Kentaro Nakano

University of Tsukuba, Tsukuba, Ibaraki, Japan

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Publications (34)114.51 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Sporulation, gametogenesis in yeast, consists of meiotic nuclear division and spore morphogenesis. In the fission yeast Schizosaccharomyces pombe, four haploid nuclei produced after meiosis II are encapsulated by the forespore membrane (FSM), which is newly synthesized from spindle pole bodies (SPBs) in the cytoplasm of the mother cell as spore precursors. Although the coordination between meiosis and FSM assembly is vital for proper sporulation, the underlying mechanism remains unclear. In the present study, we identified a novel meiosis-specific protein Npg1, which has been implicated in the efficient formation of spores and spore viability. The accumulation and organization of the FSM was compromised in npg1-null cells, leading to the error-prone envelopment of nuclei. Npg1 first emerged as internuclear dots and translocated to the SPBs before the FSM assembled. Genetic analysis revealed that Npg1 worked with the FSM proteins Spo3 and Meu14. These results suggest a possible signaling link from the nucleus to the meiotic SPBs in order to associate the onset of FSM assembly with meiosis II, which ensures the successful partitioning of gametic nuclei.
    Journal of cell science. 08/2014;
  • Masak Takaine, Osamu Numata, Kentaro Nakano
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    ABSTRACT: During cytokinesis in many eukaryotic cells, myosin-II concentrates at the equatorial cortex with actin filaments (F-actin) and is supposed to generate forces to divide the cell into two, which is called the contractile ring (CR) hypothesis. Several lines of evidence indicate that the myosin-II is recruited independently of F-actin and interacts specifically with the equatorial F-actin. Molecular details of these mechanisms are still unknown. We used the fission yeast Schizosaccharomyces pombe to investigate the regulation of myosin-II localization. We demonstrate that the CR myosin-II was composed of F-actin-dependent and -independent fractions by simultaneously observing F-actin and myosin. The F-actin-independent fraction was visualized as cortical dots in the absence of F-actin. IQGAP Rng2, an indispensable element of CR, was implicated in maintenance of the F-actin-independent fraction of myosin-II, whereas anillin Mid1 was required for assembly but not for maintenance of the fraction. In the CR of the rng2 mutant, myosin-II was less concentrated, unstable, and nonhomogeneous, which often resulted in cytokinesis failure. These results suggest that Rng2 tethers myosin-II to the cortex along the CR independently of F-actin to provide a sufficient concentration. The robust localization of myosin-II would ensure successful cytokinesis.
    Genes to Cells 12/2013; · 2.73 Impact Factor
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    ABSTRACT: In eukaryotic cells that multiply by binary fission, the interaction of actin filaments with myosin II in the contractile ring is widely recognized to generate force for membrane ingression into the cleavage furrow; however, the expression of myosin II is restricted in animals, yeast, fungi, and amoeba (collectively, unikonts). No corresponding motor protein capable of forming mini-filaments that could exert sufficient tension to cleave the cell body is found in bikonts, consisting of planta, algae, and most protozoa; however, cells in some bikont lineages multiply by binary fission, as do animal cells. Of these, the ciliate Tetrahymena is known to form an actin ring beneath the division furrow in cytokinesis. Here, we investigated the role of filamentous actin in the cytokinesis of Tetrahymena pyriformis by treating synchronized dividing cells with an actin-inhibiting drug, Latrunculin-A. Video microscopic observation of live cells undergoing cytokinesis was performed, and contrary to expectation, we found that initiation of furrow ingression and its progress are not suppressed under the inhibitory condition of actin polymerization in Tetrahymena cells. We suggest that an actin filament-independent mechanism of binary fission may have been acquired during the evolution in this organism.
    ZOOLOGICAL SCIENCE 12/2013; 30(12):1044-1049. · 1.08 Impact Factor
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    ABSTRACT: ADF/cofilin is a highly conserved actin-modulating protein. Reorganization of the actin cytoskeleton in vivo through severing and depolymerizing F-actin by this protein is essential for various cellular events such as endocytosis, phagocytosis, cytokinesis and cell migration. We show that in the ciliate Tetrahymena thermophila, the ADF/cofilin-homologue, Adf73p, associates with actin on nascent food vacuoles. Overexpression of Adf73p disrupted the proper localization of actin and inhibited the formation of food vacuoles. In vitro, recombinant Adf73p promoted the depolymerization of filaments made of T. thermophila actin, Act1p. Knockout cells lacking ADF73 gene are viable but grow extremely slowly and have a severely decreased rate of food vacuole formation. Knockout cells have abnormal aggregates of actin in the cytoplasm. Surprisingly, unlike in animals and yeasts, in Tetrahymena, ADF/cofilin is not required for cytokinesis. Thus, the Tetrahymena model shows promise for future studies on the role of ADF/cofilin in vivo.
    Eukaryotic Cell 05/2013; · 3.59 Impact Factor
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    ABSTRACT: Mitochondria activation factor (MAF) is a high-molecular-weight polyphenol purified from black tea that activates mitochondrial respiration. It increased the mitochondrial membrane potential and motility of sea urchin sperm, by up to 8%, to the same extent as sperm-activating peptides (SAPs) secreted by the egg. Unlike SAPs, MAF had no effect on sperm swimming behavior, suggesting that the mechanism of sperm activation by MAF is different from that of SAPs.
    Bioscience Biotechnology and Biochemistry 12/2012; · 1.27 Impact Factor
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    ABSTRACT: Proper cell morphogenesis requires the co-ordination of cell polarity, cytoskeletal organization and vesicle trafficking. The Schizosaccharomyces pombe mutant pob1-664 has a curious lemon-like shape, the basis of which is not understood. Here, we found abundant vesicle accumulation in these cells, suggesting that Pob1 plays a role in vesicle trafficking. We identified Rho3 as a multicopy suppressor of this phenotype. Because Rho3 function is related to For3, an actin-polymerizing protein, and Sec8, a component of the exocyst complex, we analyzed their functional relationship with Pob1. Pob1 was essential for the formation of actin cables (by interacting with For3) and for the polarized localization of Sec8. Although neither For3 nor Sec8 is essential for polarized growth, their simultaneous disruption prevented tip growth and yielded a lemon-like cell morphology similar to pob1-664. Thus, Pob1 may ensure cylindrical cell shape of S. pombe by coupling actin-mediated vesicle transport and exocyst-mediated vesicle tethering during secretory vesicle targeting.
    Traffic 03/2011; 12(6):726-39. · 4.65 Impact Factor
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    ABSTRACT: Myosins are eukaryotic actin-dependent molecular motors that play important roles in many cellular events. The function of each myosin is determined by a variety of functional domains in its tail region. In some major model organisms, the functions and properties of myosins have been investigated based on their amino acid sequences. However, in protists, myosins have been little studied beyond the level of genome sequences. We therefore investigated the mRNA expression levels and amino acid sequences of 13 myosin genes in the ciliate Tetrahymena thermophila. This study is an overview of myosins in T. thermophila, which has no typical myosins, such as class I, II, or V myosins. We showed that all 13 myosins were expressed in vegetative cells. Furthermore, these myosins could be divided into 3 subclasses based on four functional domains in their tail regions. Subclass 1 comprised of 8 myosins has both MyTH4 and FERM domains, and has a potential to function in vesicle transport or anchoring between membrane and actin filaments. Subclass 2 comprised of 4 myosins has RCC1 (regulator of chromosome condensation 1) domains, which are found only in some protists, and may have unconventional features. Subclass 3 is comprised of one myosin, which has a long coiled-coil domain like class II myosin. In addition, phylogenetic analysis on the basis of motor domains showed that T. thermophila myosins are separated into two clusters: one consists of subclasses 1 and 2, and the other consists of subclass 3.
    Gene 02/2011; 480(1-2):10-20. · 2.20 Impact Factor
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    ABSTRACT: Cytokinesis is the final stage of the cell cycle, and ensures completion of both genome segregation and organelle distribution to the daughter cells. Cytokinesis requires the cell to solve a spatial problem (to divide in the correct place, orthogonally to the plane of chromosome segregation) and a temporal problem (to coordinate cytokinesis with mitosis). Defects in the spatiotemporal control of cytokinesis may cause cell death, or increase the risk of tumor formation [Fujiwara et al., 2005 (Fujiwara T, Bandi M, Nitta M, Ivanova EV, Bronson RT, Pellman D. 2005. Cytokinesis failure generating tetraploids promotes tumorigenesis in p53-null cells. Nature 437:1043–1047); reviewed by Ganem et al., 2007 (Ganem NJ, Storchova Z, Pellman D. 2007. Tetraploidy, aneuploidy and cancer. Curr Opin Genet Dev 17:157–162.)]. Asymmetric cytokinesis, which permits the generation of two daughter cells that differ in their shape, size and properties, is important both during development, and for cellular homeostasis in multicellular organisms [reviewed by Li, 2007 (Li R. 2007. Cytokinesis in development and disease: variations on a common theme. Cell Mol Life Sci 64:3044–3058)]. The principal focus of this review will be the mechanisms of cytokinesis in the mitotic cycle of the yeast Schizosaccharomyces pombe. This simple model has contributed significantly to our understanding of how the cell cycle is regulated, and serves as an excellent model for studying aspects of cytokinesis. Here we will discuss the state of our knowledge of how the contractile ring is assembled and disassembled, how it contracts, and what we know of the regulatory mechanisms that control these events and assure their coordination with chromosome segregation.
    Cytoskeleton 02/2011; 68(2):69-88. · 2.87 Impact Factor
  • Yasuharu Kushida, Kentaro Nakano, Osamu Numata
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    ABSTRACT: To reveal the molecular systems involved in the division of a cell and its contents during cell proliferation is one of the major subjects in cell biology. Although cytoskeletal organization during mitosis has been well studied, consensus on the molecular basis of amitosis has not been achieved. Here we adapted an immunofluorescence method and investigated the cellular localization of γ-tubulin and microtubules (MTs) in dividing Tetrahymena. Although the macronucleus (Mac) lacks a bipolar spindle, γ-tubulin and MTs are specifically detected in the dividing Mac and show a marked change in the pattern of localization. First, γ-tubulin and MTs appear in whole Mac, then, γ-tubulin gathers at the center of the Mac where the aster-like structure of MTs forms. On Mac expansion, MTs associated with numerous dots of γ-tubulin are reorganized into longitudinally arranged bundles, suggesting that the mutual sliding of each filament and polymerization of MTs may induce Mac expansion. Moreover, normal Mac expansion and equal segregation of the Mac are severely disturbed when γ-tubulin is shut off. We propose that γ-tubulin-mediated MT assembly is required in Mac amitosis of Tetrahymena.
    Cytoskeleton 02/2011; 68(2):89-96. · 2.87 Impact Factor
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    ABSTRACT: Actin-depolymerizing factor (ADF)/cofilin is widely expressed in eukaryotes and plays a central role in reorganizing the actin cytoskeleton by disassembling actin filaments. The ADF-homologous domain (ADF-H) is conserved in several other actin-modulating proteins such as twinfilin, Abp1/drebrin, and coactosin. Although these proteins interact with actin via ADF-H, their effects on actin are not identical to each other. Here, we report a novel ADF/cofilin-super family protein, Gmf1 (Glia maturation factor-like protein 1), from the fission yeast Schizosaccharomyces pombe. Gmf1 is a component of actin patches, which are located on the cell cortex and required for endocytosis, and may be involved in the control of the disassembly of actin patches since its overexpression diminishes them. We provide evidence that Gmf1 binds weakly if at all to actin, but it associates with actin-related protein (Arp) 2/3 complex and suppresses its functions such as the promotion of actin polymerization and branching filaments. Importantly, Arp2/3 complex-suppressing activity is conserved among GMF-family proteins from other organisms. Given the functional plasticity of ADF-H, GMF-family proteins possibly have changed their target from conventional actin to Arps through molecular evolution.
    Cytoskeleton 06/2010; 67(6):373-82. · 2.87 Impact Factor
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    ABSTRACT: Small GTPase Rab (products of ras genes from rat brain) is a widely conserved molecular switch among eukaryotes and regulates membrane trafficking pathways. It is generally considered that the number of Rab encoded in the genome correlates with multicellularity; however, we found that unicellular ciliates Tetrahymena thermophila (Tt) and Paramecium tetraurelia (Pt) possess many more Rab genes in their genome than the 64 HsRab genes in the human genome. We succeeded in isolating 86 cDNA clones of 88 TtRab genes in the Tetrahymena genome. By comparing the amino acid sequence of Rab in humans and the budding yeast Saccharomyces cerevisiae, 42 TtRab belonged to subfamilies functionally characterized and designated as conventional Rab, while the remaining 44 TtRab were considered to be species-specific. To examine the diversity of Rab in ciliates, we searched for Rab genes in the genome database of P. tetraurelia. Overall, 229 PtRab genes were found and categorized as 157 conventional and 72 species-specific PtRab, respectively. Among them, nine PtRab genes showed high homology to seven TtRab, suggesting the conservation of ciliate-specific Rab. These data suggested that the range of Rab is markedly amplified and diversified in ciliates, which may support the elaborate cellular structures and vigorous phagocytosis of those organisms.
    Journal of Eukaryotic Microbiology 01/2010; 57(5):389-99. · 2.16 Impact Factor
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    Masak Takaine, Osamu Numata, Kentaro Nakano
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    ABSTRACT: The contractile ring (CR) consists of bundled actin filaments and myosin II; however, the actin-bundling factor remains elusive. We show that the fission yeast Schizosaccharomyces pombe IQGAP Rng2 is involved in the generation of CR F-actin and required for its arrangement into a ring. An N-terminal fragment of Rng2 is necessary for the function of Rng2 and is localized to CR F-actin. In vitro the fragment promotes actin polymerization and forms linear arrays of F-actin, which are resistant to the depolymerization induced by the actin-depolymerizing factor Adf1. Our findings indicate that Rng2 is involved in the generation of CR F-actin and simultaneously bundles the filaments and regulates its dynamics by counteracting the effects of Adf1, thus enabling the reconstruction of CR F-actin bundles, which provides an insight into the physical properties of the building blocks that comprise the CR.
    The EMBO Journal 09/2009; 28(20):3117-31. · 9.82 Impact Factor
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    ABSTRACT: Actin-depolymerizing factor (ADF)/cofilin is a well-conserved actin-modulating protein, which induces reorganization of the actin cytoskeleton by severing and depolymerizing F-actin. ADF/cofilin also binds to G-actin and inhibits nucleotide exchange, and hence, is supposed to regulate the nucleotide-bound state of the cellular G-actin pool cooperating with profilin, another well-conserved G-actin-binding protein that promotes nucleotide exchange. In this report, we investigated the biochemical properties of the ADF/cofilin-like protein Adf73p from ciliate Tetrahymena thermophila. Adf73p also binds to both G- and F-actin and severs and depolymerizes F-actin. Unlike canonical ADF/cofilin, however, Adf73p accelerates nucleotide exchange on actin and allows repolymerization of disassembled actin. These results suggest that the actin cytoskeleton of T. thermophila is regulated by Adf73p in a different way from those of mammals, plants, and yeasts.
    Biochemical and Biophysical Research Communications 09/2009; 390(1):54-9. · 2.28 Impact Factor
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    ABSTRACT: Phagocytosis is a fundamental cellular event for the uptake of nutrients from the environment in several kinds of eukaryote. Most ciliates egest waste and undigested materials in food vacuoles (FVs) through a cytoproct, which is a specific organelle for defecation. It is considered that FV egestion is initiated by fusion between the FV membrane and plasma membrane in a cytoproct and completed with retrieval of the membrane into a cytoplasmic space. In addition, electron microscopy indicated that microfilaments might be involved in the recycling process of the FV membrane in ciliates over 30 years ago; however, there is no conclusive evidence. Here we demonstrated actin organization on FV near a cytoproct in Tetrahymena thermophila by using a marker for a cytoproct. Moreover, it was revealed that cells treated with actin cytoskeletal inhibitor, Latrunculin B, might be suppressed for membrane retrieval in a cytoproct following FV egestion. On the other hand, the actin structures, likely to be the site of membrane retrieval, were frequently observed in the cells treated with cytoplasmic microtubules inhibitor, Nocodazole. We concluded that actin filaments were probably required for recycling of the FV membrane in a cytoproct although the role was not essential for FV egestion. In addition, it was possible that microtubules might be involved in transportation of recycling vesicles of FV coated with F-actin.
    Cell Motility and the Cytoskeleton 06/2009; 66(7):371-7. · 4.19 Impact Factor
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    Kentaro Nakano, Issei Mabuchi
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    ABSTRACT: Actin-capping protein (CP) is a heterodimeric protein which is expressed in various eukaryotic cells. CP binds to the barbed end of the actin filaments in vitro and inhibits both the association and dissociation of actin monomers at this end. However, the cellular role of CP has not been uncovered. Here we investigated the function of CP in fission yeast cells. The fission yeast CP is composed of Acp1 and Acp2. It was found that Acp2 accumulated as cortical dots at the cell ends during interphase and the mid-region of mitotic cells, which disappeared in the absence of Acp1 or F-actin. Acp1 and Acp2, when co-over-expressed, decreased F-actin structures in cells, and cytokinesis was often interrupted in these cells. On the other hand, disruption of one of the CP genes affected the distribution of F-actin patches at cell ends and decreased the rate of actin depolymerization in vivo. Moreover, genetic analysis showed that CP controls actin dynamics together with ADF/cofilin and profilin. In addition, CP is likely involved in assembling the F-actin contractile ring and F-actin patch with F-actin-crosslinking proteins.
    Genes to Cells 09/2006; 11(8):893-905. · 2.73 Impact Factor
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    Kentaro Nakano, Issei Mabuchi
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    ABSTRACT: The role of the actin-depolymerizing factor (ADF)/cofilin-family protein Adf1 in cytokinesis of fission yeast cells was studied. Adf1 was required for accumulation of actin at the division site by depolymerizing actin at the cell ends, assembly of the contractile ring through severing actin filaments, and maintenance of the contractile ring once formed. Genetic and cytological analyses suggested that it collaborates with profilin and capping protein in the mitotic reorganization of the actin cytoskeleton. Furthermore, it was unexpectedly found that Adf1 and myosin-II also collaborate in assembling the contractile ring. Tropomyosin was shown to antagonize the function of Adf1 in the contractile ring. We propose that formation and maintenance of the contractile ring are achieved by a balanced collaboration of these proteins.
    Molecular Biology of the Cell 05/2006; 17(4):1933-45. · 4.60 Impact Factor
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    Tadashi Mutoh, Kentaro Nakano, Issei Mabuchi
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    ABSTRACT: The Rho GTPase acts as a binary molecular switch by converting between a GDP-bound inactive and a GTP-bound active conformational state. The guanine nucleotide exchange factors (GEFs) are critical activators of Rho. Rho1 has been shown to regulate actin cytoskeleton and cell wall synthesis in the fission yeast Schizosaccharomyces pombe. Here we studied function of fission yeast RhoGEFs, Rgf1, Rgf2, and Rgf3. It was shown that these proteins have similar molecular structures, and function as GEFs for Rho1. Disruption of either rgf1 or rgf2 did not show a serious effect on the cell. On the other hand, disruption of rgf3 caused severe defects in contractile ring formation, F-actin patch localization, and septation during cytokinesis. Rgf1 and Rgf2 were localized to the cell ends during interphase and the septum. Rgf3 formed a ring at the division site, which was located outside the contractile ring and inside the septum where Rho1 was accumulated. In summary, Rgf1 and Rgf2 show functional redundancy, and roles of these RhoGEFs are likely to be different from that of Rgf3. Rho1 is likely to be activated by Rgf3 at the division site, and involved in contractile ring formation and/or maintenance and septation.
    Genes to Cells 01/2006; 10(12):1189-202. · 2.73 Impact Factor
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    Kentaro Nakano, Fumihide Bunai, Osamu Numata
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    ABSTRACT: We identified a novel actin-modulating protein Stg 1 in the fission yeast Schizosaccharomyces pombe. Stg 1 is similar to mammalian SM22/transgelin, and biochemical experiments showed that Stg 1 crosslinked F-actin. Microscopic observation suggested that Stg 1 was a component of actin patch. Overexpression of Stg 1 caused a defect in cytokinesis by suppressing the formation of a contractile ring and formation of abnormal aggregates of F-actin in the ends and mid-region of cells. Although distribution of the actin cytoskeleton was not affected by disrupting Stg 1(+), genetic interaction suggested that Stg 1 was likely involved in controlling the organization of the actin cytoskeleton in cell morphogenesis and cytokinesis in fission yeast.
    FEBS Letters 12/2005; 579(28):6311-6. · 3.58 Impact Factor
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    Kentaro Nakano, Ritsuko Arai, Issei Mabuchi
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    ABSTRACT: The small GTPase Rho1 plays an essential role in controlling the organization of the actin cytoskeleton and synthesis of the cell wall in the fission yeast Schizosaccharomyces pombe. Here we studied the role of Rho5 whose primary structure is very similar to that of Rho1. It was found that elevated expression of Rho5 was able to compensate for the lethality of cells lacking Rho1. Rho5 was localized to the ends of interphase cells and the mid-region of mitotic cells. Overexpression of Rho5 caused depolarization of F-actin patches and abnormal formation of the cell wall, as did Rho1. Although rho5(+) was not essential for maintaining the cell shape, rho1 rho5-double null cells showed more severe defects in cell viability than rho1-null cells. Thus, it is likely that Rho5 has an overlapping function with Rho1 in controlling cell growth and division in S. pombe.
    FEBS Letters 10/2005; 579(23):5181-6. · 3.58 Impact Factor
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    ABSTRACT: Rho family small GTPases have been shown to be involved in various cellular activities, including the organization of actin cytoskeleton in eukaryotic cells. There are six rho genes in the fission yeast Schizosaccharomyces pombe. Cdc42 is known to control the polarity of the cell. Rho1, Rho2 and Rho3 play important roles in controlling cell shape and septation. On the other hand, Rho4 and Rho5 have not yet been characterized. Here we report the function of rho4+ in fission yeast. Gene disruption revealed that rho4+ is not essential for cell growth. However, rho4-null cells were abnormally elongated and had multiple septa of irregular shape at 37 degrees C. In these cells, F-actin patches were randomly localized all over the cell periphery, and cytoplasmic microtubules (MTs) were misoriented. On the other hand, the exogenous expression of a constitutively active Rho4-G23V or Rho4-Q74L in wild-type cells induced depolarization of F-actin patches and cytoplasmic MTs. Rho4 was localized to the cell periphery during interphase and septum during mitosis. Both the binding of GTP and isoprenylation of its C-terminus were necessary for the localization. Furthermore, the localization of Rho4 was likely to be controlled by Rho GAP and Rho GDI. Rho4 may control cell morphogenesis and septation by regulating both the actin cytoskeleton and cytoplasmic MTs.
    Genes to Cells 05/2003; 8(4):357-70. · 2.73 Impact Factor

Publication Stats

783 Citations
114.51 Total Impact Points

Institutions

  • 2005–2013
    • University of Tsukuba
      Tsukuba, Ibaraki, Japan
  • 2010
    • National Institute of Infectious Diseases, Tokyo
      Edo, Tōkyō, Japan
  • 1995–2006
    • The University of Tokyo
      • College of Art and Science & Graduate School of Arts and Sciences
      Tokyo, Tokyo-to, Japan
  • 1998
    • Hiroshima University
      • Graduate School of Engineering
      Hiroshima-shi, Hiroshima-ken, Japan