The RHO1 gene encodes a homologue of mammalian remains to be clarified whether these target proteins of RhoA small G-protein in the yeast Saccharomyces Rho are involved in the reorganization of actin cytocerevisiae. Rho1p is required for bud formation and is skeleton. Very recently, one of these proteins, Rho kinase, localized at a bud tip or a cytokinesis site. We have recently shown that Bni1p is a potential target of has been shown to inhibit the myosin phosphatase activity Rho1p. Bni1p shares the FH1 and FH2 domains with (Kimura et al., 1996), although its physiological significproteins involved in cytokinesis or establishment of ance remains to be clarified. cell polarity. In S.cerevisiae, there is an open reading Cells of the budding yeast Saccharomyces cerevisiae frame (YIL159W) which encodes another protein grow by budding for cell division, and the actin cytohaving the FH1 and FH2 domains and we have named skeleton plays a pivotal role in the budding process this gene BNR1 (BNI1 Related). Bnr1p interacts with (Drubin, 1991 Johnson and Pringle, 1990). mutant shows a severe temperature-sensitive growth RHO1 is a homologue of the mammalian RhoA gene and phenotype. Cells of the bni1 bnr1 mutant arrested we have shown that the rho1 mutants are deficient in at the restrictive temperature are deficient in bud the budding process (Yamochi et al., 1994). Moreover, emergence, exhibit a random distribution of cortical immunofluorescence microscopic studies indicate that actin patches and often become multinucleate. TheseRho1p is localized at the growth site with cortical actin phenotypes are similar to those of the mutant of patches, including the presumptive budding site, the bud PFY1, which encodes profilin, an actin-binding protein.tip and the cytokinesis site (Yamochi et al., 1994). These Moreover, yeast two-hybrid and biochemical studies results suggest that RHO1 regulates the processes of bud demonstrate that Bni1p and Bnr1p interact directly formation. Concerning the downstream targets of Rho1p, with profilin at the FH1 domains. These results indicate we have shown that one of them is a homologue of that Bni1p and Bnr1p are potential targets of the Rho mammalian protein kinase C, Pkc1p (Nonaka et al., 1995), family members, interact with profilin and regulate which regulates cell wall integrity through the activation the reorganization of actin cytoskeleton.of the MAP kinase cascade (Levin and Errede, 1995). We Keywords: actin cytoskeleton/profilin/Rho have also shown that another target of Rho1p is 1,3-β-glucan synthase (Drgonová et al., 1996;Qadota et al., 1996), which is involved in cell wall synthesis. Very recently, we have identified BNI1 as a third potential target

Both E-cadherin, a cell-cell adhesion molecule, and cMet, the hepatocyte growth factor (HGF)/scatter factor (SF) receptor, were colocalized at cell-cell adhesion sites of MDCK cells. HGF/SF or a phorbol ester, 12-Otetradecanoylphorbol-13-acetate (TPA), induced disruption of cell-cell adhesion, which was accompanied by endocytosis of both E-cadherin and c-Met. Reduction of medium Ca 2+ to a micromolar range showed the same e ects. Re-increase in medium Ca 2+ to a millimolar range formed cell-cell adhesion, which was accompanied by exocytosis of E-cadherin and c-Met, followed by their re-colocalization at the cell-cell adhesion sites. These results suggest that E-cadherin and c-Met are colocalized at cell-cell adhesion sites and undergo co-endo-exocytosis. We have previously shown that TPA does not induce disruption of cell-cell adhesion and subsequent scattering of MDCK cells stably expressing a dominant active mutant of RhoA or Rac1 small G protein or a dominant negative mutant of Rab5 small G protein. In these cell lines, the HGF-or TPA-induced coendocytosis of E-cadherin and c-Met was inhibited, but the coendocytosis of E-cadherin and c-Met in response to reduction of medium Ca 2+ was not a ected. Wortmannin, an inhibitor of phosphoinositide (PI) 3-kinase, inhibited the HGF-induced disruption of cell-cell junction and endocytosis of E-cadherin and c-Met, but not the TPA-induced ones. These results suggest that disruption of cell-cell adhesion is involved in the HGF-or TPAinduced coendocytosis of E-cadherin and c-Met in MDCK cells, and that the Rho and Rab family members indirectly regulate this coendocytosis. In addition, coendocytosis of E-cadherin and c-Met in response to HGF is partly mediated by PI 3-kinase. The cross-talk between cell-cell and cell-matrix adherens junctions is discussed.

Proteins containing the formin homology (FH) domains FH1 and FH2 are involved in cytokinesis or establishment of cell polarity in a variety of organisms. We have shown that the FH proteins Bni1p and Bnr1p are potential targets of the Rho family small GTP-binding proteins and bind to an actin-binding protein, profilin, at their proline-rich FH1 domains to regulate reorganization of the actin cytoskeleton in the yeast Saccharomyces cerevisiae. We found here that a novel Src homology 3 (SH3) domain-containing protein, encoded by YMR032w, interacted with Bnr1p in a GTP-Rho4p-dependent manner through the FH1 domain of Bnr1p and the SH3 domain of Ymr032wp. Ymr032wp weakly bound to Bni1p. Ymr032wp was homologous to cdc15p, which is involved in cytokinesis in Schizosaccharomyces pombe, and we named this gene HOF1 (homolog of cdc 15). Both Bnr1p and Hof1p were localized at the bud neck, and both the bnr1 and hof1 mutations showed synthetic lethal interactions with the bni1 mutation. The hof1 mutant cells showed phenotypes similar to those of the septin mutants, indicating that HOF1 is involved in cytokinesis. These results indicate that Bnr1p directly interacts with Hof1p as well as with profilin to regulate cytoskeletal functions in S. cerevisiae.The Rho family belongs to the small G protein superfamily and regulates various cell functions through reorganization of the actin cytoskeleton (for reviews, see Refs. 1 and 2). Many potential targets of Rho have been identified (for a review, see Ref.3), but it has not yet been thoroughly clarified how Rho regulates reorganization of the actin cytoskeleton through these targets.The actin cytoskeleton plays a pivotal role in the budding processes in the yeast Saccharomyces cerevisiae (for a review, see Ref. 4). This yeast has the Rho family members, including RHO1, RHO2, RHO3, RHO4, and CDC42, which are involved in the budding processes (for reviews, see Refs. 4 and 5). We have isolated BNI1 as a potential target of RHO1, which links RHO1 with the actin cytoskeleton (6). BNI1 has subsequently been shown to be a potential target of CDC42, RHO3, and RHO4 (7). BNR1 is a BNI1-related gene and is a potential target of RHO4 (8). Bni1p and Bnr1p are members of the FH 1 family of proteins, which are defined by the presence of two formin homology domains, the proline-rich FH1 domain and the FH2 domain. The FH proteins play an important role in the actin cytoskeleton-dependent processes, including cytokinesis and establishment of cell polarity (for reviews, see Refs. 9 and 10). We have recently shown that Bni1p interacts with elongation factor 1␣, which binds to and bundles actin filaments (11), and that Spa2p is required for the localization of Bni1p at the bud tip (12). Bni1p and Bnr1p, at their FH1 domains, bind to an actin monomer-binding protein, profilin, which is implicated in actin polymerization (7,8). A proline-rich sequence also interacts with an SH3 domain, which is found in a wide variety of proteins, ranging from cytoskeletal components to signal transducing enzymes (for a ...

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The processes of the formation of dialkylbenzenes from monoalkylbenzene, such as disproportionation and alkylation, are among the most important in the chemical industry. These processes were carried out using solid acid catalysts in earlier times. The activity of these catalysts, such as silica±alumina, was low. Since the late 1960s, ZSM-5 catalysts have been extensively studied because of their much higher selectivity for para-isomers, which are the most valuable compounds for commercial use. However, para-selectivity significantly decreases because of the acid sites on the external surface and the size of the pore openings. Hence, surface modification and pore size control have been proposed in order to enhance the selectivity.[1±9]On the other hand, zeolites have been studied for membrane separation techniques, [10±13] as well as for catalysis, for a long time. In the last decades, combined chemical reactors with zeolite membranes have been of great interest and a lot of applications have been reported.[14±21] Compared with conventional chemical reactors, membrane reactors have great advantages, such as higher selectivity and/or yield of products, simplification of processes, and inhibition of catalyst poisoning. However, the main issue of membrane reactors is the lower permeation rate than the reaction rate. Due to the low permeation flux, a significantly large membrane area and thin zeolite membrane are required. However, the synthesis of large zeolite membranes without any defects, like pinholes or cracks, and a method of controlling the membrane thicknesshave not yet been established. Recently, we have proposed a particle level membrane reactor. A catalyst particle has been coated with a permselective membrane. [22,23] The platinum-loaded TiO 2 particles (particle size = 0.6 mm) coated with a silicalite-1 membrane showed high product selectivity in the hydrogenation of hex-1-ene (1-Hex) and 3,3-dimethylbut-1-ene (3,3-DMB) mixtures. The 1-Hex/3,3-DMB selectivity was 20 because of selective permeation of 1-Hex through the silicalite-1 membrane.[23] Silica± alumina catalyst particles (particle size = 1 mm) were coated with silicalite-1 membranes (silicalite/silica±alumina) and used for the disproportionation of toluene to produce xylene isomers. The silicalite-1 coatings on catalyst particles enhanced the para-selectivity [22] because of selective removal of p-xylene through the silicalite-1 membrane. Toluene conversion, however, significantly decreased from 1.5 to 0.08 % with the coating because the thickness of the silicalite-1 membrane was large (40 lm), which limited the diffusion of the products. In addition, the catalytic activity of silica±alumina was not very high.To solve these problems, in this study we have developed a novel composite catalyst consisting of a zeolite crystal with an inactive thin layer. A silicalite-1 layer was grown on protonexchanged ZSM-5 crystals (silicalite/H-ZSM-5). The conventional zeolite films on porous supports as well as on the particles [22,23] mentioned above have co...

Proteins containing formin homology domains, FH1 and FH2, are involved in cytokinesis or establishment of cell polarity in a variety of organisms. Bni1p and Bnr1p are FH proteins and potential targets of the Rho family small GTP-binding proteins in S. cerevisiae. We have shown that Bnr1p is localized at the bud neck to interact with Hof1p, involved in cytokinesis. We report here that the overexpression of BNR1 causes a cytokinesis de®ciency which is similar to the phenotypes of the septin mutants, including cdc3, cdc10, cdc11, and cdc12. The region required for the septin mutant phenotypes was mapped to Bnr1p (35 ± 500), which coincided with the region required for the bud-neck localization. To further isolate a gene interacting with BNI1 or BNR1, a multicopy suppressor of the bni1 bnr1 mutant was isolated. This gene encoded Smy1p, a kinesin-related protein. Bnr1p, but not Bni1p, directly interacted with the C-terminal region of Smy1p. The Smy1p-interacting region of Bnr1p was mapped to a region containing the FH2 domain. Bnr1p also directly interacted with Bud6p, a novel actin-binding protein. Bnr1p is thus a multifunctional protein which interacts with the septin system, a microtubule-dependent motor protein, and the actin system, to regulate cytoskeletal functions in S. cerevisiae.

The RHO1 gene encodes a homolog of mammalian RhoA small G protein in the yeast Saccharomyces cerevisiae. We have shown that Bni1p is one of the downstream targets of Rho1p and regulates reorganization of the actin cytoskeleton through the interaction with pro®lin, an actin monomer-binding protein. A Bni1p-binding protein was anity puri®ed from the yeast cytosol fraction and was identi®ed to be Tef1p/ Tef2p, translation elongation factor 1a (EF1a). EF1a is an essential component of the protein synthetic machinery and also possesses the actin ®lament (F-actin)-binding and -bundling activities. EF1a bound to the 186 amino acids region of Bni1p, located between the FH1 domain, the proline-rich pro®lin-binding domain, and the FH2 domain, of which function is not known. The binding of Bni1p to EF1a inhibited its F-actin-binding and -bundling activities. The BNI1 gene deleted in the EF1a-binding region did not suppress the bni1 bnr1 mutation in which the actin organization was impaired. These results suggest that the Rho1p ± Bni1p system regulates reorganization of the actin cytoskeleton through the interaction with both EF1a and pro®lin.

To apply motor proteins as natural nanomolecular machines to transporting systems in nanotechnology, complete temporal control over ON/OFF switching of the motility is necessary. We have studied the photoresponsive inhibition properties of azobenzene-tethered peptides for regulation of kinesin-microtubule motility. Although a compound containing a peptide having an amino acid sequence derived from the kinesin's C-terminus (a known inhibitor of kinesin's motor domain) and also featuring a terminal azobenzene unit exhibited an inhibition effect, the phototunability of this behavior upon irradiation with UV or visible light was only moderate. Unexpectedly, newly synthesized peptides featuring the reverse sequence of amino acids of the C-terminus of kinesin exhibited excellent photoresponsive inhibition. In particular, azobenzene-CONH-IPKAIQASHGR completely stopped and started the motility of kinesin-microtubules in its trans- and cis-rich states, respectively, obtained after irradiation with visible and UV light, respectively. A gliding motility system containing this photoresponsive inhibitor allowed in situ control of the motion of microtubules on a kinesin-coated glass substrate. It is expected that the present results on the photoresponsive nanomotor system open up new opportunities to design nanotransportation systems.