3 transferase, or C3-treated RhoA, show no cortical contractility or furrow formation in most animal cell types studied 3, 4, 5. Vertebrates have two additional proteins highly related to RhoA (RhoB and RhoC). All three proteins are subject to inactivation by the C3 enzyme. Deletion of the gene encoding RhoB (the most divergent of the three) is not lethal, but defects in vascular development occur, although mitotic defects were not reported [6]. Deletion of the gene encoding RhoC reveals that it too is not essential for cell proliferation – rather, RhoC regulates migration of specific cell types, in particular, tumor cells [7]. The possible role of redundancy among RhoA, B and C in development has not yet been addressed. However, the cytokinesis phenotypes induced by depletion of RhoA imply that it is essential for cytokinesis [8].

Section snippets

RhoA effectors

Activated RhoA orchestrates cytokinesis by binding and regulating specific protein effectors that profoundly affect the actomyosin contractile network (Figure 2). The key RhoA effectors involved in cytokinesis are members of the formin family, plus Rho-dependent kinase (ROCK, also known as ROK) and Citron kinase (Citron K). Briefly, formin promotes the polymerization of actin necessary for formation of the contractile ring. ROCK regulates myosin, which is a crucial component of the contractile

RhoA localization

Although there is compelling evidence that active RhoA is required for furrow formation, ingression and stabilization, this requirement does not necessarily imply that RhoA function is spatially restricted. However, this is an appealing possibility, and several independent means have been developed that allow activated RhoA localization during cytokinesis to be assessed 8, 41 (Box 2). These results indicate that, indeed, active RhoA concentrates at the site of furrow formation. Importantly,

Spatiotemporal regulation of RhoA

The ability to detect active RhoA during cytokinesis has allowed a preliminary dissection of the mechanism that generates the zone of active RhoA that drives furrow formation. The RhoGEF ECT2/Pebble is a crucial upstream regulator that is required for cytokinesis in all metazoans analyzed to date (see [46] for review), and, accordingly, it is required for the localization of active RhoA to the equatorial cortex.

The activity of ECT2 appears to be subject to both spatial and temporal regulation

Inactivation of RhoA

Like many other GTPases, RhoA is believed to act as a molecular switch, and the RhoGEF ECT2 switches RhoA on and allows it to engage its effectors and drive furrow formation. At late stages of division, cells become less contractile 55, 56 and the signal from probes that detect active RhoA is attenuated 8, 41, indicating that RhoA is inactivated. What are the mechanisms responsible for switching RhoA off? This question is not yet fully answered. Two mechanisms could cooperate to inactivate

Concluding remarks

There is a plethora of evidence that RhoA and its regulators and effectors play essential roles in cytokinesis in many different types of cells. However, there might be situations in which alternative pathways can substitute for the canonical pathway that is dependent on the actomyosin-based contractile ring. In Dictyostelium, for example, myosin is essential for division in non-adherent cells [64], but cells lacking myosin can divide if they are attached to a substrate [65]. Likewise, yeast

Acknowledgements

We thank of W. Bement (University of Wisconsin, USA) and G. Dassow (University of Washington, USA) for the Xenopus image in Figure 3.

References (83)

  • S. Romero

    Formin is a processive motor that requires profilin to accelerate actin assembly and associated ATP hydrolysis

    Cell

    (2004)
  • T. Otomo

    Structural basis of Rho GTPase-mediated activation of the formin mDia1

    Mol. Cell

    (2005)
  • F. Matsumura

    Regulation of myosin II during cytokinesis in higher eukaryotes

    Trends Cell Biol.

    (2005)
  • M. Amano

    Phosphorylation and activation of myosin by Rho-associated kinase (Rho-kinase)

    J. Biol. Chem.

    (1996)
  • C.G. Winter

    Drosophila Rho-associated kinase (Drok) links Frizzled-mediated planar cell polarity signaling to the actin cytoskeleton

    Cell

    (2001)
  • F. Di Cunto

    Defective neurogenesis in citron kinase knockout mice by altered cytokinesis and massive apoptosis

    Neuron

    (2000)
  • A. Echard

    Terminal cytokinesis events uncovered after an RNAi screen

    Curr. Biol.

    (2004)
  • Y. Nishimura

    Localization of Rho GTPase in sea urchin eggs

    FEBS Lett.

    (1998)
  • S. Yonemura

    Rho localization in cells and tissues

    Exp. Cell Res.

    (2004)
  • W.G. Somers et al.

    A RhoGEF and Rho family GTPase-activating protein complex links the contractile ring to cortical microtubules at the onset of cytokinesis

    Dev. Cell

    (2003)
  • S. Saito

    Deregulation and mislocalization of the cytokinesis regulator ECT2 activate the Rho signaling pathways leading to malignant transformation

    J. Biol. Chem.

    (2004)
  • J.E. Kim

    The tandem BRCT domains of Ect2 are required for both negative and positive regulation of Ect2 in cytokinesis

    J. Biol. Chem.

    (2005)
  • R. Dechant et al.

    Centrosome separation and central spindle assembly act in redundant pathways that regulate microtubule density and trigger cleavage furrow formation

    Dev. Cell

    (2003)
  • J.C. Canman

    The role of pre- and post-anaphase microtubules in the cytokinesis phase of the cell cycle

    Curr. Biol.

    (2000)
  • M. Mishima

    Central spindle assembly and cytokinesis require a kinesin-like protein/RhoGAP complex with microtubule bundling activity

    Dev. Cell

    (2002)
  • A. Toure

    MgcRacGAP, a new human GTPase-activating protein for Rac and Cdc42 similar to Drosophila rotund RacGAP gene product, is expressed in male germ cells

    J. Biol. Chem.

    (1998)
  • Y. Minoshima

    Phosphorylation by Aurora B converts MgcRacGAP to a RhoGAP during cytokinesis

    Dev. Cell

    (2003)
  • H. Yoshizaki

    Cell type-specific regulation of RhoA activity during cytokinesis

    J. Biol. Chem.

    (2004)
  • N. Tolliday

    Rho1 directs formin-mediated actin ring assembly during budding yeast cytokinesis

    Curr. Biol.

    (2002)
  • F. Chen

    Cdc42 is required for PIP(2)-induced actin polymerization and early development but not for cell viability

    Curr. Biol.

    (2000)
  • D.L. Graham

    A method to measure the interaction of Rac/Cdc42 with their binding partners using fluorescence resonance energy transfer between mutants of green fluorescent protein

    Anal. Biochem.

    (2001)
  • R. Dvorsky et al.

    Always look on the bright site of Rho: structural implications for a conserved intermolecular interface

    EMBO Rep.

    (2004)
  • K. Kishi

    Regulation of cytoplasmic division of Xenopus embryo by rho p21 and its inhibitory GDP/GTP exchange protein (rho GDI)

    J. Cell Biol.

    (1993)
  • V. Jantsch-Plunger

    CYK-4: A Rho family GTPase activating protein (GAP) required for central spindle formation and cytokinesis

    J. Cell Biol.

    (2000)
  • A.X. Liu

    RhoB is dispensable for mouse development, but it modifies susceptibility to tumor formation as well as cell adhesion and growth factor signaling in transformed cells

    Mol. Cell. Biol.

    (2001)
  • A. Hakem

    RhoC is dispensable for embryogenesis and tumor initiation but essential for metastasis

    Genes Dev.

    (2005)
  • O. Yuce

    An ECT2–centralspindlin complex regulates the localization and function of RhoA

    J. Cell Biol.

    (2005)
  • M.G. Giansanti

    Cooperative interactions between the central spindle and the contractile ring during Drosophila cytokinesis

    Genes Dev.

    (1998)
  • M. Evangelista

    Formins direct Arp2/3-independent actin filament assembly to polarize cell growth in yeast

    Nat. Cell Biol.

    (2002)
  • D. Pruyne

    Role of formins in actin assembly: nucleation and barbed-end association

    Science

    (2002)
  • I. Sagot

    Yeast formins regulate cell polarity by controlling the assembly of actin cables

    Nat. Cell Biol.

    (2002)
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