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Sample records for 1-2man rho alpha

  1. Constraints on the CKM Angle alpha in the B to rho rho Decays

    SciTech Connect

    Li, H.

    2004-11-03

    Using a data sample of 122 million {Upsilon}(4S) {yields} B{bar B} decays collected with BABAR detector at the PEP-II asymmetric B factory at SLAC, we measure the time-dependent-asymmetry parameters of the longitudinally polarized component in the B{sup 0} {yields} {rho}{sup +}{rho}{sup -} decay as C{sub L} = -0.23 {+-} 0.24(stat) {+-} 0.14(syst) and S{sub L} = -0.19 {+-} 0.33(stat) {+-} 0.11(syst). The B{sup 0} {yields} {rho}{sup 0}{rho}{sup 0} decay mode is also searched for in a data sample of about 227 million B{bar B} pairs. No significant signal is observed, and an upper limit of 1.1 x 10{sup -6} (90% C.L.) on the branching fraction is set. The penguin contribution to the CKM angle {alpha} uncertainty is measured to be 11{sup o}. All results are preliminary.

  2. Limit on the B0 to rho0rho0 Branching Fraction and Implications for the CKM Angle alpha

    SciTech Connect

    Aubert, B.

    2005-01-03

    The authors search for the decay B{sup 0} {yields} {rho}{sup 0}{rho}{sup 0} in a data sample of about 227 million {Upsilon}(4S) {yields} B{bar B} decays collected with the BABAR detector at the PEP-II asymmetric-energy e{sup +}e{sup -} collider at SLAC. They find no significant signal and set an upper limit of 1.1 x 10{sup -6} at 90% CL on the branching fraction. As a result, the uncertainty due to penguin contributions on the CKM unitarity angle {alpha} measured in B {yields} {rho}{rho} decays is 11{sup o} at 68% CL.

  3. Improved Measurement of B^ \\to\\rho^ \\rho^0 and Determination of the Quark-Mixing Phase Angle~\\alpha

    SciTech Connect

    Aubert, B.; Karyotakis, Y.; Lees, J.P.; Poireau, V.; Prencipe, E.; Prudent, X.; Tisserand, V.; Tico, J.Garra; Grauges, E.; Lopez, L.; Palano, A.; Pappagallo, M.; Eigen, G.; Stugu, B.; Sun, L.; Battaglia, M.; Brown, D.N.; Kerth, L.T.; Kolomensky, Yu.G.; Lynch, G.; Osipenkov, I.L.; /Annecy, LAPP /Barcelona U., ECM /INFN, Bari /Bari U. /Bergen U. /LBL, Berkeley /Birmingham U. /Ruhr U., Bochum /British Columbia U. /Brunel U. /Novosibirsk, IYF /UC, Irvine /UCLA /UC, Riverside /UC, San Diego /UC, Santa Barbara /UC, Santa Cruz /Caltech /Cincinnati U. /Colorado U. /Colorado State U.

    2009-07-14

    The authors present improved measurements of the branching fraction {Beta}, the longitudinal polarization fraction f{sub L}, and the direct CP asymmetry A{sub CP} in the B meson decay channel B{sup +} {yields} {rho}{sup +}{rho}{sup 0}. The data sample was collected with the BABAR detector at SLAC. The results are {Beta}(B{sup +} {yields} {rho}{sup +}{rho}{sup 0}) = (23.7 {+-} 1.4 {+-} 1.4) x 10{sup -6}, f{sub L} = 0.950 {+-} 0.015 {+-} 0.006, and A{sub CP} = -0.054 {+-} 0.055 {+-} 0.010, where the uncertainties are statistical and systematic, respectively. Based on these results, they perform an isospin analysis and determine the CKM weak phase angle {alpha} to be (92.4{sub -6.5}{sup +6.0}){sup 0}.

  4. Improved Measurement of the Cabibbo-Kobayashi-Maskawa angle alpha using B0(B) --> rho+rho- decays.

    PubMed

    Aubert, B; Barate, R; Boutigny, D; Couderc, F; Karyotakis, Y; Lees, J P; Poireau, V; Tisserand, V; Zghiche, A; Grauges, E; Palano, A; Pappagallo, M; Pompili, A; Chen, J C; Qi, N D; Rong, G; Wang, P; Zhu, Y S; Eigen, G; Ofte, I; Stugu, B; Abrams, G S; Borgland, A W; Breon, A B; Brown, D N; Button-Shafer, J; Cahn, R N; Charles, E; Day, C T; Gill, M S; Gritsan, A V; Groysman, Y; Jacobsen, R G; Kadel, R W; Kadyk, J; Kerth, L T; Kolomensky, Yu G; Kukartsev, G; Lynch, G; Mir, L M; Oddone, P J; Orimoto, T J; Pripstein, M; Roe, N A; Ronan, M T; Wenzel, W A; Barrett, M; Ford, K E; Harrison, T J; Hart, A J; Hawkes, C M; Morgan, S E; Watson, A T; Fritsch, M; Goetzen, K; Held, T; Koch, H; Lewandowski, B; Pelizaeus, M; Peters, K; Schroeder, T; Steinke, M; Boyd, J T; Burke, J P; Chevalier, N; Cottingham, W N; Kelly, M P; Cuhadar-Donszelmann, T; Hearty, C; Knecht, N S; Mattison, T S; McKenna, J A; Thiessen, D; Khan, A; Kyberd, P; Teodorescu, L; Blinov, A E; Blinov, V E; Bukin, A D; Druzhinin, V P; Golubev, V B; Ivanchenko, V N; Kravchenko, E A; Onuchin, A P; Serednyakov, S I; Skovpen, Yu I; Solodov, E P; Yushkov, A N; Best, D; Bondioli, M; Bruinsma, M; Chao, M; Eschrich, I; Kirkby, D; Lankford, A J; Mandelkern, M; Mommsen, R K; Roethel, W; Stoker, D P; Buchanan, C; Hartfiel, B L; Weinstein, A J R; Foulkes, S D; Gary, J W; Long, O; Shen, B C; Wang, K; Zhang, L; Del Re, D; Hadavand, H K; Hill, E J; Macfarlane, D B; Paar, H P; Rahatlou, S; Sharma, V; Berryhill, J W; Campagnari, C; Cunha, A; Dahmes, B; Hong, T M; Lu, A; Mazur, M A; Richman, J D; Verkerke, W; Beck, T W; Eisner, A M; Flacco, C J; Heusch, C A; Kroseberg, J; Lockman, W S; Nesom, G; Schalk, T; Schumm, B A; Seiden, A; Spradlin, P; Williams, D C; Wilson, M G; Albert, J; Chen, E; Dubois-Felsmann, G P; Dvoretskii, A; Hitlin, D G; Narsky, I; Piatenko, T; Porter, F C; Ryd, A; Samuel, A; Yang, S; Andreassen, R; Jayatilleke, S; Mancinelli, G; Meadows, B T; Sokoloff, M D; Blanc, F; Bloom, P; Chen, S; Ford, W T; Nauenberg, U; Olivas, A; Rankin, P; Ruddick, W O; Smith, J G; Ulmer, K A; Zhang, J; Chen, A; Eckhart, E A; Harton, J L; Soffer, A; Toki, W H; Wilson, R J; Zeng, Q; Spaan, B; Altenburg, D; Brandt, T; Brose, J; Dickopp, M; Feltresi, E; Hauke, A; Klose, V; Lacker, H M; Maly, E; Nogowski, R; Otto, S; Petzold, A; Schott, G; Schubert, J; Schubert, K R; Schwierz, R; Sundermann, J E; Bernard, D; Bonneaud, G R; Grenier, P; Schrenk, S; Thiebaux, Ch; Vasileiadis, G; Verderi, M; Bard, D J; Clark, P J; Gradl, W; Muheim, F; Playfer, S; Xie, Y; Andreotti, M; Azzolini, V; Bettoni, D; Bozzi, C; Calabrese, R; Cibinetto, G; Luppi, E; Negrini, M; Piemontese, L; Sarti, A; Anulli, F; Baldini-Ferroli, R; Calcaterra, A; de Sangro, R; Finocchiaro, G; Patteri, P; Peruzzi, I M; Piccolo, M; Zallo, A; Buzzo, A; Capra, R; Contri, R; Lo Vetere, M; Macri, M; Monge, M R; Passaggio, S; Patrignani, C; Robutti, E; Santroni, A; Tosi, S; Bailey, S; Brandenburg, G; Chaisanguanthum, K S; Morii, M; Won, E; Dubitzky, R S; Langenegger, U; Marks, J; Schenk, S; Uwer, U; Bhimji, W; Bowerman, D A; Dauncey, P D; Egede, U; Gaillard, J R; Morton, G W; Nash, J A; Nikolich, M B; Taylor, G P; Charles, M J; Grenier, G J; Mallik, U; Mohapatra, A K; Cochran, J; Crawley, H B; Eyges, V; Meyer, W T; Prell, S; Rosenberg, E I; Rubin, A E; Yi, J; Arnaud, N; Davier, M; Giroux, X; Grosdidier, G; Höcker, A; Le Diberder, F; Lepeltier, V; Lutz, A M; Petersen, T C; Pierini, M; Plaszczynski, S; Rodier, S; Roudeau, P; Schune, M H; Stocchi, A; Wormser, G; Cheng, C H; Lange, D J; Simani, M C; Wright, D M; Bevan, A J; Chavez, C A; Coleman, J P; Forster, I J; Fry, J R; Gabathuler, E; Gamet, R; George, K A; Hutchcroft, D E; Parry, R J; Payne, D J; Touramanis, C; Cormack, C M; Di Lodovico, F; Brown, C L; Cowan, G; Flack, R L; Flaecher, H U; Green, M G; Jackson, P S; McMahon, T R; Ricciardi, S; Salvatore, F; Brown, D; Davis, C L; Allison, J; Barlow, N R; Barlow, R J; Hodgkinson, M C; Lafferty, G D; Naisbit, M T; Williams, J C; Chen, C; Farbin, A; Hulsbergen, W D; Jawahery, A; Kovalskyi, D; Lae, C K; Lillard, V; Roberts, D A; Blaylock, G; Dallapiccola, C; Hertzbach, S S; Kofler, R; Koptchev, V B; Moore, T B; Saremi, S; Staengle, H; Willocq, S; Cowan, R; Koeneke, K; Sciolla, G; Sekula, S J; Taylor, F; Yamamoto, R K; Kim, H; Patel, P M; Robertson, S H; Lazzaro, A; Lombardo, V; Palombo, F; Bauer, J M; Cremaldi, L; Eschenburg, V; Godang, R; Kroeger, R; Reidy, J; Sanders, D A; Summers, D J; Zhao, H W; Brunet, S; Côté, D; Taras, P; Viaud, B; Nicholson, H; Cavallo, N; De Nardo, G; Fabozzi, F; Gatto, C; Lista, L; Monorchio, D; Paolucci, P; Piccolo, D; Sciacca, C; Baak, M; Bulten, H; Raven, G; Snoek, H L; Wilden, L; Jessop, C P; Losecco, J M; Allmendinger, T; Benelli, G; Gan, K K; Honscheid, K; Hufnagel, D; Jackson, P D; Kagan, H; Kass, R; Pulliam, T; Rahimi, A M; Ter-Antonyan, R; Wong, Q K; Brau, J; Frey, R; Igonkina, O; Lu, M; Potter, C T; Sinev, N B; Strom, D; Torrence, E; Colecchia, F; Dorigo, A; Galeazzi, F; Margoni, M; Morandin, M; Posocco, M; Rotondo, M; Simonetto, F; Stroili, R; Voci, C; Benayoun, M; Briand, H; Chauveau, J; David, P; Del Buono, L; de la Vaissière, Ch; Hamon, O; John, M J J; Leruste, Ph; Malclès, J; Ocariz, J; Roos, L; Therin, G; Behera, P K; Gladney, L; Guo, Q H; Panetta, J; Biasini, M; Covarelli, R; Pioppi, M; Angelini, C; Batignani, G; Bettarini, S; Bucci, F; Calderini, G; Carpinelli, M; Forti, F; Giorgi, M A; Lusiani, A; Marchiori, G; Morganti, M; Neri, N; Paoloni, E; Rama, M; Rizzo, G; Simi, G; Walsh, J; Haire, M; Judd, D; Paick, K; Wagoner, D E; Biesiada, J; Danielson, N; Elmer, P; Lau, Y P; Lu, C; Olsen, J; Smith, A J S; Telnov, A V; Bellini, F; Cavoto, G; D'Orazio, A; Di Marco, E; Faccini, R; Ferrarotto, F; Ferroni, F; Gaspero, M; Li Gioi, L; Mazzoni, M A; Morganti, S; Piredda, G; Polci, F; Tehrani, F Safai; Voena, C; Christ, S; Schröder, H; Wagner, G; Waldi, R; Adye, T; De Groot, N; Franek, B; Gopal, G P; Olaiya, E O; Wilson, F F; Aleksan, R; Emery, S; Gaidot, A; Ganzhur, S F; Giraud, P-F; Graziani, G; de Monchenault, G Hamel; Kozanecki, W; Legendre, M; London, G W; Mayer, B; Vasseur, G; Yèche, Ch; Zito, M; Purohit, M V; Weidemann, A W; Wilson, J R; Yumiceva, F X; Abe, T; Allen, M T; Aston, D; Bartoldus, R; Berger, N; Boyarski, A M; Buchmueller, O L; Claus, R; Convery, M R; Cristinziani, M; Dingfelder, J C; Dong, D; Dorfan, J; Dujmic, D; Dunwoodie, W; Fan, S; Field, R C; Glanzman, T; Gowdy, S J; Hadig, T; Halyo, V; Hast, C; Hryn'ova, T; Innes, W R; Kazuhito, S; Kelsey, M H; Kim, P; Kocian, M L; Leith, D W G S; Libby, J; Luitz, S; Luth, V; Lynch, H L; Marsiske, H; Messner, R; Muller, D R; O'grady, C P; Ozcan, V E; Perazzo, A; Perl, M; Ratcliff, B N; Roodman, A; Salnikov, A A; Schindler, R H; Schwiening, J; Snyder, A; Soha, A; Stelzer, J; Strube, J; Su, D; Sullivan, M K; Thompson, J M; Va'vra, J; Wagner, S R; Weaver, M; Wisniewski, W J; Wittgen, M; Wright, D H; Yarritu, A K; Young, C C; Burchat, P R; Edwards, A J; Majewski, S A; Petersen, B A; Roat, C; Ahmed, M; Ahmed, S; Alam, M S; Ernst, J A; Saeed, M A; Saleem, M; Wappler, F R; Bugg, W; Krishnamurthy, M; Spanier, S M; Eckmann, R; Ritchie, J L; Satpathy, A; Schwitters, R F; Izen, J M; Kitayama, I; Lou, X C; Ye, S; Bianchi, F; Bona, M; Gallo, F; Gamba, D; Bomben, M; Bosisio, L; Cartaro, C; Cossutti, F; Della Ricca, G; Dittongo, S; Grancagnolo, S; Lanceri, L; Poropat, P; Vitale, L; Vuagnin, G; Martinez-Vidal, F; Panvini, R S; Banerjee, Sw; Bhuyan, B; Brown, C M; Fortin, D; Hamano, K; Kowalewski, R; Roney, J M; Sobie, R J; Back, J J; Harrison, P F; Latham, T E; Mohanty, G B; Band, H R; Chen, X; Cheng, B; Dasu, S; Datta, M; Eichenbaum, A M; Flood, K T; Graham, M; Hollar, J J; Johnson, J R; Kutter, P E; Li, H; Liu, R; Mellado, B; Mihalyi, A; Pan, Y; Prepost, R; Tan, P; von Wimmersperg-Toeller, J H; Wu, J; Wu, S L; Yu, Z; Greene, M G; Neal, H

    2005-07-22

    We present results from an analysis of B(0)B(0)--> rho(+)rho(-) using 232 x 10(6) Gamma (4S) --> BB decays collected with the BABAR detector at the PEP-II asymmetric-energy B factory at SLAC. We measure the longitudinal polarization fraction f(L) = 0.978 +/- 0.014(stat) + 0.021 / -0.029(syst) and the CP-violating parameters S(L)= -0.33 +/- 0.24(stat) + 0.08 / -0.14(syst) and C(L)= -0.03 +/- 0.18(stat) +/- 0.09(syst). Using an isospin analysis of B --> rhorho decays, we determine the unitarity triangle parameter alpha. The solution compatible with the standard model is alpha = (100 +/- 13) degrees.

  5. Measuring {alpha} in B(t){yields}{rho}{sup {+-}}{pi}{sup {+-}}

    SciTech Connect

    Gronau, M.; Zupan, J.

    2004-10-01

    Defining a most economical parametrization of time-dependent B{yields}{rho}{sup {+-}}{pi}{sup {+-}} decays, including a measurable phase {alpha}{sub eff} which equals the weak phase {alpha} in the limit of vanishing penguin amplitudes, we propose two ways for determining {alpha} in this processes. We explain the limitation of one method, assuming only that two relevant tree amplitudes factorize and that their relative strong phase {delta}{sub t} is negligible. The other method, based on broken flavor SU(3), permits a determination of {alpha} in B{sup 0}{yields}{rho}{sup {+-}}{pi}{sup {+-}} in an over constrained system using also rate measurements of B{sup 0,+}{yields}K*{pi} and B{sup 0,+}{yields}{rho}K. Current data are shown to restrict two ratios of penguin and tree amplitudes r{sub {+-}} to a narrow range around 0.2 and to imply an upper bound |{alpha}{sub eff}-{alpha}|<15 deg. . Assuming that {delta}{sub t} is much smaller than 90 deg. , we find {alpha}=(93{+-}16) deg. and (102{+-}20) deg. using BABAR and BELLE results for B(t){yields}{rho}{sup {+-}}{pi}{sup {+-}}. Avoiding this assumption for completeness, we demonstrate the reduction of discrete ambiguities in {alpha} with increased statistics and show that SU(3) breaking effects are effectively second order in r{sub {+-}}.

  6. Overexpression of Rho GDP-dissociation inhibitor alpha predicts poor survival in oral squamous cell carcinoma.

    PubMed

    Chiang, Wei-Fan; Ho, Hsu-Chueh; Chang, Hua-Yuan; Chiu, Chien-Chih; Chen, Yuh-Ling; Hour, Tzyh-Chyuan; Chuang, Shu-Ju; Wu, Yu-Jen; Chen, Hau-Ren; Chen, Jen-Hao; Liu, Shyun-Yeu; Lu, Chin-Li; Chen, Jeff Yi-Fu

    2011-06-01

    Oral cancer has emerged as one of the fastest growing malignancies in Taiwan. However, biomarkers that reliably predict clinical outcomes have yet to be identified. This study was aimed to identify tumor-associated proteins that could be prognostic biomarkers for oral cancer. We compared the protein expression between oral squamous cell carcinoma (OSCC) tissues and adjacent non-cancerous matched tissues (NCMTs) by proteomics. We found that Rho GDP-dissociation inhibitor alpha (RhoGDIα) was differentially expressed in frozen cancerous samples and OSCC cell lines but not in NCMTs. Furthermore, our results indicated that RhoGDIα was selectively upregulated in 78 OSCC tissue sections (p<0.001), and this high expression was significantly correlated with increased tumor size (p<0.05) and poor overall survival (p<0.01). There was a trend that RhoGDIα expression was localized in the cytoplasm of cancer cells but was localized in the plasma membrane of NCMTs. Finally, expression of RhoGDIα was validated to be an independent prognostic indicator for overall survival (p<0.01). These results have identified a novel biomarker that may be useful for prediction of poor prognosis in OSCC patients.

  7. Limit on the B0-->rho0rho0 branching fraction and implications for the Cabibbo-Kobayashi-Maskawa angle alpha.

    PubMed

    Aubert, B; Barate, R; Boutigny, D; Couderc, F; Karyotakis, Y; Lees, J P; Poireau, V; Tisserand, V; Zghiche, A; Grauges-Pous, E; Palano, A; Pompili, A; Chen, J C; Qi, N D; Rong, G; Wang, P; Zhu, Y S; Eigen, G; Ofte, I; Stugu, B; Abrams, G S; Borgland, A W; Breon, A B; Brown, D N; Button-Shafer, J; Cahn, R N; Charles, E; Day, C T; Gill, M S; Gritsan, A V; Groysman, Y; Jacobsen, R G; Kadel, R W; Kadyk, J; Kerth, L T; Kolomensky, Yu G; Kukartsev, G; Lynch, G; Mir, L M; Oddone, P J; Orimoto, T J; Pripstein, M; Roe, N A; Ronan, M T; Wenzel, W A; Barrett, M; Ford, K E; Harrison, T J; Hart, A J; Hawkes, C M; Morgan, S E; Watson, A T; Fritsch, M; Goetzen, K; Held, T; Koch, H; Lewandowski, B; Pelizaeus, M; Peters, K; Schroeder, T; Steinke, M; Boyd, J T; Burke, J P; Chevalier, N; Cottingham, W N; Kelly, M P; Latham, T E; Wilson, F F; Cuhadar-Donszelmann, T; Hearty, C; Knecht, N S; Mattison, T S; McKenna, J A; Thiessen, D; Khan, A; Kyberd, P; Teodorescu, L; Blinov, A E; Blinov, V E; Druzhinin, V P; Golubev, V B; Ivanchenko, V N; Kravchenko, E A; Onuchin, A P; Serednyakov, S I; Skovpen, Yu I; Solodov, E P; Yushkov, A N; Best, D; Bruinsma, M; Chao, M; Eschrich, I; Kirkby, D; Lankford, A J; Mandelkern, M; Mommsen, R K; Roethel, W; Stoker, D P; Buchanan, C; Hartfiel, B L; Weinstein, A J R; Foulkes, S D; Gary, J W; Long, O; Shen, B C; Wang, K; Del Re, D; Hadavand, H K; Hill, E J; Macfarlane, D B; Paar, H P; Rahatlou, Sh; Sharma, V; Berryhill, J W; Campagnari, C; Cunha, A; Dahmes, B; Hong, T M; Lu, A; Mazur, M A; Richman, J D; Verkerke, W; Beck, T W; Eisner, A M; Flacco, C J; Heusch, C A; Kroseberg, J; Lockman, W S; Nesom, G; Schalk, T; Schumm, B A; Seiden, A; Spradlin, P; Williams, D C; Wilson, M G; Albert, J; Chen, E; Dubois-Felsmann, G P; Dvoretskii, A; Hitlin, D G; Narsky, I; Piatenko, T; Porter, F C; Ryd, A; Samuel, A; Yang, S; Jayatilleke, S; Mancinelli, G; Meadows, B T; Sokoloff, M D; Blanc, F; Bloom, P; Chen, S; Ford, W T; Nauenberg, U; Olivas, A; Rankin, P; Ruddick, W O; Smith, J G; Ulmer, K A; Zhang, J; Zhang, L; Chen, A; Eckhart, E A; Harton, J L; Soffer, A; Toki, W H; Wilson, R J; Zeng, Q; Spaan, B; Altenburg, D; Brandt, T; Brose, J; Dickopp, M; Feltresi, E; Hauke, A; Lacker, H M; Maly, E; Nogowski, R; Otto, S; Petzold, A; Schott, G; Schubert, J; Schubert, K R; Schwierz, R; Sundermann, J E; Bernard, D; Bonneaud, G R; Grenier, P; Schrenk, S; Thiebaux, Ch; Vasileiadis, G; Verderi, M; Bard, D J; Clark, P J; Muheim, F; Playfer, S; Xie, Y; Andreotti, M; Azzolini, V; Bettoni, D; Bozzi, C; Calabrese, R; Cibinetto, G; Luppi, E; Negrini, M; Piemontese, L; Sarti, A; Anulli, F; Baldini-Ferroli, R; Calcaterra, A; de Sangro, R; Finocchiaro, G; Patteri, P; Peruzzi, I M; Piccolo, M; Zallo, A; Buzzo, A; Capra, R; Contri, R; Crosetti, G; Lo Vetere, M; Macri, M; Monge, M R; Passaggio, S; Patrignani, C; Robutti, E; Santroni, A; Tosi, S; Bailey, S; Brandenburg, G; Chaisanguanthum, K S; Morii, M; Won, E; Dubitzky, R S; Langenegger, U; Marks, J; Uwer, U; Bhimji, W; Bowerman, D A; Dauncey, P D; Egede, U; Gaillard, J R; Morton, G W; Nash, J A; Nikolich, M B; Taylor, G P; Charles, M J; Grenier, G J; Mallik, U; Mohapatra, A K; Cochran, J; Crawley, H B; Lamsa, J; Meyer, W T; Prell, S; Rosenberg, E I; Rubin, A E; Yi, J; Arnaud, N; Davier, M; Giroux, X; Grosdidier, G; Höcker, A; Le Diberder, F; Lepeltier, V; Lutz, A M; Petersen, T C; Pierini, M; Plaszczynski, S; Schune, M H; Wormser, G; Cheng, C H; Lange, D J; Simani, M C; Wright, D M; Bevan, A J; Chavez, C A; Coleman, J P; Forster, I J; Fry, J R; Gabathuler, E; Gamet, R; Hutchcroft, D E; Parry, R J; Payne, D J; Touramanis, C; Cormack, C M; Di Lodovico, F; Brown, C L; Cowan, G; Flack, R L; Flaecher, H U; Green, M G; Jackson, P S; McMahon, T R; Ricciardi, S; Salvatore, F; Winter, M A; Brown, D; Davis, C L; Allison, J; Barlow, N R; Barlow, R J; Hodgkinson, M C; Lafferty, G D; Naisbit, M T; Williams, J C; Chen, C; Farbin, A; Hulsbergen, W D; Jawahery, A; Kovalskyi, D; Lae, C K; Lillard, V; Roberts, D A; Blaylock, G; Dallapiccola, C; Hertzbach, S S; Kofler, R; Koptchev, V B; Moore, T B; Saremi, S; Staengle, H; Willocq, S; Cowan, R; Koeneke, K; Sciolla, G; Sekula, S J; Taylor, F; Yamamoto, R K; Patel, P M; Robertson, S H; Lazzaro, A; Lombardo, V; Palombo, F; Bauer, J M; Cremaldi, L; Eschenburg, V; Godang, R; Kroeger, R; Reidy, J; Sanders, D A; Summers, D J; Zhao, H W; Brunet, S; Côté, D; Taras, P; Nicholson, H; Cavallo, N; Fabozzi, F; Gatto, C; Lista, L; Monorchio, D; Paolucci, P; Piccolo, D; Sciacca, C; Baak, M; Bulten, H; Raven, G; Snoek, H L; Wilden, L; Jessop, C P; Losecco, J M; Allmendinger, T; Benelli, G; Gan, K K; Honscheid, K; Hufnagel, D; Kagan, H; Kass, R; Pulliam, T; Rahimi, A M; Ter-Antonyan, R; Wong, Q K; Brau, J; Frey, R; Igonkina, O; Lu, M; Potter, C T; Sinev, N B; Strom, D; Torrence, E; Colecchia, F; Dorigo, A; Galeazzi, F; Margoni, M; Morandin, M; Posocco, M; Rotondo, M; Simonetto, F; Stroili, R; Voci, C; Benayoun, M; Briand, H; Chauveau, J; David, P; Del Buono, L; de la Vaissière, Ch; Hamon, O; John, M J J; Leruste, Ph; Malclès, J; Ocariz, J; Roos, L; Therin, G; Behera, P K; Gladney, L; Guo, Q H; Panetta, J; Biasini, M; Covarelli, R; Pioppi, M; Angelini, C; Batignani, G; Bettarini, S; Bondioli, M; Bucci, F; Calderini, G; Carpinelli, M; Forti, F; Giorgi, M A; Lusiani, A; Marchiori, G; Morganti, M; Neri, N; Paoloni, E; Rama, M; Rizzo, G; Simi, G; Walsh, J; Haire, M; Judd, D; Paick, K; Wagoner, D E; Danielson, N; Elmer, P; Lau, Y P; Lu, C; Miftakov, V; Olsen, J; Smith, A J S; Telnov, A V; Bellini, F; Cavoto, G; D'Orazio, A; Di Marco, E; Faccini, R; Ferrarotto, F; Ferroni, F; Gaspero, M; Li Gioi, L; Mazzoni, M A; Morganti, S; Piredda, G; Polci, F; Safai Tehrani, F; Voena, C; Christ, S; Schröder, H; Wagner, G; Waldi, R; Adye, T; De Groot, N; Franek, B; Gopal, G P; Olaiya, E O; Aleksan, R; Emery, S; Gaidot, A; Ganzhur, S F; Giraud, P-F; Graziani, G; de Monchenault, G Hamel; Kozanecki, W; Legendre, M; London, G W; Mayer, B; Vasseur, G; Yèche, Ch; Zito, M; Purohit, M V; Weidemann, A W; Wilson, J R; Yumiceva, F X; Abe, T; Aston, D; Bartoldus, R; Berger, N; Boyarski, A M; Buchmueller, O L; Claus, R; Convery, M R; Cristinziani, M; De Nardo, G; Dingfelder, J C; Dong, D; Dorfan, J; Dujmic, D; Dunwoodie, W; Fan, S; Field, R C; Glanzman, T; Gowdy, S J; Hadig, T; Halyo, V; Hast, C; Hryn'ova, T; Innes, W R; Kelsey, M H; Kim, P; Kocian, M L; Leith, D W G S; Libby, J; Luitz, S; Luth, V; Lynch, H L; Marsiske, H; Messner, R; Muller, D R; O'grady, C P; Ozcan, V E; Perazzo, A; Perl, M; Ratcliff, B N; Roodman, A; Salnikov, A A; Schindler, R H; Schwiening, J; Snyder, A; Soha, A; Stelzer, J; Strube, J; Su, D; Sullivan, M K; Va'vra, J; Wagner, S R; Weaver, M; Wisniewski, W J; Wittgen, M; Wright, D H; Yarritu, A K; Young, C C; Burchat, P R; Edwards, A J; Majewski, S A; Petersen, B A; Roat, C; Ahmed, M; Ahmed, S; Alam, M S; Ernst, J A; Saeed, M A; Saleem, M; Wappler, F R; Bugg, W; Krishnamurthy, M; Spanier, S M; Eckmann, R; Kim, H; Ritchie, J L; Satpathy, A; Schwitters, R F; Izen, J M; Kitayama, I; Lou, X C; Ye, S; Bianchi, F; Bona, M; Gallo, F; Gamba, D; Bosisio, L; Cartaro, C; Cossutti, F; Ricca, G Della; Dittongo, S; Grancagnolo, S; Lanceri, L; Poropat, P; Vitale, L; Vuagnin, G; Martinez-Vidal, F; Panvini, R S; Banerjee, Sw; Bhuyan, B; Brown, C M; Fortin, D; Hamano, K; Jackson, P D; Kowalewski, R; Roney, J M; Sobie, R J; Back, J J; Harrison, P F; Mohanty, G B; Band, H R; Chen, X; Cheng, B; Dasu, S; Datta, M; Eichenbaum, A M; Flood, K T; Graham, M; Hollar, J J; Johnson, J R; Kutter, P E; Li, H; Liu, R; Mihalyi, A; Pan, Y; Prepost, R; Tan, P; von Wimmersperg-Toeller, J H; Wu, J; Wu, S L; Yu, Z; Greene, M G; Neal, H

    2005-04-08

    We search for the decay B0-->rho(0)rho(0) in a data sample of about 227x10(6) Upsilon(4S)-->BB decays collected with the BABAR detector at the PEP-II asymmetric-energy e(+)e(-) collider at SLAC. We find no significant signal and set an upper limit of 1.1x10(-6) at 90% C.L. on the branching fraction. As a result, the uncertainty due to penguin contributions on the Cabibbo-Kobayashi-Maskawa unitarity angle alpha measured in B-->rhorho decays is decreased to 11 degrees at 68% C.L.

  8. A role for the Rho-p160 Rho coiled-coil kinase axis in the chemokine stromal cell-derived factor-1alpha-induced lymphocyte actomyosin and microtubular organization and chemotaxis.

    PubMed

    Vicente-Manzanares, Miguel; Cabrero, José Román; Rey, Mercedes; Pérez-Martínez, Manuel; Ursa, Angeles; Itoh, Kazuyuki; Sánchez-Madrid, Francisco

    2002-01-01

    The possible involvement of the Rho-p160ROCK (Rho coiled-coil kinase) pathway in the signaling induced by the chemokine Stromal cell-derived factor (SDF)-1alpha has been studied in human PBL. SDF-1alpha induced activation of RhoA, but not that of Rac. RhoA activation was followed by p160ROCK activation mediated by RhoA, which led to myosin light chain (MLC) phosphorylation, which was dependent on RhoA and p160ROCK activities. The kinetics of MLC activation was similar to that of RhoA and p160ROCK. The role of this cascade in overall cell morphology and functional responses to the chemokine was examined employing different chemical inhibitors. Inhibition of either RhoA or p160ROCK did not block SDF-1alpha-induced short-term actin polymerization, but induced the formation of long spikes arising from the cell body, which were found to be microtubule based. This morphological change was associated with an increase in microtubule instability, which argues for an active microtubule polymerization in the formation of these spikes. Inhibition of the Rho-p160ROCK-MLC kinase signaling cascade at different steps blocked lymphocyte migration and the chemotaxis induced by SDF-1alpha. Our results indicate that the Rho-p160ROCK axis plays a pivotal role in the control of the cell shape as a step before lymphocyte migration toward a chemotactic gradient.

  9. Antibodies against Manα1,2-Manα1,2-Man oligosaccharide structures recognize envelope glycoproteins from HIV-1 and SIV strains

    PubMed Central

    Luallen, Robert J; Agrawal-Gamse, Caroline; Fu, Hu; Smith, David F; Doms, Robert W; Geng, Yu

    2010-01-01

    Design of an envelope glycoprotein (Env)-based vaccine against human immunodeficiency virus type-1 (HIV-1) is complicated by the large number of N-linked glycans that coat the protein and serve as a barrier to antibody-mediated neutralization. Compared to normal mammalian glycoproteins, high-mannose-type glycans are disproportionately represented on the gp120 subunit of Env. These N-glycans serve as a target for a number of anti-HIV molecules that bind terminal α1,2-linked mannose residues, including lectins and the monoclonal antibody 2G12. We created a Saccharomyces cerevisiae glycosylation mutant, Δmnn1Δmnn4, to expose numerous terminal Manα1,2-Man residues on endogenous hypermannosylated glycoproteins in the yeast cell wall. Immunization of rabbits with whole cells from this mutant induced antibodies that bound to a broad range of Env proteins, including clade A, B, and C of HIV and simian immunodeficiency virus (SIV). The gp120 binding activity of these immune sera was due to mannose-specific immunoglobulin, as removal of high-mannose glycans and α1,2-linked mannoses from gp120 abrogated serum binding. Glycan array analysis with purified IgG demonstrated binding mainly to glycans with Manα1,2-Manα1,2-Man trisaccharides. Altogether, these data demonstrate the immunogenicity of exposed polyvalent Manα1,2-Manα1,2-Man structures on the yeast cell wall mannan and their ability to induce antibodies that bind to the HIV Env protein. The yeast strain and sera from this study will be useful tools for determining the type of mannose-specific response that is needed to develop neutralizing antibodies to the glycan shield of HIV. PMID:19920089

  10. Measurement of the CKM Angle Alpha at the BABAR Detector Using B Meson Decays to Rho Final States

    SciTech Connect

    Mihalyi, Attila; /Wisconsin U., Madison

    2006-10-16

    This thesis contains the results of an analysis of B{sup 0} {yields} {rho}{sup +}{rho}{sup -} using 232 million {Upsilon}(4S) {yields} B{bar B} decays collected with the BABAR detector at the PEP-II asymmetric-energy B Factory at SLAC. From a fitted signal yield of 617 {+-} 52 events, the longitudinal polarizations fraction, f{sub L}, of the decay is measured to be 0.978 {+-} 0.014(stat){sub -0.029}{sup +0.021}(syst). The nearly fully longitudinal dominance of the B{sup 0} {yields} {rho}{sup +}{rho}{sup -} decay allows for a measurement of the time dependent CP parameters S{sub L} and C{sub L}, where the first parameter is sensitive to mixing induced CP violation and the second one to direct CP violation. From the same signal yield, these values are found to be S{sub L} = -0.33 {+-} 0.24(stat){sub -0.14}{sup +0.08}(syst) and C{sub L} = - 0.03 {+-} 0.18(stat) {+-} 0.09(syst). The CKM angle {alpha} is then determined, using these results and the branching fractions and polarizations of the decays B{sup 0} {yields} {rho}{sup 0}{rho}{sup 0} and B{sup +} {yields} {rho}{sup +}{rho}{sup 0}. This measurement is done with an isospin analysis, in which a triangle is constructed from the isospin amplitudes of these three decay modes. A {chi}{sup 2} expression that includes the measured quantities expressed as the lengths of the sides of the isospin triangles is constructed and minimized to determine a confidence level on {alpha}. Selecting the solution compatible with the Standard Model, one obtains {alpha} = 100{sup o} {+-} 13{sup o}.

  11. Study of the decay B0(B- 0)-->rho+rho-, and constraints on the Cabibbo-Kobayashi-Maskawa angle alpha.

    PubMed

    Aubert, B; Barate, R; Boutigny, D; Couderc, F; Gaillard, J-M; Hicheur, A; Karyotakis, Y; Lees, J P; Tisserand, V; Zghiche, A; Palano, A; Pompili, A; Chen, J C; Qi, N D; Rong, G; Wang, P; Zhu, Y S; Eigen, G; Ofte, I; Stugu, B; Abrams, G S; Borgland, A W; Breon, A B; Brown, D N; Button-Shafer, J; Cahn, R N; Charles, E; Day, C T; Gill, M S; Gritsan, A V; Groysman, Y; Jacobsen, R G; Kadel, R W; Kadyk, J; Kerth, L T; Kolomensky, Yu G; Kukartsev, G; LeClerc, C; Lynch, G; Merchant, A M; Mir, L M; Oddone, P J; Orimoto, T J; Pripstein, M; Roe, N A; Ronan, M T; Shelkov, V G; Wenzel, W A; Ford, K; Harrison, T J; Hawkes, C M; Morgan, S E; Watson, A T; Fritsch, M; Goetzen, K; Held, T; Koch, H; Lewandowski, B; Pelizaeus, M; Steinke, M; Boyd, J T; Chevalier, N; Cottingham, W N; Kelly, M P; Latham, T E; Wilson, F F; Cuhadar-Donszelmann, T; Hearty, C; Knecht, N S; Mattison, T S; McKenna, J A; Thiessen, D; Khan, A; Kyberd, P; Teodorescu, L; Blinov, V E; Bukin, A D; Druzhinin, V P; Golubev, V B; Ivanchenko, V N; Kravchenko, E A; Onuchin, A P; Serednyakov, S I; Skovpen, Yu I; Solodov, E P; Yushkov, A N; Best, D; Bruinsma, M; Chao, M; Eschrich, I; Kirkby, D; Lankford, A J; Mandelkern, M; Mommsen, R K; Roethel, W; Stoker, D P; Buchanan, C; Hartfiel, B L; Gary, J W; Shen, B C; Wang, K; del Re, D; Hadavand, H K; Hill, E J; MacFarlane, D B; Paar, H P; Rahatlou, Sh; Sharma, V; Berryhill, J W; Campagnari, C; Dahmes, B; Levy, S L; Long, O; Lu, A; Mazur, M A; Richman, J D; Verkerke, W; Beck, T W; Eisner, A M; Heusch, C A; Lockman, W S; Schalk, T; Schmitz, R E; Schumm, B A; Seiden, A; Spradlin, P; Williams, D C; Wilson, M G; Albert, J; Chen, E; Dubois-Felsmann, G P; Dvoretskii, A; Hitlin, D G; Narsky, I; Piatenko, T; Porter, F C; Ryd, A; Samuel, A; Yang, S; Jayatilleke, S; Mancinelli, G; Meadows, B T; Sokoloff, M D; Abe, T; Blanc, F; Bloom, P; Chen, S; Ford, W T; Nauenberg, U; Olivas, A; Rankin, P; Smith, J G; Zhang, J; Zhang, L; Chen, A; Harton, J L; Soffer, A; Toki, W H; Wilson, R J; Zeng, Q L; Altenburg, D; Brandt, T; Brose, J; Colberg, T; Dickopp, M; Feltresi, E; Hauke, A; Lacker, H M; Maly, E; Müller-Pfefferkorn, R; Nogowski, R; Otto, S; Petzold, A; Schubert, J; Schubert, K R; Schwierz, R; Spaan, B; Sundermann, J E; Bernard, D; Bonneaud, G R; Brochard, F; Grenier, P; Schrenk, S; Thiebaux, Ch; Vasileiadis, G; Verderi, M; Bard, D J; Clark, P J; Lavin, D; Muheim, F; Playfer, S; Xie, Y; Andreotti, M; Azzolini, V; Bettoni, D; Bozzi, C; Calabrese, R; Cibinetto, G; Luppi, E; Negrini, M; Piemontese, L; Sarti, A; Treadwell, E; Baldini-Ferroli, R; Calcaterra, A; de Sangro, R; Finocchiaro, G; Patteri, P; Piccolo, M; Zallo, A; Buzzo, A; Capra, R; Contri, R; Crosetti, G; Lo Vetere, M; Macri, M; Monge, M R; Passaggio, S; Patrignani, C; Robutti, E; Santroni, A; Tosi, S; Bailey, S; Brandenburg, G; Morii, M; Won, E; Dubitzky, R S; Langenegger, U; Bhimji, W; Bowerman, D A; Dauncey, P D; Egede, U; Gaillard, J R; Morton, G W; Nash, J A; Taylor, G P; Grenier, G J; Mallik, U; Cochran, J; Crawley, H B; Lamsa, J; Meyer, W T; Prell, S; Rosenberg, E I; Yi, J; Davier, M; Grosdidier, G; Höcker, A; Laplace, S; Le Diberder, F; Lepeltier, V; Lutz, A M; Petersen, T C; Plaszczynski, S; Schune, M H; Tantot, L; Wormser, G; Cheng, C H; Lange, D J; Simani, M C; Wright, D M; Bevan, A J; Coleman, J P; Fry, J R; Gabathuler, E; Gamet, R; Parry, R J; Payne, D J; Sloane, R J; Touramanis, C; Back, J J; Cormack, C M; Harrison, P F; Mohanty, G B; Brown, C L; Cowan, G; Flack, R L; Flaecher, H U; Green, M G; Marker, C E; McMahon, T R; Ricciardi, S; Salvatore, F; Vaitsas, G; Winter, M A; Brown, D; Davis, C L; Allison, J; Barlow, N R; Barlow, R J; Hart, P A; Hodgkinson, M C; Lafferty, G D; Lyon, A J; Williams, J C; Farbin, A; Hulsbergen, W D; Jawahery, A; Kovalskyi, D; Lae, C K; Lillard, V; Roberts, D A; Blaylock, G; Dallapiccola, C; Flood, K T; Hertzbach, S S; Kofler, R; Koptchev, V B; Moore, T B; Saremi, S; Staengle, H; Willocq, S; Cowan, R; Sciolla, G; Taylor, F; Yamamoto, R K; Mangeol, D J J; Patel, P M; Robertson, S H; Lazzaro, A; Palombo, F; Bauer, J M; Cremaldi, L; Eschenburg, V; Godang, R; Kroeger, R; Reidy, J; Sanders, D A; Summers, D J; Zhao, H W; Brunet, S; Côté, D; Taras, P; Nicholson, H; Cavallo, N; Fabozzi, F; Gatto, C; Lista, L; Monorchio, D; Paolucci, P; Piccolo, D; Sciacca, C; Baak, M; Bulten, H; Raven, G; Wilden, L; Jessop, C P; LoSecco, J M; Gabriel, T A; Allmendinger, T; Brau, B; Gan, K K; Honscheid, K; Hufnagel, D; Kagan, H; Kass, R; Pulliam, T; Rahimi, A M; Ter-Antonyan, R; Wong, Q K; Brau, J; Frey, R; Igonkina, O; Potter, C T; Sinev, N B; Strom, D; Torrence, E; Colecchia, F; Dorigo, A; Galeazzi, F; Margoni, M; Morandin, M; Posocco, M; Rotondo, M; Simonetto, F; Stroili, R; Tiozzo, G; Voci, C; Benayoun, M; Briand, H; Chauveau, J; David, P; de la Vaissière, Ch; Del Buono, L; Hamon, O; John, M J J; Leruste, Ph; Ocariz, J; Pivk, M; Roos, L; T'Jampens, S; Therin, G; Manfredi, P F; Re, V; Behera, P K; Gladney, L; Guo, Q H; Panetta, J; Anulli, F; Biasini, M; Peruzzi, I M; Pioppi, M; Angelini, C; Batignani, G; Bettarini, S; Bondioli, M; Bucci, F; Calderini, G; Carpinelli, M; Del Gamba, V; Forti, F; Giorgi, M A; Lusiani, A; Marchiori, G; Martinez-Vidal, F; Morganti, M; Neri, N; Paoloni, E; Rama, M; Rizzo, G; Sandrelli, F; Walsh, J; Haire, M; Judd, D; Paick, K; Wagoner, D E; Danielson, N; Elmer, P; Lau, Y P; Lu, C; Miftakov, V; Olsen, J; Smith, A J S; Telnov, A V; Bellini, F; Cavoto, G; Faccini, R; Ferrarotto, F; Ferroni, F; Gaspero, M; Li Gioi, L; Mazzoni, M A; Morganti, S; Pierini, M; Piredda, G; Safai Tehrani, F; Voena, C; Christ, S; Wagner, G; Waldi, R; Adye, T; De Groot, N; Franek, B; Geddes, N I; Gopal, G P; Olaiya, E O; Aleksan, R; Emery, S; Gaidot, A; Ganzhur, S F; Giraud, P-F; Hamel de Monchenault, G; Kozanecki, W; Langer, M; Legendre, M; London, G W; Mayer, B; Schott, G; Vasseur, G; Yèche, Ch; Zito, M; Purohit, M V; Weidemann, A W; Yumiceva, F X; Aston, D; Bartoldus, R; Berger, N; Boyarski, A M; Buchmueller, O L; Convery, M R; Cristinziani, M; De Nardo, G; Dong, D; Dorfan, J; Dujmic, D; Dunwoodie, W; Elsen, E E; Fan, S; Field, R C; Glanzman, T; Gowdy, S J; Hadig, T; Halyo, V; Hast, C; Hryn'ova, T; Innes, W R; Kelsey, M H; Kim, P; Kocian, M L; Leith, D W G S; Libby, J; Luitz, S; Luth, V; Lynch, H L; Marsiske, H; Messner, R; Muller, D R; O'Grady, C P; Ozcan, V E; Perazzo, A; Perl, M; Petrak, S; Ratcliff, B N; Roodman, A; Salnikov, A A; Schindler, R H; Schwiening, J; Simi, G; Snyder, A; Soha, A; Stelzer, J; Su, D; Sullivan, M K; Va'vra, J; Wagner, S R; Weaver, M; Weinstein, A J R; Wisniewski, W J; Wittgen, M; Wright, D H; Yarritu, A K; Young, C C; Burchat, P R; Edwards, A J; Meyer, T I; Petersen, B A; Roat, C; Ahmed, S; Alam, M S; Ernst, J A; Saeed, M A; Saleem, M; Wappler, F R; Bugg, W; Krishnamurthy, M; Spanier, S M; Eckmann, R; Kim, H; Ritchie, J L; Satpathy, A; Schwitters, R F; Izen, J M; Kitayama, I; Lou, X C; Ye, S; Bianchi, F; Bona, M; Gallo, F; Gamba, D; Borean, C; Bosisio, L; Cartaro, C; Cossutti, F; Della Ricca, G; Dittongo, S; Grancagnolo, S; Lanceri, L; Poropat, P; Vitale, L; Vuagnin, G; Panvini, R S; Banerjee, Sw; Brown, C M; Fortin, D; Jackson, P D; Kowalewski, R; Roney, J M; Band, H R; Dasu, S; Datta, M; Eichenbaum, A M; Graham, M; Hollar, J J; Johnson, J R; Kutter, P E; Li, H; Liu, R; Di Lodovico, F; Mihalyi, A; Mohapatra, A K; Pan, Y; Prepost, R; Rubin, A E; Sekula, S J; Tan, P; von Wimmersperg-Toeller, J H; Wu, J; Wu, S L; Yu, Z; Neal, H

    2004-12-03

    Using a data sample of 89 x 10(6) Upsilon(4S)-->BB decays collected with the BABAR detector at the PEP-II asymmetric B Factory at SLAC, we measure the B0(B (0))-->rho(+)rho(-) branching fraction as [30+/-4(stat)+/-5(syst)]x10(-6) and a longitudinal polarization fraction of f(L)=0.99+/-0.03(stat)+0.04-0.03(syst). We measure the time-dependent-asymmetry parameters of the longitudinally polarized component of this decay as C(L)=-0.17+/-0.27(stat)+/-0.14(syst) and S(L)=-0.42+/-0.42(stat)+/-0.14(syst). We exclude values of alpha between 19 degrees and 71 degrees (90% C.L.).

  12. Activation of p115-RhoGEF Requires Direct Association of G[alpha subscript 13] and the Dbl Homology Domain

    SciTech Connect

    Chen, Zhe; Guo, Liang; Hadas, Jana; Gutowski, Stephen; Sprang, Stephen R.; Sternweis, Paul C.

    2012-09-05

    RGS-containing RhoGEFs (RGS-RhoGEFs) represent a direct link between the G{sub 12} class of heterotrimeric G proteins and the monomeric GTPases. In addition to the canonical Dbl homology (DH) and pleckstrin homology domains that carry out the guanine nucleotide exchange factor (GEF) activity toward RhoA, these RhoGEFs also possess RGS homology (RH) domains that interact with activated {alpha} subunits of G{sub 12} and G{sub 13}. Although the GEF activity of p115-RhoGEF (p115), an RGS-RhoGEF, can be stimulated by G{alpha}{sub 13}, the exact mechanism of the stimulation has remained unclear. Using combined studies with small angle x-ray scattering, biochemistry, and mutagenesis, we identify an additional binding site for activated G{alpha}{sub 13} in the DH domain of p115. Small angle x-ray scattering reveals that the helical domain of G{alpha}{sub 13} docks onto the DH domain, opposite to the surface of DH that binds RhoA. Mutation of a single tryptophan residue in the {alpha}3b helix of DH reduces binding to activated G{alpha}{sub 13} and ablates the stimulation of p115 by G{alpha}{sub 13}. Complementary mutations at the predicted DH-binding site in the {alpha}B-{alpha}C loop of the helical domain of G{alpha}{sub 13} also affect stimulation of p115 by G{alpha}{sub 13}. Although the GAP activity of p115 is not required for stimulation by G{alpha}{sub 13}, two hydrophobic motifs in RH outside of the consensus RGS box are critical for this process. Therefore, the binding of G{alpha}{sub 13} to the RH domain facilitates direct association of G{alpha}{sub 13} to the DH domain to regulate its exchange activity. This study provides new insight into the mechanism of regulation of the RGS-RhoGEF and broadens our understanding of G protein signaling.

  13. Expected accuracy in a measurement of the CKM angle alpha using a Dalitz plot analysis of B0 ---> rho pi decays in the BTeV project

    SciTech Connect

    Shestermanov, K.E.; Vasiliev, A.N; Butler, J.; Derevschikov, A.A.; Kasper, P.; Kiselev, V.V.; Kravtsov, V.I.; Kubota, Y.; Kutschke, R.; Matulenko, Y.A.; Minaev, N.G.; /Serpukhov, IHEP /Fermilab /Minnesota U. /Syracuse U. /INFN, Milan

    2005-12-01

    A precise measurement of the angle {alpha} in the CKM triangle is very important for a complete test of Standard Model. A theoretically clean method to extract {alpha} is provided by B{sup 0} {yields} {rho}{pi} decays. Monte Carlo simulations to obtain the BTeV reconstruction efficiency and to estimate the signal to background ratio for these decays were performed. Finally the time-dependent Dalitz plot analysis, using the isospin amplitude formalism for tre and penguin contributions, was carried out. It was shown that in one year of data taking BTeV could achieve an accuracy on {alpha} better than 5{sup o}.

  14. Cooling augments vasoconstriction mediated by 5-HT1 and alpha2-adrenoceptors in the isolated equine digital vein: involvement of Rho kinase.

    PubMed

    Zerpa, Hector; Berhane, Yoel; Elliott, Jonathan; Bailey, Simon R

    2007-08-27

    The vasculature of the equine digit fulfils an important role in thermoregulation. In other species, it has been found that cooling may enhance the response of cutaneous vessels to 5-hydroxytryptamine (5-HT) and alpha(2)-adrenoceptor agonists. Translocation of alpha(2)-adrenoceptors to the smooth muscle cell membrane, mediated by Rho kinase, is thought to be involved in the cooling-enhanced response in mouse tail arteries. However, little is known about the effect of cooling on 5-HT receptor function. The present investigation compared the response of 5-bromo-6-(2-imidazolin-2-ylamino) quinoxaline (UK14304:1 nM to 30 microM), methoxamine (0.1 nM to 30 microM; in the presence of yohimbine 0.1 microM), 5-carboxamidotryptamine (5-CT; 0.1 nM to 10 microM) and alpha-methyl 5-HT (0.1 nM to 10 microM) in the isolated equine digital vein at 30 degrees C and 22 degrees C. The effect of the Rho kinase inhibitor, fasudil (1 microM), and the recovery of the response after the irreversible blockade of surface receptors with phenoxybenzamine (10 microM) or 2-ethoxy-1-ethoxycarbonyl-1,2-dihydroquinoline (EEDQ;10 microM), was established. Moderate cooling significantly increased the maximum response to alpha-methyl 5-HT, 5-CT and UK14304 and shifted their response curves to the left. Cooling also augmented the phenoxybenzamine- and EEDQ-resistant response to UK14304 and 5-CT, respectively. Fasudil had no effect on the contractile response at 30 degrees C, but completely abrogated the effect of cooling on the response to 5-CT and UK14304. The response to methoxamine was not significantly affected by cooling. These results suggest that Rho kinase plays an important role in the cooling-enhanced response mediated by 5-HT(1B/D) receptors and alpha(2)-adrenoceptors. The exact mechanism by which Rho/Rho kinase enhances the functional responses mediated by these receptors in these vessels has yet to be determined.

  15. Constitutive activation of NF-kappa B and secretion of interleukin-8 induced by the G protein-coupled receptor of Kaposi's sarcoma-associated herpesvirus involve G alpha(13) and RhoA.

    PubMed

    Shepard, L W; Yang, M; Xie, P; Browning, D D; Voyno-Yasenetskaya, T; Kozasa, T; Ye, R D

    2001-12-07

    The Kaposi's sarcoma herpesvirus (KSHV) open reading frame 74 encodes a G protein-coupled receptor (GPCR) for chemokines. Exogenous expression of this constitutively active GPCR leads to cell transformation and vascular overgrowth characteristic of Kaposi's sarcoma. We show here that expression of KSHV-GPCR in transfected cells results in constitutive transactivation of nuclear factor kappa B (NF-kappa B) and secretion of interleukin-8, and this response involves activation of G alpha(13) and RhoA. The induced expression of a NF-kappa B luciferase reporter was partially reduced by pertussis toxin and the G beta gamma scavenger transducin, and enhanced by co-expression of G alpha(13) and to a lesser extent, G alpha(q). These results indicate coupling of KSHV-GPCR to multiple G proteins for NF-kappa B activation. Expression of KSHV-GPCR led to stress fiber formation in NIH 3T3 cells. To examine the involvement of the G alpha(13)-RhoA pathway in KSHV-GPCR-mediated NF-kappa B activation, HeLa cells were transfected with KSHV-GPCR alone and in combination with the regulator of G protein signaling (RGS) from p115RhoGEF or a dominant negative RhoA(T19N). Both constructs, as well as the C3 exoenzyme from Clostritium botulinum, partially reduced NF-kappa B activation by KSHV-GPCR, and by a constitutively active G alpha(13)(Q226L). KSHV-GPCR-induced NF-kappa B activation is accompanied by increased secretion of IL-8, a function mimicked by the activated G alpha(13) but not by an activated G alpha(q)(Q209L). These results suggest coupling of KSHV-GPCR to the G alpha(13)-RhoA pathway in addition to other G proteins.

  16. Hop rho iso-alpha acids, berberine, vitamin D3 and vitamin K1 favorably impact biomarkers of bone turnover in postmenopausal women in a 14-week trial.

    PubMed

    Holick, Michael F; Lamb, Joseph J; Lerman, Robert H; Konda, Veera R; Darland, Gary; Minich, Deanna M; Desai, Anuradha; Chen, Tai C; Austin, Melissa; Kornberg, Jacob; Chang, Jyh-Lurn; Hsi, Alex; Bland, Jeffrey S; Tripp, Matthew L

    2010-05-01

    Osteoporosis is a major health issue facing postmenopausal women. Increased production of pro-inflammatory cytokines resulting from declining estrogen leads to increased bone resorption. Nutrition can have a positive impact on osteoporosis prevention and amelioration. The objective of this study was to investigate the impact of targeted phytochemicals and nutrients essential for bone health on bone turnover markers in healthy postmenopausal women. In this 14-week, single-blinded, 2-arm placebo-controlled pilot study, all women were instructed to consume a modified Mediterranean-style low-glycemic-load diet and to engage in limited aerobic exercise; 17 randomized to the placebo and 16 to the treatment arm (receiving 200 mg hop rho iso-alpha acids, 100 mg berberine sulfate trihydrate, 500 IU vitamin D(3) and 500 microg vitamin K(1), twice daily). Thirty-two women completed the study. Baseline nutrient intake did not differ between arms. At 14 weeks, the treatment arm exhibited an estimated 31% mean reduction (P = 0.02) in serum osteocalcin (a marker of bone turnover), whereas the placebo arm exhibited a 19% increase (P = 0.03) compared to baseline. Serum 25-hydroxyvitamin D (25(OH)D) increased by 13% (P = 0.24) in the treatment arm and decreased by 25% (P < 0.01) in the placebo arm. The between-arm differences for OC and 25(OH)D were statistically significant. Serum IGF-I was increased in both arms, but the increase was more significant in the treatment arm at 14 weeks (P < 0.01). Treatment with hop rho iso-alpha acids, berberine sulfate trihydrate, vitamin D(3) and vitamin K(1) produced a more favorable bone biomarker profile that supports a healthy bone metabolism.

  17. Affinophoresis of pea lectin and fava bean lectin with an anionic affinophore, bearing rho-aminophenyl-alpha-D-mannoside as an affinity ligand.

    PubMed

    Shimura, K; Kasai, K

    1987-07-29

    Affinophoresis is an electrophoretic separation technique for biological polymers with the aid of an affinophore, which is a macromolecular polyelectrolyte bearing affinity ligands. The affinophore migrates rapidly in an electric field, and consequently the electrophoretic mobility of molecules having an affinity for the ligand is specifically changed. An anionic affinophore-bearing mannosyl residue was synthesized for the affinophoresis of lectins. rho-Aminophenyl-alpha-D-mannopyranoside and aminomethanesulphonic acid were coupled to about one-tenth and one-fifth, respectively, of the carboxyl groups of succinyl-poly-L-lysine with an average degree of polymerization of 120 by the use of a water-soluble carbodiimide. Extracts of seeds of pea (Pisum sativum) or fava bean (Vicia fava) were subjected to two-dimensional agarose gel electrophoresis, in which the first dimension was ordinary agarose gel electrophoresis and the second dimension was affinophoresis with the affinophore. The separated proteins were stained with Coomassie Blue R250. The lectins in both seed extracts were separated from a diagonal line formed by other proteins in the extracts. About 10 ng of the separated pea lectin was detected on a nitrocellulose blot by immunostaining with a horseradish peroxidase-conjugated second antibody.

  18. Crystallization and preliminary X-ray analysis of the Man(α1-2)Man-specific lectin from Bowringia mildbraedii in complex with its carbohydrate ligand

    SciTech Connect

    Garcia-Pino, Abel; Loris, Remy; Wyns, Lode; Buts, Lieven

    2005-10-01

    The lectin from the Nigerian legume B. mildbraedii was crystallized in complex with Man(α1-2)Man and data were collected to a resolution of 1.90 Å using synchrotron radiation. The lectin from Bowringia mildbraedii seeds crystallizes in the presence of the disaccharide Man(α1-2)Man. The best crystals grow at 293 K within four weeks after a pre-incubation at 277 K to induce nucleation. A complete data set was collected to a resolution of 1.90 Å using synchrotron radiation. The crystals belong to space group I222, with unit-cell parameters a = 66.06, b = 86.35, c = 91.76 Å, and contain one lectin monomer in the asymmetric unit.

  19. Identification of critical residues in G(alpha)13 for stimulation of p115RhoGEF activity and the structure of the G(alpha)13-p115RhoGEF regulator of G protein signaling homology (RH) domain complex.

    PubMed

    Hajicek, Nicole; Kukimoto-Niino, Mutsuko; Mishima-Tsumagari, Chiemi; Chow, Christina R; Shirouzu, Mikako; Terada, Takaho; Patel, Maulik; Yokoyama, Shigeyuki; Kozasa, Tohru

    2011-06-10

    RH-RhoGEFs are a family of guanine nucleotide exchange factors that contain a regulator of G protein signaling homology (RH) domain. The heterotrimeric G protein Gα(13) stimulates the guanine nucleotide exchange factor (GEF) activity of RH-RhoGEFs, leading to activation of RhoA. The mechanism by which Gα(13) stimulates the GEF activity of RH-RhoGEFs, such as p115RhoGEF, has not yet been fully elucidated. Here, specific residues in Gα(13) that mediate activation of p115RhoGEF are identified. Mutation of these residues significantly impairs binding of Gα(13) to p115RhoGEF as well as stimulation of GEF activity. These data suggest that the exchange activity of p115RhoGEF is stimulated allosterically by Gα(13) and not through its interaction with a secondary binding site. A crystal structure of Gα(13) bound to the RH domain of p115RhoGEF is also presented, which differs from a previously crystallized complex with a Gα(13)-Gα(i1) chimera. Taken together, these data provide new insight into the mechanism by which p115RhoGEF is activated by Gα(13).

  20. Search for the Decay B{sup 0} --> {rho}{sup 0} {rho}{sup 0}

    SciTech Connect

    Aubert, B

    2004-08-16

    The B{sup 0} --> {rho}{sup 0}{rho}{sup 0} decay mode is searched for in a data sample of about 227 million {Upsilon}(4S) --> B{bar B} decays collected with the BABAR detector at the PEP-II asymmetric B factory at SLAC. No significant signal is observed, and an upper limit of 1.1x10{sup -6} (90% C.L.) on the branching fraction is set. Implications on the penguin contribution and constraints on the CKM angle {alpha} with B --> {rho}{rho} decays are discussed. All results are preliminary.

  1. Structures of sugar chains of the subunits of an alpha-amylase inhibitor from Phaseolus vulgaris white kidney beans.

    PubMed

    Yamaguchi, H; Funaoka, H; Iwamoto, H

    1992-03-01

    The structures of asparagine-linked oligosaccharides in the subunits of an alpha-amylase inhibitor from the white kidney bean (Phaseolus vulgaris) were determined. Glycopeptides obtained from each subunit were treated with hydrazine, then N-acetylated. The oligosaccharides thus liberated were labeled with 2-aminopyridine at their reducing ends and purified by gel-permeation, reverse-phase, and size-fractionation HPLC. The structures of seven oligosaccharides from the alpha-subunit and eight oligosaccharides from the beta-subunit were determined by a combination of composition and molecular size analyses, exo- and endoglycosidase digestions, partial acetolysis, and 1H-NMR spectroscopy. The major glycan chains in the alpha-subunit were Man alpha 1-6(Man alpha 1-3)Man alpha 1-6(Man alpha 1-2Man alpha 1-3)-Man beta 1-4GlcNAc beta 1-4GlcNAc and (Man alpha 1-2)Man alpha 1-6(Man alpha 1-2Man alpha 1-3)Man alpha 1-6 (Man alpha 1-2Man alpha 1-2Man alpha 1-3)Man beta 1-4GlcNAc beta 1-4GlcNAc, while a glycan chain Man alpha 1-6(Man alpha 1-3)(Xyl beta 1-2)Man beta 1-4GlcNAc beta 1-4GlcNAc comprised more than 70% of the sugar moiety of the beta-subunit.

  2. Optimized mixture of hops rho iso-alpha acids-rich extract and acacia proanthocyanidins-rich extract reduces insulin resistance in 3T3-L1 adipocytes and improves glucose and insulin control in db/db mice

    PubMed Central

    Darland, Gary; Konda, Veera Reddy; Pacioretty, Linda M.; Chang, Jyh-Lurn; Bland, Jeffrey S.; Babish, John G.

    2012-01-01

    Rho iso-alpha acids-rich extract (RIAA) from Humulus lupulus (hops) and proanthocyanidins-rich extracts (PAC) from Acacia nilotica exert anti-inflammatory and anti-diabetic activity in vitro and in vivo. We hypothesized that a combination of these two extracts would exert enhanced effects in vitro on inflammatory markers and insulin signaling, and on nonfasting glucose and insulin in db/db mice. Over 49 tested combinations, RIAA:PAC at 5:1 (6.25 µg/mL) exhibited the greatest reductions in TNFα-stimulated lipolysis and IL-6 release in 3T3-L1 adipocytes, comparable to 5 µg/mL troglitazone. Pretreatment of 3T3-L1 adipocytes with this combination (5 µg/mL) also led to a 3-fold increase in insulin-stimulated glucose uptake that was comparable to 5 µg/mL pioglitazone or 901 µg/mL aspirin. Finally, db/db mice fed with RIAA:PAC at 5:1 (100 mg/kg) for 7 days resulted in 22% decrease in nonfasting glucose and 19% decrease in insulin that was comparable to 0.5 mg/kg rosiglitazone and better than 100 mg/kg metformin. RIAA:PAC mixture may have the potential to be an alternative when conventional therapy is undesirable or ineffective, and future research exploring its long-term clinical application is warranted. PMID:23198019

  3. Recent Results on the CKM Angle Alpha

    SciTech Connect

    Mihalyi, A.; /Wisconsin U., Madison

    2005-10-18

    The method to measure the CKM angle {alpha} and the modes sensitive to it are discussed. It is shown that the B {yields} {rho}{rho} decays provide the most stringent constraint on {alpha}, which is found to be {alpha} = 96{sup o} {+-} 10{sup o}(stat) {+-} 4{sup o}(syst){+-} 13{sup o}(penguin).

  4. Rho GAPs and GEFs

    PubMed Central

    van Buul, Jaap D; Geerts, Dirk; Huveneers, Stephan

    2014-01-01

    Within blood vessels, endothelial cell–cell and cell–matrix adhesions are crucial to preserve barrier function, and these adhesions are tightly controlled during vascular development, angiogenesis, and transendothelial migration of inflammatory cells. Endothelial cellular signaling that occurs via the family of Rho GTPases coordinates these cell adhesion structures through cytoskeletal remodelling. In turn, Rho GTPases are regulated by GTPase-activating proteins (GAPs) and guanine nucleotide exchange factors (GEFs). To understand how endothelial cells initiate changes in the activity of Rho GTPases, and thereby regulate cell adhesion, we will discuss the role of Rho GAPs and GEFs in vascular biology. Many potentially important Rho regulators have not been studied in detail in endothelial cells. We therefore will first overview which GAPs and GEFs are highly expressed in endothelium, based on comparative gene expression analysis of human endothelial cells compared with other tissue cell types. Subsequently, we discuss the relevance of Rho GAPs and GEFs for endothelial cell adhesion in vascular homeostasis and disease. PMID:24622613

  5. Measurement of the angle alpha at BABAR

    SciTech Connect

    Perez, A.; /Orsay, LAL

    2009-06-25

    The authors present recent measurements of the CKM angle {alpha} using data collected by the BABAR detector at the PEP-II asymmetric-energy e{sup +}e{sup -} collider at the SLAC National Accelerator Laboratory, operating at the {Upsilon}(4S) resonance. They present constraints on {alpha} from B {yields} {pi}{pi}, B {yields} {rho}{rho} and B {yields} {rho}{pi} decays.

  6. The Rho Kappa Spirit

    ERIC Educational Resources Information Center

    McCullagh, Mary T.

    2012-01-01

    Many years ago, Christopher Columbus High School opened a chapter of Rho Kappa, the Social Studies Honor Society developed through the Florida Council for the Social Studies (FCSS). As department chair and member of FCSS, the author was thrilled to be able to offer their students opportunities in the Honor Society for the Social Studies. They…

  7. RhoA/Rho-Kinase in the Cardiovascular System.

    PubMed

    Shimokawa, Hiroaki; Sunamura, Shinichiro; Satoh, Kimio

    2016-01-22

    Twenty years ago, Rho-kinase was identified as an important downstream effector of the small GTP-binding protein, RhoA. Thereafter, a series of studies demonstrated the important roles of Rho-kinase in the cardiovascular system. The RhoA/Rho-kinase pathway is now widely known to play important roles in many cellular functions, including contraction, motility, proliferation, and apoptosis, and its excessive activity induces oxidative stress and promotes the development of cardiovascular diseases. Furthermore, the important role of Rho-kinase has been demonstrated in the pathogenesis of vasospasm, arteriosclerosis, ischemia/reperfusion injury, hypertension, pulmonary hypertension, and heart failure. Cyclophilin A is secreted by vascular smooth muscle cells and inflammatory cells and activated platelets in a Rho-kinase-dependent manner, playing important roles in a wide range of cardiovascular diseases. Thus, the RhoA/Rho-kinase pathway plays crucial roles under both physiological and pathological conditions and is an important therapeutic target in cardiovascular medicine. Recently, functional differences between ROCK1 and ROCK2 have been reported in vitro. ROCK1 is specifically cleaved by caspase-3, whereas granzyme B cleaves ROCK2. However, limited information is available on the functional differences and interactions between ROCK1 and ROCK2 in the cardiovascular system in vivo. Herein, we will review the recent advances about the importance of RhoA/Rho-kinase in the cardiovascular system.

  8. Measurements of Branching Ratios And Search for CP Violation in the Modes B0 to Rho Pi, Rho K

    SciTech Connect

    Laplace, Sandrine; /Paris U., VI-VII

    2006-09-18

    The BABAR experiment, at the PEP-II collider at SLAC, has been studying since 1999 CP violation in the B meson system. After the precise measurement of sin2{beta}, one is now concentrating on measuring the angles {alpha} and {gamma} of the unitarity triangle. The work presented in this thesis concerns the measurement of the angle {alpha} in the B{sup 0} {yields} {rho}{pi} mode.

  9. Rho GTPases in embryonic development

    PubMed Central

    Duquette, Philippe M; Lamarche-Vane, Nathalie

    2014-01-01

    In the last decade, several mouse models for RhoA, Rac1, and Cdc42 have emerged and have contributed a great deal to understanding the precise functions of Rho GTPases at early stages of development. This review summarizes our current knowledge of various mouse models of tissue-specific ablation of Cdc42, Rac1, and RhoA with emphasis on early embryogenesis, epithelial and skin morphogenesis, tubulogenesis, development of the central nervous system, and limb development. PMID:25483305

  10. Search for the Decay B^0 -> a^\\pm_1 \\rho^\\mp

    SciTech Connect

    Aubert, B.

    2006-05-10

    The authors present a search for the rare B-meson decay B{sup 0} {yields} {alpha}{sub 1}{sup {+-}}{rho}{sup {-+}} with {alpha}{sub 1}{sup {+-}} {yields} {pi}{sup +}{pi}{sup -}{pi}{sup {+-}}. We use (110 {+-} 1.2) x 10{sup 6} {Upsilon}(4S) {yields} B{bar B} decays collected with the BABAR detector at the PEp-II asymmetric-energy B Factory at SLAC. They obtain an upper limit of 30 x 10{sup -6} (90% C.L.) for the branching fraction product {Beta}(B{sup 0} {yields} {alpha}{sub 1}{sup {+-}}{rho}{sup {-+}}) {Beta}({alpha}{sub 1}{sup {+-}} {yields} {pi}{sup +}{pi}{sup -}{pi}{sup {+-}}), where they assume that the {alpha}{sub 1}{sup {+-}} decays exclusively to {rho}{sup 0}{pi}{sup {+-}}.

  11. Bacterial Cytotoxins Target Rho GTPases

    NASA Astrophysics Data System (ADS)

    Schmidt, Gudula; Aktories, Klaus

    1998-06-01

    Low molecular mass GTPases of the Rho family, which are involved in the regulation of the actin cytoskeleton and in various signal transduction processes, are the eukaryotic targets of bacterial protein toxins. The toxins covalently modify Rho proteins by ADP ribosylation, glucosylation, and deamidation, thereby inactivating and activating the GTPases.

  12. RhoB regulates the function of macrophages in the hypoxia-induced inflammatory response

    PubMed Central

    Huang, Gaoxiang; Su, Jie; Zhang, Mingzhuo; Jin, Yiduo; Wang, Yan; Zhou, Peng; Lu, Jian

    2017-01-01

    Immune cells, particularly macrophages, play critical roles in the hypoxia-induced inflammatory response. The small GTPase RhoB is usually rapidly induced by a variety of stimuli and has been described as an important regulator of cytoskeletal organization and vesicle and membrane receptor trafficking. However, it is unknown whether RhoB is involved in the hypoxia-induced inflammatory response. Here, we investigated the effect of hypoxia on the expression of RhoB and the mechanism and significance of RhoB expression in macrophages. We found that hypoxia significantly upregulated the expression of RhoB in RAW264.7 cells, mouse peritoneal macrophages, and the spleen of rats. Hypoxia-induced expression of RhoB was significantly blocked by a specific inhibitor of hypoxia-inducible factor-1α (HIF-1α), c-Jun N-terminal kinase (JNK), or extracellular-signal regulated protein kinase (ERK), indicating that hypoxia-activated HIF-1α, JNK, and ERK are involved in the upregulation of RhoB by hypoxia. Knockdown of RhoB expression not only significantly suppressed basal production of interleukin-1 beta (IL-1β), interleukin 6 (IL-6), and tumor necrosis factor alpha (TNF-α) in normoxia but also more markedly decreased the hypoxia-stimulated production of these cytokines. Furthermore, we showed that RhoB increased nuclear factor-kappa B (NF-κB) activity, and the inhibition of NF-κB transcriptional activity significantly decreased the RhoB-increased mRNA levels of IL-1β, IL-6, and TNF-α. Finally, we demonstrated that RhoB enhanced cell adhesion and inhibited cell migration in normoxia and hypoxia. Taken together, these results suggest that RhoB plays an important role in the hypoxia-induced activation of macrophages and the inflammatory response. PMID:26388235

  13. RhoB regulates the function of macrophages in the hypoxia-induced inflammatory response.

    PubMed

    Huang, Gaoxiang; Su, Jie; Zhang, Mingzhuo; Jin, Yiduo; Wang, Yan; Zhou, Peng; Lu, Jian

    2017-03-01

    Immune cells, particularly macrophages, play critical roles in the hypoxia-induced inflammatory response. The small GTPase RhoB is usually rapidly induced by a variety of stimuli and has been described as an important regulator of cytoskeletal organization and vesicle and membrane receptor trafficking. However, it is unknown whether RhoB is involved in the hypoxia-induced inflammatory response. Here, we investigated the effect of hypoxia on the expression of RhoB and the mechanism and significance of RhoB expression in macrophages. We found that hypoxia significantly upregulated the expression of RhoB in RAW264.7 cells, mouse peritoneal macrophages, and the spleen of rats. Hypoxia-induced expression of RhoB was significantly blocked by a specific inhibitor of hypoxia-inducible factor-1α (HIF-1α), c-Jun N-terminal kinase (JNK), or extracellular-signal regulated protein kinase (ERK), indicating that hypoxia-activated HIF-1α, JNK, and ERK are involved in the upregulation of RhoB by hypoxia. Knockdown of RhoB expression not only significantly suppressed basal production of interleukin-1 beta (IL-1β), interleukin 6 (IL-6), and tumor necrosis factor alpha (TNF-α) in normoxia but also more markedly decreased the hypoxia-stimulated production of these cytokines. Furthermore, we showed that RhoB increased nuclear factor-kappa B (NF-κB) activity, and the inhibition of NF-κB transcriptional activity significantly decreased the RhoB-increased mRNA levels of IL-1β, IL-6, and TNF-α. Finally, we demonstrated that RhoB enhanced cell adhesion and inhibited cell migration in normoxia and hypoxia. Taken together, these results suggest that RhoB plays an important role in the hypoxia-induced activation of macrophages and the inflammatory response.Cellular & Molecular Immunology advance online publication, 21 September 2015; doi:10.1038/cmi.2015.78.

  14. Rho GTPases, phosphoinositides, and actin

    PubMed Central

    Croisé, Pauline; Estay-Ahumada, Catherine; Gasman, Stéphane; Ory, Stéphane

    2014-01-01

    Rho GTPases are well known regulators of the actin cytoskeleton that act by binding and activating actin nucleators. They are therefore involved in many actin-based processes, including cell migration, cell polarity, and membrane trafficking. With the identification of phosphoinositide kinases and phosphatases as potential binding partners or effectors, Rho GTPases also appear to participate in the regulation of phosphoinositide metabolism. Since both actin dynamics and phosphoinositide turnover affect the efficiency and the fidelity of vesicle transport between cell compartments, Rho GTPases have emerged as critical players in membrane trafficking. Rho GTPase activity, actin remodeling, and phosphoinositide metabolism need to be coordinated in both space and time to ensure the progression of vesicles along membrane trafficking pathways. Although most molecular pathways are still unclear, in this review, we will highlight recent advances made in our understanding of how Rho-dependent signaling pathways organize actin dynamics and phosphoinositides and how phosphoinositides potentially provide negative feedback to Rho GTPases during endocytosis, exocytosis and membrane exchange between intracellular compartments. PMID:24914539

  15. Rho A and the Rho kinase pathway regulate fibroblast contraction: Enhanced contraction in constitutively active Rho A fibroblast cells

    SciTech Connect

    Nobe, Koji; Nobe, Hiromi; Yoshida, Hiroko; Kolodney, Michael S.; Paul, Richard J.; Honda, Kazuo

    2010-08-20

    Research highlights: {yields} Mechanisms of fibroblast cell contraction in collagen matrix. {yields} Assessed an isometric force development using 3D-reconstituted-fibroblast fiber. {yields} Constitutively active Rho A induced the over-contraction of fibroblast cells. {yields} Rho A and Rho kinase pathway has a central role in fibroblast cell contraction. -- Abstract: Fibroblast cells play a central role in the proliferation phase of wound healing processes, contributing to force development. The intracellular signaling pathways regulating this non-muscle contraction are only partially understood. To study the relations between Rho A and contractile responses, constitutively active Rho A (CA-Rho A) fibroblast cells were reconstituted into fibers and the effects of calf serum (CS) on isometric force were studied. CS-induced force in CA-Rho A fibroblast fibers was twice as large as that in wild type (NIH 3T3) fibroblast fibers. During this response, the translocation of Rho A from the cytosol to the membrane was detected by Rho A activity assays and Western blot analysis. Pre-treatment with a Rho specific inhibitor (C3-exoenzyme) suppressed translocation as well as contraction. These results indicate that Rho A activation is essential for fibroblast contraction. The Rho kinase inhibitor ( (Y27632)) inhibited both NIH 3T3 and CA-Rho A fibroblast fiber contractions. Activation of Rho A is thus directly coupled with Rho kinase activity. We conclude that the translocation of Rho A from the cytosol to the membrane and the Rho kinase pathway can regulate wound healing processes mediated by fibroblast contraction.

  16. Rho GTPases, oxidation, and cell redox control

    PubMed Central

    Hobbs, G Aaron; Zhou, Bingying; Cox, Adrienne D; Campbell, Sharon L

    2014-01-01

    While numerous studies support regulation of Ras GTPases by reactive oxygen and nitrogen species, the Rho subfamily has received considerably less attention. Over the last few years, increasing evidence is emerging that supports the redox sensitivity of Rho GTPases. Moreover, as Rho GTPases regulate the cellular redox state by controlling enzymes that generate and convert reactive oxygen and nitrogen species, redox feedback loops likely exist. Here, we provide an overview of cellular oxidants, Rho GTPases, and their inter-dependence. PMID:24809833

  17. Arachidonic acid-induced Ca2+ sensitization of smooth muscle contraction through activation of Rho-kinase.

    PubMed

    Araki, S; Ito, M; Kureishi, Y; Feng, J; Machida, H; Isaka, N; Amano, M; Kaibuchi, K; Hartshorne, D J; Nakano, T

    2001-02-01

    Arachidonic acid activates isolated Rho-kinase and contracts permeabilized smooth muscle fibres. Various assays were carried out to examine the mechanism of this activation. Native Rho-kinase was activated 5-6 times by arachidonic acid but an N-terminal, constitutively-active fragment of Rho-kinase, expressed as a glutathione-S-transferase (GST) fusion protein and including the catalytic subunit (GST-Rho-kinase-CAT), was not. GST-Rho-kinase-CAT was inhibited by a C-terminal fragment of Rho-kinase and arachidonic acid removed this inhibition. These results suggest that the C-terminal part of Rho-kinase, containing the RhoA binding site and the pleckstrin homology domain, acts as an autoinhibitor. It is suggested further that activation by arachidonic acid is due to its binding to the autoinhibitory region and subsequent release from the catalytic site. Arachidonic acid, at concentrations greater than 30 microM, increases force in alpha-toxin-permeabilized femoral artery but not in Triton X-100-skinned fibres. The content of Rho-kinase in the latter was lower than in alpha-toxin-treated or intact fibres. The arachidonic acid-induced contraction was not observed at a pCa above 8.0 and was inhibited by Y-27632 and wortmannin, inhibitors of Rho-kinase and myosin light-chain kinase (MLCK), respectively. The activation of Rho-kinase and subsequent phosphorylation of the myosin phosphatase target subunit inhibits myosin phosphatase and increases myosin phosphorylation.

  18. RhoG regulates anoikis through a phosphatidylinositol 3-kinase-dependent mechanism

    SciTech Connect

    Yamaki, Nao; Negishi, Manabu; Katoh, Hironori . E-mail: hirokato@pharm.kyoto-u.ac.jp

    2007-08-01

    In normal epithelial cells, cell-matrix interaction is required for cell survival and proliferation, whereas disruption of this interaction causes epithelial cells to undergo apoptosis called anoikis. Here we show that the small GTPase RhoG plays an important role in the regulation of anoikis. HeLa cells are capable of anchorage-independent cell growth and acquire resistance to anoikis. We found that RNA interference-mediated knockdown of RhoG promoted anoikis in HeLa cells. Previous studies have shown that RhoG activates Rac1 and induces several cellular functions including promotion of cell migration through its effector ELMO and the ELMO-binding protein Dock180 that function as a Rac-specific guanine nucleotide exchange factor. However, RhoG-induced suppression of anoikis was independent of the ELMO- and Dock180-mediated activation of Rac1. On the other hand, the regulation of anoikis by RhoG required phosphatidylinositol 3-kinase (PI3K) activity, and constitutively active RhoG bound to the PI3K regulatory subunit p85{alpha} and induced the PI3K-dependent phosphorylation of Akt. Taken together, these results suggest that RhoG protects cells from apoptosis caused by the loss of anchorage through a PI3K-dependent mechanism, independent of its activation of Rac1.

  19. Quantitative Analysis of Prenylated RhoA Interaction with Its Chaperone, RhoGDI*

    PubMed Central

    Tnimov, Zakir; Guo, Zhong; Gambin, Yann; Nguyen, Uyen T. T.; Wu, Yao-Wen; Abankwa, Daniel; Stigter, Anouk; Collins, Brett M.; Waldmann, Herbert; Goody, Roger S.; Alexandrov, Kirill

    2012-01-01

    Small GTPases of the Rho family regulate cytoskeleton remodeling, cell polarity, and transcription, as well as the cell cycle, in eukaryotic cells. Membrane delivery and recycling of the Rho GTPases is mediated by Rho GDP dissociation inhibitor (RhoGDI), which forms a stable complex with prenylated Rho GTPases. We analyzed the interaction of RhoGDI with the active and inactive forms of prenylated and unprenylated RhoA. We demonstrate that RhoGDI binds the prenylated form of RhoA·GDP with unexpectedly high affinity (Kd = 5 pm). The very long half-life of the complex is reduced 25-fold on RhoA activation, with a concomitant reduction in affinity (Kd = 3 nm). The 2.8-Å structure of the RhoA·guanosine 5′-[β,γ-imido] triphosphate (GMPPNP)·RhoGDI complex demonstrated that complex formation forces the activated RhoA into a GDP-bound conformation in the absence of nucleotide hydrolysis. We demonstrate that membrane extraction of Rho GTPase by RhoGDI is a thermodynamically favored passive process that operates through a series of progressively tighter intermediates, much like the one that is mediated by RabGDI. PMID:22628549

  20. Deregulation of Rho GTPases in cancer

    PubMed Central

    Porter, Andrew P.; Papaioannou, Alexandra; Malliri, Angeliki

    2016-01-01

    ABSTRACT In vitro and in vivo studies and evidence from human tumors have long implicated Rho GTPase signaling in the formation and dissemination of a range of cancers. Recently next generation sequencing has identified direct mutations of Rho GTPases in human cancers. Moreover, the effects of ablating genes encoding Rho GTPases and their regulators in mouse models, or through pharmacological inhibition, strongly suggests that targeting Rho GTPase signaling could constitute an effective treatment. In this review we will explore the various ways in which Rho signaling can be deregulated in human cancers. PMID:27104658

  1. Plasma membrane restricted RhoGEF activity is sufficient for RhoA-mediated actin polymerization

    PubMed Central

    van Unen, Jakobus; Reinhard, Nathalie R.; Yin, Taofei; Wu, Yi I.; Postma, Marten; Gadella, Theodorus W.J.; Goedhart, Joachim

    2015-01-01

    The small GTPase RhoA is involved in cell morphology and migration. RhoA activity is tightly regulated in time and space and depends on guanine exchange factors (GEFs). However, the kinetics and subcellular localization of GEF activity towards RhoA are poorly defined. To study the mechanism underlying the spatiotemporal control of RhoA activity by GEFs, we performed single cell imaging with an improved FRET sensor reporting on the nucleotide loading state of RhoA. By employing the FRET sensor we show that a plasma membrane located RhoGEF, p63RhoGEF, can rapidly activate RhoA through endogenous GPCRs and that localized RhoA activity at the cell periphery correlates with actin polymerization. Moreover, synthetic recruitment of the catalytic domain derived from p63RhoGEF to the plasma membrane, but not to the Golgi apparatus, is sufficient to activate RhoA. The synthetic system enables local activation of endogenous RhoA and effectively induces actin polymerization and changes in cellular morphology. Together, our data demonstrate that GEF activity at the plasma membrane is sufficient for actin polymerization via local RhoA signaling. PMID:26435194

  2. Epigallocatechin-3-gallate, a green-tea polyphenol, suppresses Rho signaling in TWNT-4 human hepatic stellate cells.

    PubMed

    Higashi, Nobuhiko; Kohjima, Motoyuki; Fukushima, Marie; Ohta, Satoshi; Kotoh, Kazuhiro; Enjoji, Munechika; Kobayashi, Naoya; Nakamuta, Makoto

    2005-06-01

    Epigallocatechin-3-gallate (EGCG), a major constituent of the polyphenoids in green tea, has been reported to possess a wide range of biologic activities, including antifibrogenesis. Activated hepatic stellate cells (HSCs) are central to hepatic fibrosis, and Rho (a small GTPase)-signaling pathways have been implicated in the activation and proliferation of HSCs. In this study, we investigated the effect of EGCG on Rho-signaling pathways in activated human HSC-derived TWNT-4 cells. EGCG inhibited stress-fiber formation, an indicator of Rho activation, and changed the distribution of alpha-smooth-muscle actin. These inhibitory effects of EGCG were restored by overexpression of constitutively active Rho. A pull-down assay revealed that activated Rho (GTP-bound state) was strongly inhibited by ECGC and accompanied by suppressed phosphorylation of focal adhesion kinase, which is a regulator of Rho-signaling pathways. 5-Bromo-2'-deoxy-uridine incorporation demonstrated that ECGC (100 micromol/L suppressed cell growth by 80%, and terminal deoxynucleotidyl transferase viotin-deoxyruidine triphosphate nick end-labeling revealed that EGCG (100 micromol/L) caused apoptosis in half of the total cells. EGCG also strongly inhibited lysophoaphatidic acid (an activator of Rho) and induced phosphorylation of mitogen-activated protein kinases (Erk1/2, c-jun kinase, and p38). These findings demonstrate that EGCG regulates the structure and growth of HSCs by way of Rho-signaling pathways and suggest that EGCG has therapeutic potential in the setting of liver fibrosis.

  3. Pathophysiological effects of RhoA and Rho-associated kinase on cardiovascular system.

    PubMed

    Cai, Anping; Li, Liwen; Zhou, Yingling

    2016-01-01

    In past decades, growing evidence from basic and clinical researches reveal that small guanosine triphosphate binding protein ras homolog gene family, member A (RhoA) and its main effector Rho-associated kinase (ROCK) play central and complex roles in cardiovascular systems, and increasing RhoA and ROCK activity is associated with a broad range of cardiovascular diseases such as congestive heart failure, atherosclerosis, and hypertension. Favorable outcomes have been observed with ROCK inhibitors treatment. In this review, we briefly summarize the pathophysiological roles of RhoA/ROCK signaling pathway on cardiovascular system, displaying the potential benefits in the cardiovascular system with controlling RhoA/ROCK signaling pathway.

  4. Rho-directed forces in collective migration.

    PubMed

    Friedl, Peter; Wolf, Katarina; Zegers, Mirjam M

    2014-03-01

    Collective cell migration depends on multicellular mechanocoupling between leader and follower cells to coordinate traction force and position change. Co-registration of Rho GTPase activity and forces in migrating epithelial cell sheets now shows how RhoA controls leader-follower cell hierarchy, multicellular cytoskeletal contractility and mechanocoupling, to prevent ectopic leading edges and to move the cell sheet forward.

  5. Rho GTPases and cancer cell transendothelial migration.

    PubMed

    Reymond, Nicolas; Riou, Philippe; Ridley, Anne J

    2012-01-01

    Small Rho GTPases are major regulators of actin cytoskeleton dynamics and influence cell shape and migration. The expression of several Rho GTPases is often up-regulated in tumors and this frequently correlates with a poor prognosis for patients. Migration of cancer cells through endothelial cells that line the blood vessels, called transendothelial migration or extravasation, is a critical step during the metastasis process. The use of siRNA technology to target specifically each Rho family member coupled with imaging techniques allows the roles of individual Rho GTPases to be investigated. In this chapter we describe methods to assess how Rho GTPases affect the different steps of cancer cell transendothelial cell migration in vitro.

  6. RhoGDIα-dependent balance between RhoA and RhoC is a key regulator of cancer cell tumorigenesis

    PubMed Central

    Giang Ho, T. T.; Stultiens, Audrey; Dubail, Johanne; Lapière, Charles M.; Nusgens, Betty V.; Colige, Alain C.; Deroanne, Christophe F.

    2011-01-01

    RhoGTPases are key signaling molecules regulating main cellular functions such as migration, proliferation, survival, and gene expression through interactions with various effectors. Within the RhoA-related subclass, RhoA and RhoC contribute to several steps of tumor growth, and the regulation of their expression affects cancer progression. Our aim is to investigate their respective contributions to the acquisition of an invasive phenotype by using models of reduced or forced expression. The silencing of RhoC, but not of RhoA, increased the expression of genes encoding tumor suppressors, such as nonsteroidal anti-inflammatory drug–activated gene 1 (NAG-1), and decreased migration and the anchorage-independent growth in vitro. In vivo, RhoC small interfering RNA (siRhoC) impaired tumor growth. Of interest, the simultaneous knockdown of RhoC and NAG-1 repressed most of the siRhoC-related effects, demonstrating the central role of NAG-1. In addition of being induced by RhoC silencing, NAG-1 was also largely up-regulated in cells overexpressing RhoA. The silencing of RhoGDP dissociation inhibitor α (RhoGDIα) and the overexpression of a RhoA mutant unable to bind RhoGDIα suggested that the effect of RhoC silencing is indirect and results from the up-regulation of the RhoA level through competition for RhoGDIα. This study demonstrates the dynamic balance inside the RhoGTPase network and illustrates its biological relevance in cancer progression. PMID:21757538

  7. Measurement of the B+- to rho pi0 Branching Fraction and Direct CP Asymmetry

    SciTech Connect

    Aubert, B.

    2007-01-25

    The authors present improved measurements of the branching fraction and CP asymmetry for the process B{sup {+-}} {yields} {rho}{sup {+-}}{pi}{sup 0}. The data sample corresponding to 211 fb{sup -1} comprises 232 million {Upsilon}(4S) {yields} B{bar B} decays collected with the BABAR detector at the PEP-II asymmetric B Factory at SLAC. The yield and CP asymmetry are measured using an extended maximum likelihood fitting method. The branching fraction and Cp asymmetry are found to be {Beta}(B{sup {+-}} {yields} {rho}{sup {+-}}{pi}{sup 0}) = [10.2 {+-} 1.4(stat) {+-} 0.9(syst)] x 10{sup -6} and {Alpha}{sub CP}(B{sup {+-}} {yields} {rho}{sup {+-}}{pi}{sup 0}) = -0.01 {+-} 0.13(stat) {+-} 0.02(syst).

  8. Unprecedentedly high selective adsorption of gas mixtures in rho zeolite-like metal-organic framework: a molecular simulation study.

    PubMed

    Babarao, Ravichandar; Jiang, Jianwen

    2009-08-19

    We report a molecular simulation study for the separation of industrially important gas mixtures (CO(2)/H(2), CO(2)/CH(4), and CO(2)/N(2)) in rho zeolite-like metal-organic framework (rho-ZMOF). Rho-ZMOF contains a wide-open anionic framework and charge-balancing extraframework Na(+) ions. Two types of binding sites for Na(+) ions are identified in the framework. Site I is in the single eight-membered ring, whereas site II is in the alpha-cage. Na(+) ions at site I have a stronger affinity for the framework and thus a smaller mobility. The binding sites in rho-ZMOF resemble those in its inorganic counterpart rho-zeolite. CO(2) is adsorbed predominantly over other gases because of its strong electrostatic interactions with the charged framework and the presence of Na(+) ions acting as additional adsorption sites. At ambient temperature and pressure, the CO(2) selectivities are 1800 for the CO(2)/H(2) mixture, 80 for the CO(2)/CH(4) mixture, and 500 for the CO(2)/N(2) mixture. Compared with other MOFs and nanoporous materials reported to date, rho-ZMOF exhibits unprecedentedly high selective adsorption for these gas mixtures. This work represents the first simulation study to characterize extraframework ions and examine gas separation in a charged ZMOF. The simulation results reveal that rho-ZMOF is a promising candidate for the separation of syngas, natural gas, and flue gas.

  9. Rho-associated kinase, a novel serine/threonine kinase, as a putative target for small GTP binding protein Rho.

    PubMed Central

    Matsui, T; Amano, M; Yamamoto, T; Chihara, K; Nakafuku, M; Ito, M; Nakano, T; Okawa, K; Iwamatsu, A; Kaibuchi, K

    1996-01-01

    The small GTP binding protein Rho is implicated in cytoskeletal responses to extracellular signals such as lysophosphatidic acid to form stress fibers and focal contacts. Here we have purified a Rho-interacting protein with a molecular mass of approximately 164 kDa (p164) from bovine brain. This protein bound to GTPgammaS (a non-hydrolyzable GTP analog).RhoA but not to GDP.RhoA or GTPgammaS.RhoA with a mutation in the effector domain (RhoAA37).p164 had a kinase activity which was specifically stimulated by GTPgammaS.RhoA. We obtained the cDNA encoding p164 on the basis of its partial amino acid sequences and named it Rho-associated kinase (Rho-kinase). Rho-kinase has a catalytic domain in the N-terminal portion, a coiled coil domain in the middle portion and a zinc finger-like motif in the C-terminal portion. The catalytic domain shares 72% sequence homology with that of myotonic dystrophy kinase and the coiled coil domain contains a Rho-interacting interface. When COS7 cells were cotransfected with Rho-kinase and activated RhoA, some Rho-kinase was recruited to membranes. Thus it is likely that Rho-kinase is a putative target serine/threonine kinase for Rho and serves as a mediator of the Rho-dependent signaling pathway. Images PMID:8641286

  10. Expression of GABA receptor rho subunits in rat brain.

    PubMed

    Boue-Grabot, E; Roudbaraki, M; Bascles, L; Tramu, G; Bloch, B; Garret, M

    1998-03-01

    The GABA receptor rho1, rho2, and rho3 subunits are expressed in the retina where they form bicuculline-insensitive GABA(C) receptors. We used northern blot, in situ hybridization, and RT-PCR analysis to study the expression of rho subunits in rat brains. In situ hybridization allowed us to detect rho-subunit expression in the superficial gray layer of the superior colliculus and in the cerebellar Purkinje cells. RT-PCR experiments indicated that (a) in retina and in domains that may contain functional GABA(C) receptors, rho2 and rho1 subunits are expressed at similar levels; and (b) in domains and in tissues that are unlikely to contain GABA(C) receptors, rho2 mRNA is enriched relative to rho1 mRNA. These results suggest that both rho1 and rho2 subunits are necessary to form a functional GABA(C) receptor. The use of RT-PCR also showed that, except in the superior colliculus, rho3 is expressed along with rho1 and rho2 subunits. We also raised an antibody against a peptide sequence unique to the rho1 subunit. The use of this antibody on cerebellum revealed the rat rho1 subunit in the soma and dendrites of Purkinje neurons. The allocation of GABA(C) receptor subunits to identified neurons paves the way for future electrophysiological studies.

  11. GTPase regulation: getting aRnd Rock and Rho inhibition.

    PubMed

    Chardin, Pierre

    2003-09-16

    Rnd proteins are atypical members of the Rho small G protein family that inhibit the formation of actomyosin contractile fibers via activation of RhoGAPs and inhibition of a Rho effector, the Ser/Thr kinase Rock. These mechanisms might be used to fine-tune Rho GTPase inhibition locally at sites where particular actin structures need to be made.

  12. Photoproduction of the rho meson and its magnetic moments

    SciTech Connect

    Kaneko, Hiromi; Hosaka, Atsushi; Scholten, Olaf

    2011-10-21

    We study photoproduction of {rho} meson in a model of hidden local symmetry. We introduce the {rho} meson on a hidden gauge boson and phenomenological {rho} meson-nucleon Lagrangian is constructed respecting chiral symmetry. It turns out that the {sigma}-exchange interaction plays an important role in neutral {rho} meson photoproduction to reproduce the experimental cross sections. In charged {rho} meson photoproduction, the model takes into account the {rho} meson magnetic moments from the three-point vertex in the kinetic terms. We show that the magnetic moment of the charged {rho} meson has a significant effect on the total cross sections in proportion to the photon energies.

  13. Regulation of Rho proteins by phosphorylation in the cardiovascular system.

    PubMed

    Loirand, Gervaise; Guilluy, Christophe; Pacaud, Pierre

    2006-08-01

    The small G protein Rho signaling pathways are recognized as major regulators of cardiovascular functions, and activation of Rho proteins appears to be a common component for the pathogenesis of hypertension and vascular proliferative disorders. Rho proteins are tightly regulated, and recent evidence suggests that modulation of Rho protein signaling by phosphorylation of Rho proteins provides an additional simple mechanism for coordinating Rho protein functions. This regulation by phosphorylation is particularly important in the arterial wall, where RhoA protein expressed in vascular smooth muscle cells is controlled by the endothelium through the nitric oxide/cGMP-dependent kinase pathway.

  14. RhoBTB3: A Rho GTPase-family ATPase required for endosome to Golgi transport

    PubMed Central

    Espinosa, Eric J.; Calero, Monica; Sridevi, Khambhampaty; Pfeffer, Suzanne R.

    2009-01-01

    Summary Rho GTPases are key regulators of the actin-based cytoskeleton; Rab GTPases are key regulators of membrane traffic. We report here that the atypical Rho GTPase family member, RhoBTB3, binds directly to Rab9 GTPase, and functions with Rab9 in protein transport from endosomes to the trans Golgi network. Gene replacement experiments show that RhoBTB3 function in cultured cells requires both RhoBTB3’s N-terminal, Rho-related domain, and C-terminal sequences that are important for Rab9 interaction.9 Biochemical analysis reveals that RhoBTB3 binds and hydrolyzes ATP rather than GTP. Rab9 binding opens the auto-inhibited RhoBTB3 protein to permit maximal ATP hydroysis. Because RhoBTB3 interacts with TIP47 on membranes, we propose that it may function to release this cargo selection protein from vesicles to permit their efficient docking and fusion at the Golgi. PMID:19490898

  15. Analysis of RhoA and Rho GEF activity in whole cells and the cell nucleus.

    PubMed

    Guilluy, Christophe; Dubash, Adi D; García-Mata, Rafael

    2011-12-01

    We have recently shown that a fraction of the total cellular pool of the small GTPase RhoA resides in the nucleus, and that the nuclear guanine nucleotide exchange factor (GEF) Net1 has a role in the regulation of its activity. In this protocol, we describe a method to measure both the activities of the nuclear pools of RhoA and Rho GEFs. This process required the development of a nuclear isolation protocol that is both fast and virtually free of cytosolic and membrane contaminants, as well as a redesign of existing RhoA and Rho GEF activity assays so that they work in nuclear samples. This protocol can be also used for other Rho GTPases and Rho GEFs, which have also been found in the nucleus. Completion of the procedure, including nuclear isolation and RhoA or Rho GEF activity assay, takes 1 h 40 min. We also include details of how to perform a basic assay of whole-cell extracts.

  16. Coherent rho(0) production in ultraperipheral heavy-ion collisions.

    PubMed

    Adler, C; Ahammed, Z; Allgower, C; Amonett, J; Anderson, B D; Anderson, M; Averichev, G S; Balewski, J; Barannikova, O; Barnby, L S; Baudot, J; Bekele, S; Belaga, V V; Bellwied, R; Berger, J; Bichsel, H; Bland, L C; Blyth, C O; Bonner, B E; Boucham, A; Brandin, A; Bravar, A; Cadman, R V; Caines, H; Calderón de la Barca Sánchez, M; Cardenas, A; Carroll, J; Castillo, J; Castro, M; Cebra, D; Chaloupka, P; Chattopadhyay, S; Chen, Y; Chernenko, S P; Cherney, M; Chikanian, A; Choi, B; Christie, W; Coffin, J P; Cormier, T M; Cramer, J G; Crawford, H J; Deng, W S; Derevschikov, A A; Didenko, L; Dietel, T; Draper, J E; Dunin, V B; Dunlop, J C; Eckardt, V; Efimov, L G; Emelianov, V; Engelage, J; Eppley, G; Erazmus, B; Fachini, P; Faine, V; Filimonov, K; Finch, E; Fisyak, Y; Flierl, D; Foley, K J; Fu, J; Gagliardi, C A; Gagunashvili, N; Gans, J; Gaudichet, L; Germain, M; Geurts, F; Ghazikhanian, V; Grachov, O; Grigoriev, V; Guedon, M; Gushin, E; Hallman, T J; Hardtke, D; Harris, J W; Henry, T W; Heppelmann, S; Herston, T; Hippolyte, B; Hirsch, A; Hjort, E; Hoffmann, G W; Horsley, M; Huang, H Z; Humanic, T J; Igo, G; Ishihara, A; Ivanshin, Yu I; Jacobs, P; Jacobs, W W; Janik, M; Johnson, I; Jones, P G; Judd, E G; Kaneta, M; Kaplan, M; Keane, D; Kiryluk, J; Kisiel, A; Klay, J; Klein, S R; Klyachko, A; Konstantinov, A S; Kopytine, M; Kotchenda, L; Kovalenko, A D; Kramer, M; Kravtsov, P; Krueger, K; Kuhn, C; Kulikov, A I; Kunde, G J; Kunz, C L; Kutuev, R Kh; Kuznetsov, A A; Lakehal-Ayat, L; Lamont, M A C; Landgraf, J M; Lange, S; Lansdell, C P; Lasiuk, B; Laue, F; Lebedev, A; Lednický, R; Leontiev, V M; LeVine, M J; Li, Q; Lindenbaum, S J; Lisa, M A; Liu, F; Liu, L; Liu, Z; Liu, Q J; Ljubicic, T; Llope, W J; LoCurto, G; Long, H; Longacre, R S; Lopez-Noriega, M; Love, W A; Ludlam, T; Lynn, D; Ma, J; Majka, R; Margetis, S; Markert, C; Martin, L; Marx, J; Matis, H S; Matulenko, Yu A; McShane, T S; Meissner, F; Melnick, Yu; Meschanin, A; Messer, M; Miller, M L; Milosevich, Z; Minaev, N G; Mitchell, J; Moiseenko, V A; Moore, C F; Morozov, V; de Moura, M M; Munhoz, M G; Nelson, J M; Nevski, P; Nikitin, V A; Nogach, L V; Norman, B; Nurushev, S B; Nystrand, J; Odyniec, G; Ogawa, A; Okorokov, V; Oldenburg, M; Olson, D; Paic, G; Pandey, S U; Panebratsev, Y; Panitkin, S Y; Pavlinov, A I; Pawlak, T; Perevoztchikov, V; Peryt, W; Petrov, V A; Planinic, M; Pluta, J; Porile, N; Porter, J; Poskanzer, A M; Potrebenikova, E; Prindle, D; Pruneau, C; Putschke, J; Rai, G; Rakness, G; Ravel, O; Ray, R L; Razin, S V; Reichhold, D; Reid, J G; Retiere, F; Ridiger, A; Ritter, H G; Roberts, J B; Rogachevski, O V; Romero, J L; Roy, C; Rykov, V; Sakrejda, I; Salur, S; Sandweiss, J; Saulys, A C; Savin, I; Schambach, J; Scharenberg, R P; Schmitz, N; Schroeder, L S; Schüttauf, A; Schweda, K; Seger, J; Seliverstov, D; Seyboth, P; Shahaliev, E; Shestermanov, K E; Shimanskii, S S; Shvetcov, V S; Skoro, G; Smirnov, N; Snellings, R; Sorensen, P; Sowinski, J; Spinka, H M; Srivastava, B; Stephenson, E J; Stock, R; Stolpovsky, A; Strikhanov, M; Stringfellow, B; Struck, C; Suaide, A A P; Sugarbaker, E; Suire, C; Sumbera, M; Surrow, B; Symons, T J M; Szanto de Toledo, A; Szarwas, P; Tai, A; Takahashi, J; Tang, A H; Thomas, J H; Thompson, M; Tikhomirov, V; Tokarev, M; Tonjes, M B; Trainor, T A; Trentalange, S; Tribble, R E; Trofimov, V; Tsai, O; Ullrich, T; Underwood, D G; Van Buren, G; VanderMolen, A M; Vasilevski, I M; Vasiliev, A N; Vigdor, S E; Voloshin, S A; Wang, F; Ward, H; Watson, J W; Wells, R; Westfall, G D; Whitten, C; Wieman, H; Willson, R; Wissink, S W; Witt, R; Wood, J; Xu, N; Xu, Z; Yakutin, A E; Yamamoto, E; Yang, J; Yepes, P; Yurevich, V I; Zanevski, Y V; Zborovský, I; Zhang, H; Zhang, W M; Zoulkarneev, R; Zubarev, A N

    2002-12-30

    The STAR Collaboration reports the first observation of exclusive rho(0) photoproduction, AuAu-->AuAurho(0), and rho(0) production accompanied by mutual nuclear Coulomb excitation, AuAu-->Au*Au*rho(0), in ultraperipheral heavy-ion collisions. The rho(0) have low transverse momenta, consistent with coherent coupling to both nuclei. The cross sections at sqrt[s(NN)]=130 GeV agree with theoretical predictions treating rho(0) production and Coulomb excitation as independent processes.

  17. Phylogenetic analysis of sequences from diverse bacteria with homology to the Escherichia coli rho gene.

    PubMed Central

    Opperman, T; Richardson, J P

    1994-01-01

    Genes from Pseudomonas fluorescens, Chromatium vinosum, Micrococcus luteus, Deinococcus radiodurans, and Thermotoga maritima with homology to the Escherichia coli rho gene were cloned and sequenced, and their sequences were compared with other available sequences. The species for all of the compared sequences are members of five bacterial phyla, including Thermotogales, the most deeply diverged phylum. This suggests that a rho-like gene is ubiquitous in the Bacteria and was present in their common ancestor. The comparative analysis revealed that the Rho homologs are highly conserved, exhibiting a minimum identity of 50% of their amino acid residues in pairwise comparisons. The ATP-binding domain had a particularly high degree of conservation, consisting of some blocks with sequences of residues that are very similar to segments of the alpha and beta subunits of F1-ATPase and of other blocks with sequences that are unique to Rho. The RNA-binding domain is more diverged than the ATP-binding domain. However, one of its most highly conserved segments includes a RNP1-like sequence, which is known to be involved in RNA binding. Overall, the degree of similarity is lowest in the first 50 residues (the first half of the RNA-binding domain), in the putative connector region between the RNA-binding and the ATP-binding domains, and in the last 50 residues of the polypeptide. Since functionally defective mutants for E. coli Rho exist in all three of these segments, they represent important parts of Rho that have undergone adaptive evolution. PMID:8051015

  18. Rho GTPases, Statins, and Nitric Oxide

    PubMed Central

    Rikitake, Yoshiyuki; Liao, James K.

    2009-01-01

    The lipid-lowering drugs, 3-hydroxy-3-methylgulutaryl-coenzyme A (HMG-CoA) reductase inhibitors or statins, are used in the prevention and treatment of cardiovascular diseases. Recent experimental and clinical studies suggest that statins may exert vascular protective effects beyond cholesterol reduction. For example, statins improve endothelial function by cholesterol-dependent and -independent mechanisms. The cholesterol-independent or “pleiotropic” effects of statins include the upregulation and activation of endothelial NO synthase (eNOS). Because statins inhibit an early step in the cholesterol biosynthetic pathway, they also inhibit the synthesis of isoprenoids such as farnesylpyrophosphate and geranylgeranylpyrophosphate, which are important posttranslational lipid attachments for intracellular signaling molecules such as the Rho GTPases. Indeed, decrease in Rho GTPase responses as a consequence of statin treatment increases the production and bioavailability of endothelium-derived NO. The mechanism involves, in part, Rho/Rho-kinase (ROCK)-mediated changes in the actin cytoskeleton, which leads to decreases in eNOS mRNA stability. The regulation of eNOS by Rho GTPases, therefore, may be an important mechanism underlying the cardiovascular protective effect of statins. PMID:16339495

  19. RhoGDIα Acetylation at K127 and K141 Affects Binding toward Nonprenylated RhoA.

    PubMed

    Kuhlmann, Nora; Wroblowski, Sarah; Scislowski, Lukas; Lammers, Michael

    2016-01-19

    Rho proteins are major regulators of the cytoskeleton. As most Ras-related proteins, they switch between an active, GTP-bound and an inactive, GDP-bound conformation. Rho proteins are targeted to the plasma membrane via a polybasic region and a prenyl group attached to a C-terminal cysteine residue. To distribute Rho proteins in the cell, the molecular chaperone RhoGDIα binds to the prenylated Rho proteins forming a cytosolic pool of mainly GDP-loaded Rho. Most studies characterized the interaction of prenylated Rho proteins and RhoGDIα. However, RhoGDIα was also shown to bind to nonprenylated Rho proteins with physiologically relevant micomolar affinities. Recently, it was discovered that RhoGDIα is targeted by post-translational lysine acetylation. For one site, K141, it was hypothesized that acetylation might lead to increased levels of formation of filamentous actin and filopodia in mammalian cells. The functional consequences of lysine acetylation for the interplay with nonprenylated RhoA have not been investigated. Here, we report that lysine acetylation at lysines K127 and K141 in the RhoGDIα immunoglobulin domain interferes with the interaction toward nonprenylated RhoA using a combined biochemical and biophysical approach. We determined the first crystal structure of a doubly acetylated protein, RhoGDIα, in complex with RhoA·GDP. We discover that the C-terminus of RhoA adopts a different conformation forming an intermolecular β-sheet with the RhoGDIα immunoglobulin domain.

  20. Rho resonance parameters from lattice QCD

    SciTech Connect

    Guo, Dehua; Alexandru, Andrei; Molina, Raquel; Döring, Michael

    2016-08-01

    We perform a high-precision calculation of the phase shifts for $\\pi$-$\\pi$ scattering in the I = 1, J = 1 channel in the elastic region using elongated lattices with two mass-degenerate quark favors ($N_f = 2$). We extract the $\\rho$ resonance parameters using a Breit-Wigner fit at two different quark masses, corresponding to $m_{\\pi} = 226$MeV and $m_{\\pi} = 315$MeV, and perform an extrapolation to the physical point. The extrapolation is based on a unitarized chiral perturbation theory model that describes well the phase-shifts around the resonance for both quark masses. We find that the extrapolated value, $m_{\\rho} = 720(1)(15)$MeV, is significantly lower that the physical rho mass and we argue that this shift could be due to the absence of the strange quark in our calculation.

  1. Phospholipase Cdelta3 regulates RhoA/Rho kinase signaling and neurite outgrowth.

    PubMed

    Kouchi, Zen; Igarashi, Takahiro; Shibayama, Nami; Inanobe, Shunichi; Sakurai, Kazuyuki; Yamaguchi, Hideki; Fukuda, Toshifumi; Yanagi, Shigeru; Nakamura, Yoshikazu; Fukami, Kiyoko

    2011-03-11

    Phospholipase Cδ3 (PLCδ3) is a key enzyme regulating phosphoinositide metabolism; however, its physiological function remains unknown. Because PLCδ3 is highly enriched in the cerebellum and cerebral cortex, we examined the role of PLCδ3 in neuronal migration and outgrowth. PLCδ3 knockdown (KD) inhibits neurite formation of cerebellar granule cells, and application of PLCδ3KD using in utero electroporation in the developing brain results in the retardation of the radial migration of neurons in the cerebral cortex. In addition, PLCδ3KD inhibits axon and dendrite outgrowth in primary cortical neurons. PLCδ3KD also suppresses neurite formation of Neuro2a neuroblastoma cells induced by serum withdrawal or treatment with retinoic acid. This inhibition is released by the reintroduction of wild-type PLCδ3. Interestingly, the H393A mutant lacking phosphatidylinositol 4,5-bisphosphate hydrolyzing activity generates supernumerary protrusions, and a constitutively active mutant promotes extensive neurite outgrowth, indicating that PLC activity is important for normal neurite outgrowth. The introduction of dominant negative RhoA (RhoA-DN) or treatment with Y-27632, a Rho kinase-specific inhibitor, rescues the neurite extension in PLCδ3KD Neuro2a cells. Similar effects were also detected in primary cortical neurons. Furthermore, the RhoA expression level was significantly decreased by serum withdrawal or retinoic acid in control cells, although this decrease was not observed in PLCδ3KD cells. We also found that exogenous expression of PLCδ3 down-regulated RhoA protein, and constitutively active PLCδ3 promotes the RhoA down-regulation more significantly than PLCδ3 upon differentiation. These results indicate that PLCδ3 negatively regulates RhoA expression, inhibits RhoA/Rho kinase signaling, and thereby promotes neurite extension.

  2. Conditional lethality of a yeast strain expressing human RHOA in place of RHO1.

    PubMed Central

    Qadota, H; Anraku, Y; Botstein, D; Ohya, Y

    1994-01-01

    The yeast RHO1 GTPase, which has 72% amino acid sequence identity with its human counterpart, RHOA, is essential for growth, although the reason has not been investigated. We report here that yeast strains that rely solely on expression of human RHOA in place of RHO1 are able to grow at 23 degrees C but grow neither at 37 degrees C nor in the presence of 300 mM CaCl2 even at 23 degrees C. Measurements of steady-state protein levels indicate that inability to grow at the restrictive temperature is not due to instability of the protein. Homolog scanning with the two GTPases identified a small, 27-residue region of RHO1 which, when substituted into RHOA, confers full function in yeast. This region corresponds to the alpha 3-helix loop 7 region of RAS; the same region was reported to determine specificity of function between GTPases of the RAB family, Sec4p and Ypt1p. By examining the phenotype of RHOA substitution strains at nonpermissive temperature, we found evidence suggesting that the normal function of RHO1 is to maintain osmotic integrity. Images PMID:7937763

  3. Rho activation patterns after spinal cord injury and the role of activated Rho in apoptosis in the central nervous system

    PubMed Central

    Dubreuil, Catherine I.; Winton, Matthew J.; McKerracher, Lisa

    2003-01-01

    Growth inhibitory proteins in the central nervous system (CNS) block axon growth and regeneration by signaling to Rho, an intracellular GTPase. It is not known how CNS trauma affects the expression and activation of RhoA. Here we detect GTP-bound RhoA in spinal cord homogenates and report that spinal cord injury (SCI) in both rats and mice activates RhoA over 10-fold in the absence of changes in RhoA expression. In situ Rho-GTP detection revealed that both neurons and glial cells showed Rho activation at SCI lesion sites. Application of a Rho antagonist (C3–05) reversed Rho activation and reduced the number of TUNEL-labeled cells by ∼50% in both injured mouse and rat, showing a role for activated Rho in cell death after CNS injury. Next, we examined the role of the p75 neurotrophin receptor (p75NTR) in Rho signaling. After SCI, an up-regulation of p75NTR was detected by Western blot and observed in both neurons and glia. Treatment with C3–05 blocked the increase in p75NTR expression. Experiments with p75NTR-null mutant mice showed that immediate Rho activation after SCI is p75NTR dependent. Our results indicate that blocking overactivation of Rho after SCI protects cells from p75NTR-dependent apoptosis. PMID:12860969

  4. Rho family and Rap GTPase activation assays.

    PubMed

    Jennings, Richard T; Knaus, Ulla G

    2014-01-01

    The detection of Ras superfamily GTPase activity in innate immune cells is important when studying signaling events elicited by various ligands and cellular processes. The development of high-affinity probes detecting the activated, GTP-bound form of small GTPases has significantly enhanced our understanding of initiation and termination of GTPase-regulated signaling pathways. These probes are created by fusing a high-affinity GTPase-binding domain derived from a specific downstream effector protein to glutathione S-transferase (GST). Such domains bind preferentially to the GTP-bound form of the upstream Rho or Ras GTPase. Coupling these probes to beads enables extraction of the complex and subsequent quantification of the active GTP-binding protein by immunoblotting. Although effector domains that discriminate efficiently between GDP- and GTP-bound states and highly specific antibodies are not yet available for every small GTPase, analysis of certain members of the Rho and Ras GTPase family is now routinely performed. Here, we describe affinity-based pulldown assays for detection of Rho GTPase (Rac1/2, Cdc42, RhoA/B) and Rap1/2 activity in stimulated neutrophils or macrophages.

  5. Immunological and Functional Characterization of RhoGDI3 and Its Molecular Targets RhoG and RhoB in Human Pancreatic Cancerous and Normal Cells.

    PubMed

    de León-Bautista, Mercedes Piedad; Cardenas-Aguayo, Maria Del Carmen; Casique-Aguirre, Diana; Almaraz-Salinas, Manuel; Parraguirre-Martinez, Sara; Olivo-Diaz, Angelica; Thompson-Bonilla, María Del Rocío; Vargas, Miguel

    2016-01-01

    RhoGDI proteins have been implicated in several human cancers; changes in their expression levels have shown pro- or anti-tumorigenic effects. Pancreatic Ductal Adenocarcinoma (PDAC) is a complex pathology, with poor prognosis, and most patients die shortly after diagnosis. Efforts have been focused on understanding the role of RhoGDI's in PDAC, specially, RhoGDI1 and RhoGDI2. However, the role of RhoGDI3 has not been studied in relation to cancer or to PDAC. Here, we characterized the expression and functionality of RhoGDI3 and its target GTPases, RhoG and RhoB in pancreatic cell lines from both normal pancreatic tissue and tissue in late stages of PDAC, and compared them to human biopsies. Through immunofluorescences, pulldown assays and subcellular fractionation, we found a reduction in RhoGDI3 expression in the late stages of PDAC, and this reduction correlates with tumor progression and aggressiveness. Despite the reduction in the expression of RhoGDI3 in PDAC, we found that RhoB was underexpressed while RhoG was overexpressed, suggesting that cancerous cells preserve their capacity to activate this pathway, thus these cells may be more eager to response to the stimuli needed to proliferate and become invasive unlike normal cells. Surprisingly, we found nuclear localization of RhoGDI3 in non-cancerous pancreatic cell line and normal pancreatic tissue biopsies, which could open the possibility of novel nuclear functions for this protein, impacting gene expression regulation and cellular homeostasis.

  6. Immunological and Functional Characterization of RhoGDI3 and Its Molecular Targets RhoG and RhoB in Human Pancreatic Cancerous and Normal Cells

    PubMed Central

    de León-Bautista, Mercedes Piedad; Cardenas-Aguayo, Maria del Carmen; Casique-Aguirre, Diana; Almaraz-Salinas, Manuel; Parraguirre-Martinez, Sara; Olivo-Diaz, Angelica; Thompson-Bonilla, María del Rocío

    2016-01-01

    RhoGDI proteins have been implicated in several human cancers; changes in their expression levels have shown pro- or anti-tumorigenic effects. Pancreatic Ductal Adenocarcinoma (PDAC) is a complex pathology, with poor prognosis, and most patients die shortly after diagnosis. Efforts have been focused on understanding the role of RhoGDI's in PDAC, specially, RhoGDI1 and RhoGDI2. However, the role of RhoGDI3 has not been studied in relation to cancer or to PDAC. Here, we characterized the expression and functionality of RhoGDI3 and its target GTPases, RhoG and RhoB in pancreatic cell lines from both normal pancreatic tissue and tissue in late stages of PDAC, and compared them to human biopsies. Through immunofluorescences, pulldown assays and subcellular fractionation, we found a reduction in RhoGDI3 expression in the late stages of PDAC, and this reduction correlates with tumor progression and aggressiveness. Despite the reduction in the expression of RhoGDI3 in PDAC, we found that RhoB was underexpressed while RhoG was overexpressed, suggesting that cancerous cells preserve their capacity to activate this pathway, thus these cells may be more eager to response to the stimuli needed to proliferate and become invasive unlike normal cells. Surprisingly, we found nuclear localization of RhoGDI3 in non-cancerous pancreatic cell line and normal pancreatic tissue biopsies, which could open the possibility of novel nuclear functions for this protein, impacting gene expression regulation and cellular homeostasis. PMID:27832197

  7. Rho/Rho-dependent kinase affects locomotion and actin-myosin II activity of Amoeba proteus.

    PubMed

    Kłopocka, W; Redowicz, M J

    2004-10-01

    The highly motile free-living unicellular organism Amoeba proteus has been widely used as a model to study cell motility. However, the molecular mechanisms underlying its unique locomotion are still scarcely known. Recently, we have shown that blocking the amoebae's endogenous Rac- and Rho-like proteins led to distinct and irreversible changes in the appearance of these large migrating cells as well as to a significant inhibition of their locomotion. In order to elucidate the mechanism of the Rho pathway, we tested the effects of blocking the endogenous Rho-dependent kinase (ROCK) by anti-ROCK antibodies and Y-27632, (+)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl)cyclohexanecarboxamide dihydrochloride, a specific inhibitor of ROCK, on migrating amoebae and the effect of the Rho and ROCK inhibition on the actin-activated Mg-ATPase of the cytosolic fraction of the amoebae. Amoebae microinjected with anti-ROCK inhibitors remained contracted and strongly attached to the glass surface and exhibited an atypical locomotion. Despite protruding many pseudopodia that were advancing in various directions, the amoebae could not effectively move. Immunofluorescence studies showed that ROCK-like protein was dispersed throughout the cytoplasm and was also found in the regions of actin-myosin II interaction during both isotonic and isometric contraction. The Mg-ATPase activity was about two- to threefold enhanced, indicating that blocking the Rho/Rho-dependent kinase activated myosin. It is possible then that in contrast to the vertebrate cells, the inactivation of Rho/Rho-dependent kinase in amoebae leads to the activation of myosin II and to the observed hypercontracted cells which cannot exert effective locomotion.

  8. Y-39983 downregulates RhoA/Rho-associated kinase expression during its promotion of axonal regeneration.

    PubMed

    Yang, Zijian; Wang, Jing; Liu, Xiaohong; Cheng, Yu; Deng, Lianfu; Zhong, Yisheng

    2013-03-01

    Y-39983, a selective Rho-associated kinase (ROCK) inhibitor, promotes axonal regeneration of damaged retinal ganglion cells (RGCs). The present study investigated the effects of Y-39983 on RhoA/ROCK expression during promotion of axonal regeneration using a rat optic nerve crush (ONC) model. Herein, we demonstrated that Y-39983 significantly enhanced the survival and axonal regeneration of RGCs after ONC. Using a pull‑down assay and affinity precipitation to examine the activity of RhoA, we detected the decreased expression of active-RhoA after using Y-39983. The expression of ROCK1 and ROCK2 was significantly decreased as demonstrated by RT-PCR, immunohistochemistry and western blot analysis. The downregulation of active-RhoA, ROCK1 and ROCK2 expression by Y-39983 coincided with the appearance of larger numbers of regenerating axons. In conclusion, Y-39983 downregulated the expression of active-RhoA, ROCK1 and ROCK2 during its promotion of axonal regeneration.

  9. Rho GTPases at the crossroad of signaling networks in mammals

    PubMed Central

    Wojnacki, José; Quassollo, Gonzalo; Marzolo, María-Paz; Cáceres, Alfredo

    2014-01-01

    Microtubule (MT) organization and dynamics downstream of external cues is crucial for maintaining cellular architecture and the generation of cell asymmetries. In interphase cells RhoA, Rac, and Cdc42, conspicuous members of the family of small Rho GTPases, have major roles in modulating MT stability, and hence polarized cell behaviors. However, MTs are not mere targets of Rho GTPases, but also serve as signaling platforms coupling MT dynamics to Rho GTPase activation in a variety of cellular conditions. In this article, we review some of the key studies describing the reciprocal relationship between small Rho-GTPases and MTs during migration and polarization. PMID:24691223

  10. The ubiquitin-like protein LC3 regulates the Rho-GEF activity of AKAP-Lbc.

    PubMed

    Baisamy, Laurent; Cavin, Sabrina; Jurisch, Nathalie; Diviani, Dario

    2009-10-09

    AKAP-Lbc is a member of the A-kinase anchoring protein (AKAP) family that has been recently associated with the development of pathologies, such as cardiac hypertrophy and cancer. We have previously demonstrated that, at the molecular level, AKAP-Lbc functions as a guanine nucleotide exchange factor (GEF) that promotes the specific activation of RhoA. In the present study, we identified the ubiquitin-like protein LC3 as a novel regulatory protein interacting with AKAP-Lbc. Mutagenesis studies revealed that LC3, through its NH(2)-terminal alpha-helical domain, interacts with two binding sites located within the NH(2)-terminal regulatory region of AKAP-Lbc. Interestingly, LC3 overexpression strongly reduced the ability of AKAP-Lbc to interact with RhoA, profoundly impairing the Rho-GEF activity of the anchoring protein and, as a consequence, its ability to promote cytoskeletal rearrangements associated with the formation of actin stress fibers. Moreover, AKAP-Lbc mutants that fail to interact with LC3 show a higher basal Rho-GEF activity as compared with the wild type protein and become refractory to the inhibitory effect of LC3. This suggests that LC3 binding maintains AKAP-Lbc in an inactive state that displays a reduced ability to promote downstream signaling. Collectively, these findings provide evidence for a previously uncharacterized role of LC3 in the regulation of Rho signaling and in the reorganization of the actin cytoskeleton.

  11. RhoA GTPase inhibition organizes contraction during epithelial morphogenesis

    PubMed Central

    Mason, Frank M.; Xie, Shicong; Vasquez, Claudia G.; Tworoger, Michael

    2016-01-01

    During morphogenesis, contraction of the actomyosin cytoskeleton within individual cells drives cell shape changes that fold tissues. Coordination of cytoskeletal contractility is mediated by regulating RhoA GTPase activity. Guanine nucleotide exchange factors (GEFs) activate and GTPase-activating proteins (GAPs) inhibit RhoA activity. Most studies of tissue folding, including apical constriction, have focused on how RhoA is activated by GEFs to promote cell contractility, with little investigation as to how GAPs may be important. Here, we identify a critical role for a RhoA GAP, Cumberland GAP (C-GAP), which coordinates with a RhoA GEF, RhoGEF2, to organize spatiotemporal contractility during Drosophila melanogaster apical constriction. C-GAP spatially restricts RhoA pathway activity to a central position in the apical cortex. RhoGEF2 pulses precede myosin, and C-GAP is required for pulsation, suggesting that contractile pulses result from RhoA activity cycling. Finally, C-GAP expression level influences the transition from reversible to irreversible cell shape change, which defines the onset of tissue shape change. Our data demonstrate that RhoA activity cycling and modulating the ratio of RhoGEF2 to C-GAP are required for tissue folding. PMID:27551058

  12. Rho GTPases at the crossroad of signaling networks in mammals: impact of Rho-GTPases on microtubule organization and dynamics.

    PubMed

    Wojnacki, José; Quassollo, Gonzalo; Marzolo, María-Paz; Cáceres, Alfredo

    2014-01-01

    Microtubule (MT) organization and dynamics downstream of external cues is crucial for maintaining cellular architecture and the generation of cell asymmetries. In interphase cells RhoA, Rac, and Cdc42, conspicuous members of the family of small Rho GTPases, have major roles in modulating MT stability, and hence polarized cell behaviors. However, MTs are not mere targets of Rho GTPases, but also serve as signaling platforms coupling MT dynamics to Rho GTPase activation in a variety of cellular conditions. In this article, we review some of the key studies describing the reciprocal relationship between small Rho-GTPases and MTs during migration and polarization.

  13. Small Rho-GTPases and cortical malformations

    PubMed Central

    2013-01-01

    Rho-GTPases have been found to be crucial for cytoskeleton remodelling and cell polarity, as well as key players in directed cell migration in various tissues and organs, therefore becoming good candidates for involvement in neuronal migration disorders. We recently found that genetic deletion of the small GTPase RhoA in the developing mouse cerebral cortex results in three distinct cortical malformations: a defect in the proliferation of progenitor cells during development that leads to a bigger cerebral cortex in the adult mouse, a change in the morphology of radial glial cells that results in the formation of a subcortical band heterotopia (SBH, also called Double Cortex) and an increase in the speed of migrating newborn neurons. The latter, together with the aberrant radial glial shape, is likely to be the cause of cobblestone lissencephaly, where neurons protrude beyond layer I at the pial surface of the brain. PMID:23524873

  14. Vector meson dominance and the {rho} meson

    SciTech Connect

    Benayoun, M.; OConnell, H.B.; Williams, A.G.

    1999-04-01

    We discuss the properties of vector mesons, in particular the {rho}{sup 0}, in the context of the hidden local symmetry (HLS) model. This provides a unified framework to study several aspects of the low energy QCD sector. First, we show that in the HLS model the physical photon is massless, without requiring off field diagonalization. We then demonstrate the equivalence of HLS and the two existing representations of vector meson dominance, VMD1 and VMD2, at both the tree level and one loop order. Finally the S matrix pole position is shown to provide a model and process independent means of specifying the {rho} mass and width, in contrast with the real axis prescription currently used in the Particle Data Group tables. {copyright} {ital 1999} {ital The American Physical Society}

  15. Rho kinase inhibition in diabetic kidney disease.

    PubMed

    Komers, Radko

    2013-10-01

    Small GTPases of the Rho family and their down-stream effectors Rho associated kinases (ROCKs) are the molecules that converge a spectrum of pathophysiological signals triggered by the diabetic milieu and represent promising molecular targets for nephroprotective treatment in diabetes. The review discusses recent studies exploring the consequences of diabetes-induced Rho-ROCK activation in the kidney and the effects of ROCK inhibition (ROCKi) in experimental diabetic kidney disease (DKD). Studies in models of type 1 and type 2 diabetes have indicated blood pressure-independent nephroprotective actions of ROCKi in DKD. The underlying mechanisms include attenuation of diabetes-induced increases in renal expression of prosclerotic cytokines and extracellular matrix, anti-oxidant effects and protection of mitochondrial function, resulting in slower development of glomerulosclerosis and interstitial fibrosis. The studies have also shown antiproteinuric effects of ROCKi that could be related to reductions in permeability of the glomerular barrier and beneficial effects on podocytes. Glomerular haemodynamic mechanisms might also be involved. Despite remaining questions in this field, such as the effects in podocytes later in the course of DKD, specificity of currently available ROCKi, or the roles of individual ROCK isoforms, recent evidence in experimental diabetes suggests that ROCKi might in future broaden the spectrum of treatments available for patients with DKD. This is supported by the evidence generated in models of non-diabetic kidney disease and in clinical studies in patients with various cardiovascular disorders.

  16. BAR domain proteins regulate Rho GTPase signaling

    PubMed Central

    Aspenström, Pontus

    2014-01-01

    BAR proteins comprise a heterogeneous group of multi-domain proteins with diverse biological functions. The common denominator is the Bin-Amphiphysin-Rvs (BAR) domain that not only confers targeting to lipid bilayers, but also provides scaffolding to mold lipid membranes into concave or convex surfaces. This function of BAR proteins is an important determinant in the dynamic reconstruction of membrane vesicles, as well as of the plasma membrane. Several BAR proteins function as linkers between cytoskeletal regulation and membrane dynamics. These links are provided by direct interactions between BAR proteins and actin-nucleation-promoting factors of the Wiskott-Aldrich syndrome protein family and the Diaphanous-related formins. The Rho GTPases are key factors for orchestration of this intricate interplay. This review describes how BAR proteins regulate the activity of Rho GTPases, as well as how Rho GTPases regulate the function of BAR proteins. This mutual collaboration is a central factor in the regulation of vital cellular processes, such as cell migration, cytokinesis, intracellular transport, endocytosis, and exocytosis. PMID:25483303

  17. The new Be-type star HD 147196 in the Rho Ophiuchi dark cloud region

    NASA Technical Reports Server (NTRS)

    The, P. S.; Perez, M. R.; De Winter, D.; Van Den Ancker, M. E.

    1993-01-01

    The newly discovered hot-emission line star, HD 147196 in the Rho Oph dark cloud region was observed spectroscopically and photometrically and high and low resolution IUE spectra were obtained. The finding of Irvine (1990) that this relatively bright star show its H-alpha-line in emission is confirmed. Previous H-alpha-surveys of the Rho Oph star-forming region did not detect HD 147196 as an H-alpha-emission star, meaning that it must recently be very active and has perhaps transformed itself from a B-type star at shell phase to a Be-phase. The Mg II h + k resonance lines are in absorption and they appear to be interstellar in nature, which means that either the abundance of Mg in the extended atmosphere of the star is low or that the shell is not extended enough to produce emission lines of Mg II. Photometric observations of this B8 V type star do not show any variations during at least the years covered by our monitoring or any excess of NIR radiation in its spectral energy distribution up to the M-passband at 4.8 microns.

  18. Generating Rho-0 Cells Using Mesenchymal Stem Cell Lines

    PubMed Central

    Fernández-Moreno, Mercedes; Hermida-Gómez, Tamara; Gallardo, M. Esther; Dalmao-Fernández, Andrea; Rego-Pérez, Ignacio; Garesse, Rafael

    2016-01-01

    Introduction The generation of Rho-0 cells requires the use of an immortalization process, or tumor cell selection, followed by culture in the presence of ethidium bromide (EtBr), incurring the drawbacks its use entails. The purpose of this work was to generate Rho-0 cells using human mesenchymal stem cells (hMSCs) with reagents having the ability to remove mitochondrial DNA (mtDNA) more safely than by using EtBr. Methodology Two immortalized hMSC lines (3a6 and KP) were used; 143B.TK-Rho-0 cells were used as reference control. For generation of Rho-0 hMSCs, cells were cultured in medium supplemented with each tested reagent. Total DNA was isolated and mtDNA content was measured by real-time polymerase chain reaction (PCR). Phenotypic characterization and gene expression assays were performed to determine whether 3a6 Rho-0 hMSCs maintain the same stem properties as untreated 3a6 hMSCs. To evaluate whether 3a6 Rho-0 hMSCs had a phenotype similar to that of 143B.TK-Rho-0 cells, in terms of reactive oxygen species (ROS) production, apoptotic levels and mitochondrial membrane potential (Δψm) were measured by flow cytometry and mitochondrial respiration was evaluated using a SeaHorse XFp Extracellular Flux Analyzer. The differentiation capacity of 3a6 and 3a6 Rho-0 hMSCs was evaluated using real-time PCR, comparing the relative expression of genes involved in osteogenesis, adipogenesis and chondrogenesis. Results The results showed the capacity of the 3a6 cell line to deplete its mtDNA and to survive in culture with uridine. Of all tested drugs, Stavudine (dt4) was the most effective in producing 3a6-Rho cells. The data indicate that hMSC Rho-0 cells continue to express the characteristic MSC cell surface receptor pattern. Phenotypic characterization showed that 3a6 Rho-0 cells resembled 143B.TK-Rho-0 cells, indicating that hMSC Rho-0 cells are Rho-0 cells. While the adipogenic capability was higher in 3a6 Rho-0 cells than in 3a6 cells, the osteogenic and chondrogenic

  19. A negative modulatory role for rho and rho-associated kinase signaling in delamination of neural crest cells

    PubMed Central

    Groysman, Maya; Shoval, Irit; Kalcheim, Chaya

    2008-01-01

    Background Neural crest progenitors arise as epithelial cells and then undergo a process of epithelial to mesenchymal transition that precedes the generation of cellular motility and subsequent migration. We aim at understanding the underlying molecular network. Along this line, possible roles of Rho GTPases that act as molecular switches to control a variety of signal transduction pathways remain virtually unexplored, as are putative interactions between Rho proteins and additional known components of this cascade. Results We investigated the role of Rho/Rock signaling in neural crest delamination. Active RhoA and RhoB are expressed in the membrane of epithelial progenitors and are downregulated upon delamination. In vivo loss-of-function of RhoA or RhoB or of overall Rho signaling by C3 transferase enhanced and/or triggered premature crest delamination yet had no effect on cell specification. Consistently, treatment of explanted neural primordia with membrane-permeable C3 or with the Rock inhibitor Y27632 both accelerated and enhanced crest emigration without affecting cell proliferation. These treatments altered neural crest morphology by reducing stress fibers, focal adhesions and downregulating membrane-bound N-cadherin. Reciprocally, activation of endogenous Rho by lysophosphatidic acid inhibited emigration while enhancing the above. Since delamination is triggered by BMP and requires G1/S transition, we examined their relationship with Rho. Blocking Rho/Rock function rescued crest emigration upon treatment with noggin or with the G1/S inhibitor mimosine. In the latter condition, cells emigrated while arrested at G1. Conversely, BMP4 was unable to rescue cell emigration when endogenous Rho activity was enhanced by lysophosphatidic acid. Conclusion Rho-GTPases, through Rock, act downstream of BMP and of G1/S transition to negatively regulate crest delamination by modifying cytoskeleton assembly and intercellular adhesion. PMID:18945340

  20. RhoGTPase Regulators Orchestrate Distinct Stages of Synaptic Development

    PubMed Central

    Martin-Vilchez, Samuel; Whitmore, Leanna; Asmussen, Hannelore; Zareno, Jessica; Horwitz, Rick; Newell-Litwa, Karen

    2017-01-01

    Small RhoGTPases regulate changes in post-synaptic spine morphology and density that support learning and memory. They are also major targets of synaptic disorders, including Autism. Here we sought to determine whether upstream RhoGTPase regulators, including GEFs, GAPs, and GDIs, sculpt specific stages of synaptic development. The majority of examined molecules uniquely regulate either early spine precursor formation or later maturation. Specifically, an activator of actin polymerization, the Rac1 GEF β-PIX, drives spine precursor formation, whereas both FRABIN, a Cdc42 GEF, and OLIGOPHRENIN-1, a RhoA GAP, regulate spine precursor elongation. However, in later development, a novel Rac1 GAP, ARHGAP23, and RhoGDIs inactivate actomyosin dynamics to stabilize mature synapses. Our observations demonstrate that specific combinations of RhoGTPase regulatory proteins temporally balance RhoGTPase activity during post-synaptic spine development. PMID:28114311

  1. Rho-kinase inhibition in the therapy of cardiovascular disease.

    PubMed

    Lai, Andrew; Frishman, William H

    2005-01-01

    Rho is a GTPase known to be a major mediator in the formation of stress fibers and focal adhesions, cell morphology, and smooth muscle contraction. Its role in smooth muscle contraction has led to exploration into the connection between Rho-mediated kinase activity and cardiovascular disease. The role of Rho-kinase in calcium sensitization for vascular smooth muscle contraction has recently been characterized. Inappropriate coronary artery vasoconstriction resulting from increased Rho-kinase in the vascular system is likely involved in the pathogenesis of exercise-induced myocardial ischemia, spontaneous coronary artery spasm, and hypertension. In clinical trials, Rho-kinase inhibitors such as fasudil and Y-27632 have demonstrated antiischemic, antivasospastic, and antihypertensive effects. These compounds have also exhibited the ability to blunt progression of cardiomyocyte hypertrophy and cardiac remodeling in heart failure. As such, Rho-kinase inhibition represents a potential novel therapeutic approach in cardiovascular disease.

  2. High-{rho}R Implosions for Fast-Ignition Fuel Assembly

    SciTech Connect

    Zhou, C. D.; Betti, R.; Meyerhofer, D. D.; Theobald, W.; Radha, P. B.; Smalyuk, V. A.; Glebov, V. Yu.; Stoeckl, C.; Anderson, K. S.; Sangster, T. C.; Shvarts, D.; Li, C. K.; Petrasso, R. D.; Frenje, J. A.; Seguin, F. H.

    2007-01-12

    Thick, 40 {mu}m plastic shells filled with 25-35 atm of D{sub 2} or D{sup 3}He were imploded on a low-adiabat ({alpha}{approx_equal}1.3) and with a low-implosion velocity ({approx}2x10{sup 7} cm/s) on the OMEGA laser to generate massive cores of compressed plasma with high areal densities optimal for fast ignition. The targets are driven by 20-kJ relaxation adiabat-shaping laser pulses to keep the inner portion of the shell nearly Fermi degenerate. The measured kinetic energy downshift of proton spectra is in good agreement with the theoretical predictions yielding burn-averaged areal densities of 0.130{+-}0.017 g/cm{sup 2} and peak {rho}R during the burn of about 0.24{+-}0.018 g/cm{sup 2}, the largest {rho}R measured on OMEGA to date. The same implosions with empty plastic shells are expected to reach 1.3 g/cm{sup 2} across the core (i.e., 2{rho}R) enough to stop fast electrons with energies up to 4.5 MeV typical of fast ignition scenarios.

  3. Positioning the cleavage furrow: All you need is Rho

    PubMed Central

    Liu, Zairan

    2016-01-01

    RhoA controls cleavage furrow formation during cell division, but whether RhoA suffices to orchestrate spatiotemporal dynamics of furrow formation is unknown. In this issue, Wagner and Glotzer (2016. J. Cell Biol. http://dx.doi.org/10.1083/jcb.201603025) show that RhoA activity can induce furrow formation in all cell cortex positions and cell cycle phases. PMID:27325786

  4. Insights into the Molecular Activation Mechanism of the RhoA-specific Guanine Nucleotide Exchange Factor, PDZRhoGEF

    SciTech Connect

    Bielnicki, Jakub A.; Shkumatov, Alexander V.; Derewenda, Urszula; Somlyo, Avril V.; Svergun, Dmitri I.; Derewenda, Zygmunt S.

    2012-10-09

    PDZRhoGEF (PRG) belongs to a small family of RhoA-specific nucleotide exchange factors that mediates signaling through select G-protein-coupled receptors via G{alpha}{sub 12/13} and activates RhoA by catalyzing the exchange of GDP to GTP. PRG is a multidomain protein composed of PDZ, regulators of G-protein signaling-like (RGSL), Dbl-homology (DH), and pleckstrin-homology (PH) domains. It is autoinhibited in cytosol and is believed to undergo a conformational rearrangement and translocation to the membrane for full activation, although the molecular details of the regulation mechanism are not clear. It has been shown recently that the main autoregulatory elements of PDZRhoGEF, the autoinhibitory 'activation box' and the 'GEF switch,' which is required for full activation, are located directly upstream of the catalytic DH domain and its RhoA binding surface, emphasizing the functional role of the RGSL-DH linker. Here, using a combination of biophysical and biochemical methods, we show that the mechanism of PRG regulation is yet more complex and may involve an additional autoinhibitory element in the form of a molten globule region within the linker between RGSL and DH domains. We propose a novel, two-tier model of autoinhibition where the activation box and the molten globule region act synergistically to impair the ability of RhoA to bind to the catalytic DH-PH tandem. The molten globule region and the activation box become less ordered in the PRG-RhoA complex and dissociate from the RhoA-binding site, which may constitute a critical step leading to PRG activation.

  5. Functional Cross-talk between Ras and Rho Pathways

    PubMed Central

    Jaiswal, Mamta; Dvorsky, Radovan; Amin, Ehsan; Risse, Sarah L.; Fansa, Eyad K.; Zhang, Si-Cai; Taha, Mohamed S.; Gauhar, Aziz R.; Nakhaei-Rad, Saeideh; Kordes, Claus; Koessmeier, Katja T.; Cirstea, Ion C.; Olayioye, Monilola A.; Häussinger, Dieter; Ahmadian, Mohammad R.

    2014-01-01

    The three deleted in liver cancer genes (DLC1–3) encode Rho-specific GTPase-activating proteins (RhoGAPs). Their expression is frequently silenced in a variety of cancers. The RhoGAP activity, which is required for full DLC-dependent tumor suppressor activity, can be inhibited by the Src homology 3 (SH3) domain of a Ras-specific GAP (p120RasGAP). Here, we comprehensively investigated the molecular mechanism underlying cross-talk between two distinct regulators of small GTP-binding proteins using structural and biochemical methods. We demonstrate that only the SH3 domain of p120 selectively inhibits the RhoGAP activity of all three DLC isoforms as compared with a large set of other representative SH3 or RhoGAP proteins. Structural and mutational analyses provide new insights into a putative interaction mode of the p120 SH3 domain with the DLC1 RhoGAP domain that is atypical and does not follow the classical PXXP-directed interaction. Hence, p120 associates with the DLC1 RhoGAP domain by targeting the catalytic arginine finger and thus by competitively and very potently inhibiting RhoGAP activity. The novel findings of this study shed light on the molecular mechanisms underlying the DLC inhibitory effects of p120 and suggest a functional cross-talk between Ras and Rho proteins at the level of regulatory proteins. PMID:24443565

  6. Posttranslational lipid modification of Rho family small GTPases.

    PubMed

    Mitin, Natalia; Roberts, Patrick J; Chenette, Emily J; Der, Channing J

    2012-01-01

    The Rho family comprises a major branch of the Ras superfamily of small GTPases. A majority of Rho GTPases are synthesized as inactive, cytosolic proteins. They then undergo posttranslational modification by isoprenoid or fatty acid lipids, and together with additional carboxyl-terminal sequences target Rho GTPases to specific membrane and subcellular compartments essential for function. We summarize the use of biochemical and cellular assays and pharmacologic inhibitors instrumental for the study of the role of posttranslational lipid modifications and processing in Rho GTPase biology.

  7. rho/sup 0/. omega. production in photon photon interactions

    SciTech Connect

    Derby, K.A.

    1987-08-01

    The subject of this dissertation is the production of the rho/sup 0/..omega.. final state in photon photon interactions. The production of the rho/sup 0/..omega.. final state has been of interest primarily because of its similarity to the related process ..gamma gamma.. ..-->.. rho/sup 0/rho/sup 0/. The cross section for rho/sup 0/rho/sup 0/ production demonstrates a peaking near threshold, the mechanism of which has been the subject of considerable speculation. The data sample used for the analysis was obtained using the TPC detector facility at the PEP e/sup +/e/sup -/ storage ring, and corresponds to an integrated e/sup +/e/sup -/ luminosity of 64 pb/sup -1/ at 29 GeV center of mass energy. Our estimate of the rho/sup 0/..omega.. cross section is compared to the predictions of several models which have been used to account for the observed rho/sup 0/rho/sup 0/ cross section. The experimental results are consistent with the predictions of a threshold enhancement model, as well as those of a four quark (qq anti q anti q) resonance model. However, they disagree with the predictions of a t-channel factorization approach.

  8. Search for medium modifications of the rho meson.

    PubMed

    Nasseripour, R; Wood, M H; Djalali, C; Weygand, D P; Tur, C; Mosel, U; Muehlich, P; Adams, G; Amaryan, M J; Ambrozewicz, P; Anghinolfi, M; Asryan, G; Avakian, H; Bagdasaryan, H; Baillie, N; Ball, J P; Baltzell, N A; Barrow, S; Battaglieri, M; Bedlinskiy, I; Bektasoglu, M; Bellis, M; Benmouna, N; Berman, B L; Biselli, A S; Blaszczyk, L; Bouchigny, S; Boiarinov, S; Bradford, R; Branford, D; Briscoe, W J; Brooks, W K; Bültmann, S; Burkert, V D; Butuceanu, C; Calarco, J R; Careccia, S L; Carman, D S; Carnahan, B; Casey, L; Chen, S; Cole, P L; Collins, P; Coltharp, P; Crabb, D; Crannell, H; Crede, V; Cummings, J P; Dashyan, N; De Masi, R; De Vita, R; De Sanctis, E; Degtyarenko, P V; Denizli, H; Dennis, L; Deur, A; Dharmawardane, K V; Dickson, R; Dodge, G E; Doughty, D; Dugger, M; Dytman, S; Dzyubak, O P; Egiyan, H; Egiyan, K S; El Fassi, L; Elouadrhiri, L; Eugenio, P; Fedotov, G; Feldman, G; Feuerbach, R J; Funsten, H; Garçon, M; Gavalian, G; Gilfoyle, G P; Giovanetti, K L; Girod, F X; Goetz, J T; Gordon, C I O; Gothe, R W; Griffioen, K A; Guidal, M; Guler, N; Guo, L; Gyurjyan, V; Hadjidakis, C; Hafidi, K; Hakobyan, H; Hakobyan, R S; Hanretty, C; Hardie, J; Hersman, F W; Hicks, K; Hleiqawi, I; Holtrop, M; Hyde-Wright, C E; Ilieva, Y; Ireland, D G; Ishkhanov, B S; Isupov, E L; Ito, M M; Jenkins, D; Jo, H S; Johnstone, J R; Joo, K; Juengst, H G; Kalantarians, N; Kellie, J D; Khandaker, M; Kim, W; Klein, A; Klein, F J; Klimenko, A V; Kossov, M; Krahn, Z; Kramer, L H; Kubarovsky, V; Kuhn, J; Kuhn, S E; Kuleshov, S V; Lachniet, J; Laget, J M; Langheinrich, J; Lawrence, D; Li, Ji; Livingston, K; Lu, H Y; Maccormick, M; Markov, N; Mattione, P; McAleer, S; McKinnon, B; McNabb, J W C; Mecking, B A; Mehrabyan, S; Melone, J J; Mestayer, M D; Meyer, C A; Mibe, T; Mikhailov, K; Minehart, R; Mirazita, M; Miskimen, R; Mokeev, V; Moriya, K; Morrow, S A; Moteabbed, M; Mueller, J; Munevar, E; Mutchler, G S; Nadel-Turonski, P; Niccolai, S; Niculescu, G; Niculescu, I; Niczyporuk, B B; Niroula, M R; Niyazov, R A; Nozar, M; Osipenko, M; Ostrovidov, A I; Park, K; Pasyuk, E; Paterson, C; Anefalos Pereira, S; Pierce, J; Pivnyuk, N; Pocanic, D; Pogorelko, O; Pozdniakov, S; Preedom, B M; Price, J W; Prok, Y; Protopopescu, D; Raue, B A; Riccardi, G; Ricco, G; Ripani, M; Ritchie, B G; Ronchetti, F; Rosner, G; Rossi, P; Sabatié, F; Salamanca, J; Salgado, C; Santoro, J P; Sapunenko, V; Schumacher, R A; Serov, V S; Sharabian, Y G; Sharov, D; Shvedunov, N V; Smith, E S; Smith, L C; Sober, D I; Sokhan, D; Stavinsky, A; Stepanyan, S S; Stepanyan, S; Stokes, B E; Stoler, P; Strakovsky, I I; Strauch, S; Taiuti, M; Tedeschi, D J; Tkabladze, A; Tkachenko, S; Todor, L; Ungaro, M; Vineyard, M F; Vlassov, A V; Watts, D P; Weinstein, L B; Williams, M; Wolin, E; Yegneswaran, A; Zana, L; Zhang, B; Zhang, J; Zhao, B; Zhao, Z W

    2007-12-31

    The photoproduction of vector mesons on various nuclei has been studied using the CLAS detector at Jefferson Laboratory. The vector mesons, rho, omega, and varphi, are observed via their decay to e;{+}e;{-}, in order to reduce the effects of final-state interactions in the nucleus. Of particular interest are possible in-medium effects on the properties of the rho meson. The rho mass spectrum is extracted from the data on various nuclei, 2H, C, Fe, and Ti. We observe no significant mass shift and some broadening consistent with expected collisional broadening for the rho meson.

  9. Rho kinase as a target for cerebral vascular disorders

    PubMed Central

    Bond, Lisa M; Sellers, James R; McKerracher, Lisa

    2015-01-01

    The development of novel pharmaceutical treatments for disorders of the cerebral vasculature is a serious unmet medical need. These vascular disorders are typified by a disruption in the delicate Rho signaling equilibrium within the blood vessel wall. In particular, Rho kinase overactivation in the smooth muscle and endothelial layers of the vessel wall results in cytoskeletal modifications that lead to reduced vascular integrity and abnormal vascular growth. Rho kinase is thus a promising target for the treatment of cerebral vascular disorders. Indeed, preclinical studies indicate that Rho kinase inhibition may reduce the formation/growth/rupture of both intracranial aneurysms and cerebral cavernous malformations. PMID:26062400

  10. Key role of the RhoA/Rho kinase system in pulmonary hypertension.

    PubMed

    Connolly, Michelle J; Aaronson, Philip I

    2011-02-01

    Pulmonary hypertension (PH) is a general term comprising a spectrum of pulmonary hypertensive disorders which have in common an elevation of mean pulmonary arterial pressure (mPAP). The prototypical form of the disease, termed pulmonary arterial hypertension (PAH), is a rare but lethal syndrome with a complex aetiology characterised by increased pulmonary vascular resistance (PVR) and progressive elevation of mPAP; patients generally die from heart failure. Current therapies are inadequate and median survival is less than three years. PH due to chronic hypoxia (CH) is a condition separate from PAH and is strongly associated with chronic obstructive pulmonary disease (COPD). An early event in the pathogenesis of this form of PH is hypoxic pulmonary vasoconstriction (HPV), an acute homeostatic process that maintains the ventilation-perfusion ratio during alveolar hypoxia. The mechanisms underlying HPV remain controversial, but RhoA/Rho kinase (ROK)-mediated Ca²+-sensitisation is considered important. Increasing evidence also implicates RhoA/ROK in PASMC proliferation, inflammatory cell recruitment and the regulation of cell motility, all of which are involved in the pulmonary vascular remodelling occurring in all forms of PH. ROK is therefore a potential therapeutic target in treating PH of various aetiologies. Here, we examine current concepts regarding the aetiology of PAH and also PH due to CH, focusing on the contribution that RhoA/ROK-mediated processes may make to their development and on ROK inhibitors as potential therapies.

  11. G alpha12 is targeted to the mitochondria and affects mitochondrial morphology and motility.

    PubMed

    Andreeva, Alexandra V; Kutuzov, Mikhail A; Voyno-Yasenetskaya, Tatyana A

    2008-08-01

    G alpha12 constitutes, along with G alpha13, one of the four families of alpha subunits of heterotrimeric G proteins. We found that the N terminus of G alpha12, but not those of other G alpha subunits, contains a predicted mitochondrial targeting sequence. Using confocal microscopy and cell fractionation, we demonstrated that up to 40% of endogenous G alpha12 in human umbilical vein endothelial cells colocalize with mitochondrial markers. N-terminal sequence of G alpha12 fused to GFP efficiently targeted the fusion protein to mitochondria. G alpha12 with mutated mitochondrial targeting sequence was still located in mitochondria, suggesting the existence of additional mechanisms for mitochondrial localization. Lysophosphatidic acid, one of the known stimuli transduced by G alpha12/13, inhibited mitochondrial motility, while depletion of endogenous G alpha12 increased mitochondrial motility. G alpha12Q229L variants uncoupled from RhoGEFs (but not fully functional activated G alpha12Q229L) induced transformation of the mitochondrial network into punctate mitochondria and resulted in a loss of mitochondrial membrane potential. All examined G alpha12Q229L variants reduced phosphorylation of Bcl-2 at Ser-70, while only mutants unable to bind RhoGEFs also decreased cellular levels of Bcl-2. These G alpha12 mutants were also more efficient Hsp90 interactors. These findings are the first demonstration of a heterotrimeric G protein alpha subunit specifically targeted to mitochondria and involved in the control of mitochondrial morphology and dynamics.

  12. Testing the dynamics of B{yields}{pi}{pi} and constraints on {alpha}

    SciTech Connect

    Grossman, Yuval; Hoecker, Andreas; Ligeti, Zoltan; Pirjol, Dan

    2005-11-01

    In charmless nonleptonic B decays to {pi}{pi} or {rho}{rho}, the 'color allowed' and 'color suppressed' tree amplitudes can be studied in a systematic expansion in {alpha}{sub s}(m{sub b}) and {lambda}{sub QCD}/m{sub b}. At leading order in this expansion their relative strong phase vanishes. The implications of this prediction are obscured by penguin contributions. We propose to use this prediction to test the relative importance of the various penguin amplitudes using experimental data. The present B{yields}{pi}{pi} data suggest that there are large corrections to the heavy quark limit, which can be due to power corrections to the tree amplitudes, large up-penguin amplitude, or enhanced weak annihilation. Because the penguin contributions are smaller, the heavy quark limit is more consistent with the B{yields}{rho}{rho} data, and its implications may become important for the extraction of {alpha} from this mode in the future.

  13. Spatiotemporal analysis of RhoA/B/C activation in primary human endothelial cells

    PubMed Central

    Reinhard, Nathalie R.; van Helden, Suzanne F.; Anthony, Eloise C.; Yin, Taofei; Wu, Yi I.; Goedhart, Joachim; Gadella, Theodorus W. J.; Hordijk, Peter L.

    2016-01-01

    Endothelial cells line the vasculature and are important for the regulation of blood pressure, vascular permeability, clotting and transendothelial migration of leukocytes and tumor cells. A group of proteins that that control the endothelial barrier function are the RhoGTPases. This study focuses on three homologous (>88%) RhoGTPases: RhoA, RhoB, RhoC of which RhoB and RhoC have been poorly characterized. Using a RhoGTPase mRNA expression analysis we identified RhoC as the highest expressed in primary human endothelial cells. Based on an existing RhoA FRET sensor we developed new RhoB/C FRET sensors to characterize their spatiotemporal activation properties. We found all these RhoGTPase sensors to respond to physiologically relevant agonists (e.g. Thrombin), reaching transient, localized FRET ratio changes up to 200%. These RhoA/B/C FRET sensors show localized GEF and GAP activity and reveal spatial activation differences between RhoA/C and RhoB. Finally, we used these sensors to monitor GEF-specific differential activation of RhoA/B/C. In summary, this study adds high-contrast RhoB/C FRET sensors to the currently available FRET sensor toolkit and uncover new insights in endothelial and RhoGTPase cell biology. This allows us to study activation and signaling by these closely related RhoGTPases with high spatiotemporal resolution in primary human cells. PMID:27147504

  14. Study of Branching Fractions and CP-Violating Asymmetries in B Meson Decays to Rho And Pion Final State with the BABAR Detector

    SciTech Connect

    Wu, Jinwei; /Wisconsin U., Madison

    2006-03-22

    We present measurements of branching fractions and CP-violating asymmetries in B-meson decays to {rho}{sup +}{pi}{sup 0}, {rho}{sup 0}{pi}{sup +} and {rho}{sup 0}{pi}{sup 0}. The data sample comprises 89 x 10{sup 6} {Upsilon}(4S) {yields} B{bar B} decays collected with the BABAR detector at the PEP-II asymmetric-energy B Factory at SLAC. We find the charge-averaged branching fractions {Beta}(B{sup +} {yields} {rho}{sup +}{pi}{sup 0}) = (10.9 {+-} 1.9(stat) {+-} 1.9(syst)) x 10{sup -6} and {Beta}(B{sup 0} {yields} {rho}{sup 0}{pi}{sup +}) = (9.5 {+-} 1.1 {+-} 0.9) x 10{sup -6}, and we set a 90% confidence-level upper limit {Beta}(B{sup 0} {yields} {rho}{sup 0}{pi}{sup 0}) < 2.9 x 10{sup -6}. We measure the charge asymmetries A{sub CP}{rho}{sup +}{pi}{sup 0} = 0.24 {+-} 0.16 {+-} 0.06 and {Alpha}{sub CP}{sup {rho}{sup 0}{pi}{sup +}} = -0.19 {+-} 0.11 {+-} 0.02. We also present the preliminary measurement of CP-violating asymmetries in B{sup 0} {yields} ({rho}{pi}){sup 0} {yields} {pi}{sup +}{pi}{sup -}{pi}{sup 0} decays using a time-dependent Dalitz plot analysis. The results are obtained from a data sample of 213 million {Upsilon}(4S) {yields} B{bar B} decays, collected by the BABAR detector at the PEP-II asymmetric-energy B Factory at SLAC. This analysis extends the narrow-{rho} quasi-two-body approximation used in the previous analysis, by taking into account the interference between the {rho} resonances of the three charges. We measure 16 coefficients of the bilinear form factor terms occurring in the time-dependent decay rate of the B{sup 0} meson with the use of a maximum-likelihood fit. We derive the physically relevant quantities from these coefficients. We measure the direct CP-violation parameters {Alpha}{sub {rho}{pi}} = -0.088 {+-} 0.049 {+-} 0.013 and C = 0.34 {+-} 0.11 {+-} 0.05, where the first errors are statistical and the second systematic. For the mixing-induced CP-violation parameter we find S = -0.10 {+-} 0.14 {+-} 0.04, and for the dilution and

  15. Regulation of phagocytosis by Rho GTPases.

    PubMed

    Mao, Yingyu; Finnemann, Silvia C

    2015-01-01

    Phagocytosis is defined as a cellular uptake pathway for particles of greater than 0.5 μm in diameter. Particle clearance by phagocytosis is of critical importance for tissue health and homeostasis. The ultimate goal of anti-pathogen phagocytosis is to destroy engulfed bacteria or fungi and to stimulate cell-cell signaling that mount an efficient immune defense. In contrast, clearance phagocytosis of apoptotic cells and cell debris is anti-inflammatory. High capacity clearance phagocytosis pathways are available to professional phagocytes of the immune system and the retina. Additionally, a low capacity, so-called bystander phagocytic pathway is available to most other cell types. Different phagocytic pathways are stimulated by particle ligation of distinct surface receptors but all forms of phagocytosis require F-actin recruitment beneath tethered particles and F-actin re-arrangement promoting engulfment, which are controlled by Rho family GTPases. The specificity of Rho GTPase activity during the different forms of phagocytosis by mammalian cells is the subject of this review.

  16. A novel oncogene, ost, encodes a guanine nucleotide exchange factor that potentially links Rho and Rac signaling pathways.

    PubMed Central

    Horii, Y; Beeler, J F; Sakaguchi, K; Tachibana, M; Miki, T

    1994-01-01

    Transfection of NIH3T3 cells with an osteosarcoma expression cDNA library led to the appearance of foci of morphologically transformed cells which were found to harbor a novel oncogene, ost. The ost product was activated by truncation of the N-terminal domain of the ost proto-oncogene and was highly tumorigenic in nude mouse assays. The proto-ost cDNA, isolated subsequently, encodes a predicted protein of 100 kDa containing DH (Db1 homology) and PH (pleckstrin homology) domains. Ost is mainly phosphorylated on serine and localized in the cytoplasm. Purified Ost protein catalyzed guanine nucleotide exchange on RhoA and Cdc42 among the Rho and Ras family members tested, indicating that Ost can activate these small GTP-binding proteins. Ost did not detectably associate with RhoA or Cdc42, but interacted specifically with the GTP-bound form of Rac1, suggesting that Ost can function as an effector of Rac1. These results suggest that Ost is a critical regulatory component which links pathways that signal through Rac1, RhoA and Cdc42. Of the tissues examined, expression of ost was the highest in brain and could be localized to neurons and alpha-tanycytes, suggesting that Ost may participate in axonal transport in these specialized cells. Images PMID:7957046

  17. 1,5-asymmetric induction of chirality using pi-allyltricarbonyliron lactone complexes: highly diastereoselective synthesis of alpha-functionalised carbonyl compounds.

    PubMed

    Hollowood, Christopher J; Ley, Steven V; Wright, Edward A

    2003-09-21

    Silyl enol ethers derived from ketone functionalised rho-allyltricarbonyliron lactone complexes undergo highly diastereoselective carbon-fluorine and carbon-oxygen bond formation reactions with excellent control at the alpha-stereogenic centre.

  18. miR-124-regulated RhoG

    PubMed Central

    Schumacher, Stefan; Franke, Kristin

    2013-01-01

    RhoG is a member of the Rho family of small GTPases sharing highest sequence similarity with Rac and Cdc42. Mig-2 and Mtl represent the functional equivalents of RhoG in Caenorhabditis elegans and Drosophila, respectively. RhoG has attracted great interest because it plays a central role in the regulation of cytoskeletal reorganization in various physiological and pathophysiological situations. For example, it is fundamental to phagocytotic processes, is able to regulate gene expression, cell survival and proliferation, and is involved in cell migration and in the invasion of pathogenic bacteria. The activation of Rac1 via an ELMO/Dock180 module has been elaborated to be important for RhoG signaling. Although a stimulatory role for neurite outgrowth in the pheochromocytoma PC12 cell line has been assigned to RhoG, the exact function of this GTPase for the development of the processes of primary neurons remains to be clarified. In this view, we discuss the impact of RhoG on axonal and dendritic differentiation, its role as a conductor of Rac1 and Cdc42 activity and the functional regulation of RhoG expression by the microRNA miR-124. PMID:23303397

  19. RhoA Controls Wnt Upregulation on Microstructured Titanium Surfaces

    PubMed Central

    Mazzotta, Silvia; Piergianni, Maddalena; Piemontese, Marilina; Passeri, Giovanni

    2014-01-01

    Rough topography enhances the activation of Wnt canonical signaling in vitro, and this mediates its effects on cell differentiation. However, the molecular mechanisms underlying topography-dependent control of Wnt signaling are still poorly understood. As the small GTPase RhoA controls cytoskeletal reorganization and actomyosin-induced tensional forces, we hypothesized that RhoA could affect the activation of Wnt signaling in cells on micropatterned titanium surfaces. G-LISA assay revealed that RhoA activation was higher in C2C12 cells on rough (SLA) surfaces under basal conditions than on smooth (Polished) titanium. Transfection with dominant negative RhoA decreased Wnt activation by normalized TCF-Luc activity on SLA, whilst transfection with constitutively active RhoA increased TCF-Luc activation on Polished titanium. One mM Myosin II inhibitor Blebbistatin increased RhoA activation but decreased Wnt activation on SLA surfaces, indicating that tension-generating structures are required for canonical Wnt modulation on titanium surfaces. Actin inhibitor Cytochalasin markedly enhanced RhoA and TCF-Luc activation on both surfaces and increased the expression of differentiation markers in murine osteoblastic MC3T3 cells. Taken together, these data show that RhoA is upregulated in cells on rough surfaces and it affects the activation of Wnt canonical signaling through Myosin II modulation. PMID:24949442

  20. Rock `N' Rho in Outer Hair Cell Motility

    NASA Astrophysics Data System (ADS)

    Zhang, M.; Kalinec, G.; Kalinec, F.; Billadeau, D. D.; Urrutia, R.

    2003-02-01

    RhoA, Cdc42 and Rac1, small GTPases of the Rho family, are crucial regulators of the actin cytoskeleton and mediate different types of cell motility. They also help to maintain cellular homeostasis, actively regulating the structure and mechanical properties of the cells. We investigated the expression in the guinea-pig cochlea of the serine/threonine kinase ROCK, a well-known effector of RhoA, and measured electromotile amplitude in outer hair cells (OHCs) internally perfused with C3 and Y-27632, pharmacological inhibitors of RhoA and ROCK respectively, and dominant-negative mutants of Rac1 and Cdc42. We found that a RhoA/ROCK-mediated signaling pathway is important for mechanical homeostasis of cochlear OHCs, and identified ROCK as a potential target to selectively modulate outer hair cell electromotility.

  1. The RhoGAP activity of CYK-4/MgcRacGAP functions non-canonically by promoting RhoA activation during cytokinesis

    PubMed Central

    Zhang, Donglei; Glotzer, Michael

    2015-01-01

    Cytokinesis requires activation of the GTPase RhoA. ECT-2, the exchange factor responsible for RhoA activation, is regulated to ensure spatiotemporal control of contractile ring assembly. Centralspindlin, composed of the Rho family GTPase-activating protein (RhoGAP) MgcRacGAP/CYK-4 and the kinesin MKLP1/ZEN-4, is known to activate ECT-2, but the underlying mechanism is not understood. We report that ECT-2-mediated RhoA activation depends on the ability of CYK-4 to localize to the plasma membrane, bind RhoA, and promote GTP hydrolysis by RhoA. Defects resulting from loss of CYK-4 RhoGAP activity can be rescued by activating mutations in ECT-2 or depletion of RGA-3/4, which functions as a conventional RhoGAP for RhoA. Consistent with CYK-4 RhoGAP activity contributing to GEF activation, the catalytic domains of CYK-4 and ECT-2 directly interact. Thus, counterintuitively, CYK-4 RhoGAP activity promotes RhoA activation. We propose that the most active form of the cytokinetic RhoGEF involves complex formation between ECT-2, centralspindlin and RhoA. DOI: http://dx.doi.org/10.7554/eLife.08898.001 PMID:26252513

  2. Observation of e(+)e(-) annihilation into the C = +1 hadronic final states rho(0)rho(0) and phirho(0).

    PubMed

    Aubert, B; Barate, R; Bona, M; Boutigny, D; Couderc, F; Karyotakis, Y; Lees, J P; Poireau, V; Tisserand, V; Zghiche, A; Grauges, E; Palano, A; Chen, J C; Qi, N D; Rong, G; Wang, P; Zhu, Y S; Eigen, G; Ofte, I; Stugu, B; Abrams, G S; Battaglia, M; Brown, D N; Button-Shafer, J; Cahn, R N; Charles, E; Gill, M S; Groysman, Y; Jacobsen, R G; Kadyk, J A; Kerth, L T; Kolomensky, Yu G; Kukartsev, G; Lynch, G; Mir, L M; Oddone, P J; Orimoto, T J; Pripstein, M; Roe, N A; Ronan, M T; Wenzel, W A; Del Amo Sanchez, P; Barrett, M; Ford, K E; Harrison, T J; Hart, A J; Hawkes, C M; Morgan, S E; Watson, A T; Goetzen, K; Held, T; Koch, H; Lewandowski, B; Pelizaeus, M; Peters, K; Schroeder, T; Steinke, M; Boyd, J T; Burke, J P; Cottingham, W N; Walker, D; Cuhadar-Donszelmann, T; Fulsom, B G; Hearty, C; Knecht, N S; Mattison, T S; McKenna, J A; Khan, A; Kyberd, P; Saleem, M; Sherwood, D J; Teodorescu, L; Blinov, V E; Bukin, A D; Druzhinin, V P; Golubev, V B; Onuchin, A P; Serednyakov, S I; Skovpen, Yu I; Solodov, E P; Todyshev, K Yu; Best, D S; Bondioli, M; Bruinsma, M; Chao, M; Curry, S; Eschrich, I; Kirkby, D; Lankford, A J; Lund, P; Mandelkern, M; Mommsen, R K; Roethel, W; Stoker, D P; Abachi, S; Buchanan, C; Foulkes, S D; Gary, J W; Long, O; Shen, B C; Wang, K; Zhang, L; Hadavand, H K; Hill, E J; Paar, H P; Rahatlou, S; Sharma, V; Berryhill, J W; Campagnari, C; Cunha, A; Dahmes, B; Hong, T M; Kovalskyi, D; Richman, J D; Beck, T W; Eisner, A M; Flacco, C J; Heusch, C A; Kroseberg, J; Lockman, W S; Nesom, G; Schalk, T; Schumm, B A; Seiden, A; Spradlin, P; Williams, D C; Wilson, M G; Albert, J; Chen, E; Dvoretskii, A; Fang, F; Hitlin, D G; Narsky, I; Piatenko, T; Porter, F C; Ryd, A; Samuel, A; Mancinelli, G; Meadows, B T; Sokoloff, M D; Blanc, F; Bloom, P C; Chen, S; Ford, W T; Hirschauer, J F; Kreisel, A; Nauenberg, U; Olivas, A; Ruddick, W O; Smith, J G; Ulmer, K A; Wagner, S R; Zhang, J; Chen, A; Eckhart, E A; Soffer, A; Toki, W H; Wilson, R J; Winklmeier, F; Zeng, Q; Altenburg, D D; Feltresi, E; Hauke, A; Jasper, H; Petzold, A; Spaan, B; Brandt, T; Klose, V; Lacker, H M; Mader, W F; Nogowski, R; Schubert, J; Schubert, K R; Schwierz, R; Sundermann, J E; Volk, A; Bernard, D; Bonneaud, G R; Grenier, P; Latour, E; Thiebaux, Ch; Verderi, M; Bard, D J; Clark, P J; Gradl, W; Muheim, F; Playfer, S; Robertson, A I; Xie, Y; Andreotti, M; Bettoni, D; Bozzi, C; Calabrese, R; Cibinetto, G; Luppi, E; Negrini, M; Petrella, A; Piemontese, L; Prencipe, E; Anulli, F; Baldini-Ferroli, R; Calcaterra, A; de Sangro, R; Finocchiaro, G; Pacetti, S; Patteri, P; Peruzzi, I M; Piccolo, M; Rama, M; Zallo, A; Buzzo, A; Capra, R; Contri, R; Lo Vetere, M; Macri, M M; Monge, M R; Passaggio, S; Patrignani, C; Robutti, E; Santroni, A; Tosi, S; Brandenburg, G; Chaisanguanthum, K S; Morii, M; Wu, J; Dubitzky, R S; Marks, J; Schenk, S; Uwer, U; Bhimji, W; Bowerman, D A; Dauncey, P D; Egede, U; Flack, R L; Nash, J A; Nikolich, M B; Panduro Vazquez, W; Chai, X; Charles, M J; Mallik, U; Meyer, N T; Ziegler, V; Cochran, J; Crawley, H B; Dong, L; Eyges, V; Meyer, W T; Prell, S; Rosenberg, E I; Rubin, A E; Gritsan, A V; Fritsch, M; Schott, G; Arnaud, N; Davier, M; Grosdidier, G; Höcker, A; Le Diberder, F; Lepeltier, V; Lutz, A M; Oyanguren, A; Pruvot, S; Rodier, S; Roudeau, P; Schune, M H; Stocchi, A; Wang, W F; Wormser, G; Cheng, C H; Lange, D J; Wright, D M; Chavez, C A; Forster, I J; Fry, J R; Gabathuler, E; Gamet, R; George, K A; Hutchcroft, D E; Payne, D J; Schofield, K C; Touramanis, C; Bevan, A J; Di Lodovico, F; Menges, W; Sacco, R; Cowan, G; Flaecher, H U; Hopkins, D A; Jackson, P S; McMahon, T R; Ricciardi, S; Salvatore, F; Wren, A C; Brown, D N; Davis, C L; Allison, J; Barlow, N R; Barlow, R J; Chia, Y M; Edgar, C L; Lafferty, G D; Naisbit, M T; Williams, J C; Yi, J I; Chen, C; Hulsbergen, W D; Jawahery, A; Lae, C K; Roberts, D A; Simi, G; Blaylock, G; Dallapiccola, C; Hertzbach, S S; Li, X; Moore, T B; Saremi, S; Staengle, H; Cowan, R; Sciolla, G; Sekula, S J; Spitznagel, M; Taylor, F; Yamamoto, R K; Kim, H; Patel, P M; Robertson, S H; Lazzaro, A; Lombardo, V; Palombo, F; Bauer, J M; Cremaldi, L; Eschenburg, V; Godang, R; Kroeger, R; Sanders, D A; Summers, D J; Zhao, H W; Brunet, S; Côté, D; Taras, P; Viaud, F B; Nicholson, H; Cavallo, N; De Nardo, G; Fabozzi, F; Gatto, C; Lista, L; Monorchio, D; Paolucci, P; Piccolo, D; Sciacca, C; Baak, M; Raven, G; Snoek, H L; Jessop, C P; Losecco, J M; Allmendinger, T; Benelli, G; Gan, K K; Honscheid, K; Hufnagel, D; Jackson, P D; Kagan, H; Kass, R; Rahimi, A M; Ter-Antonyan, R; Wong, Q K; Blount, N L; Brau, J; Frey, R; Igonkina, O; Lu, M; Potter, C T; Rahmat, R; Sinev, N B; Strom, D; Strube, J; Torrence, E; Galeazzi, F; Gaz, A; Margoni, M; Morandin, M; Pompili, A; Posocco, M; Rotondo, M; Simonetto, F; Stroili, R; Voci, C; Benayoun, M; Chauveau, J; David, P; Del Buono, L; de la Vaissière, Ch; Hamon, O; Hartfiel, B L; John, M J J; Malclès, J; Ocariz, J; Roos, L; Therin, G; Behera, P K; Gladney, L; Panetta, J; Biasini, M; Covarelli, R; Angelini, C; Batignani, G; Bettarini, S; Bucci, F; Calderini, G; Carpinelli, M; Cenci, R; Forti, F; Giorgi, M A; Lusiani, A; Marchiori, G; Mazur, M A; Morganti, M; Neri, N; Rizzo, G; Walsh, J J; Haire, M; Judd, D; Wagoner, D E; Biesiada, J; Danielson, N; Elmer, P; Lau, Y P; Lu, C; Olsen, J; Smith, A J S; Telnov, A V; Bellini, F; Cavoto, G; D'Orazio, A; Del Re, D; Di Marco, E; Faccini, R; Ferrarotto, F; Ferroni, F; Gaspero, M; Li Gioi, L; Mazzoni, M A; Morganti, S; Piredda, G; Polci, F; Safai Tehrani, F; Voena, C; Ebert, M; Schröder, H; Waldi, R; Adye, T; De Groot, N; Franek, B; Olaiya, E O; Wilson, F F; Emery, S; Gaidot, A; Ganzhur, S F; Hamel de Monchenault, G; Kozanecki, W; Legendre, M; Vasseur, G; Yèche, Ch; Zito, M; Chen, X R; Liu, H; Park, W; Purohit, M V; Wilson, J R; Allen, M T; Aston, D; Bartoldus, R; Bechtle, P; Berger, N; Claus, R; Coleman, J P; Convery, M R; Cristinziani, M; Dingfelder, J C; Dorfan, J; Dubois-Felsmann, G P; Dujmic, D; Dunwoodie, W; Field, R C; Glanzman, T; Gowdy, S J; Graham, M T; Halyo, V; Hast, C; Hryn'ova, T; Innes, W R; Kelsey, M H; Kim, P; Leith, D W G S; Li, S; Luitz, S; Luth, V; Lynch, H L; Macfarlane, D B; Marsiske, H; Messner, R; Muller, D R; O'grady, C P; Ozcan, V E; Perazzo, A; Perl, M; Pulliam, T; Ratcliff, B N; Roodman, A; Salnikov, A A; Schindler, R H; Schwiening, J; Snyder, A; Stelzer, J; Su, D; Sullivan, M K; Suzuki, K; Swain, S K; Thompson, J M; Va'vra, J; van Bakel, N; Weaver, M; Weinstein, A J R; Wisniewski, W J; Wittgen, M; Wright, D H; Yarritu, A K; Yi, K; Young, C C; Burchat, P R; Edwards, A J; Majewski, S A; Petersen, B A; Roat, C; Wilden, L; Ahmed, S; Alam, M S; Bula, R; Ernst, J A; Jain, V; Pan, B; Saeed, M A; Wappler, F R; Zain, S B; Bugg, W; Krishnamurthy, M; Spanier, S M; Eckmann, R; Ritchie, J L; Satpathy, A; Schilling, C J; Schwitters, R F; Izen, J M; Lou, X C; Ye, S; Bianchi, F; Gallo, F; Gamba, D; Bomben, M; Bosisio, L; Cartaro, C; Cossutti, F; Della Ricca, G; Dittongo, S; Lanceri, L; Vitale, L; Azzolini, V; Martinez-Vidal, F; Banerjee, Sw; Bhuyan, B; Brown, C M; Fortin, D; Hamano, K; Kowalewski, R; Nugent, I M; Roney, J M; Sobie, R J; Back, J J; Harrison, P F; Latham, T E; Mohanty, G B; Pappagallo, M; Band, H R; Chen, X; Cheng, B; Dasu, S; Datta, M; Flood, K T; Hollar, J J; Kutter, P E; Mellado, B; Mihalyi, A; Pan, Y; Pierini, M; Prepost, R; Wu, S L; Yu, Z; Neal, H

    2006-09-15

    We report the first observation of e(+)e(-) annihilation into states of positive C parity, namely, rho(0)rho(0) and phirho(0). The two states are observed in the pi(+)pi(-)pi(+)pi(-) and K(+)K(-)pi(+)pi(-) final states, respectively, in a data sample of 225 fb(-1) collected by the BABAR experiment at the Positron-Electron Project II e(+)e(-) storage rings at energies near sqrt[s]=10.58 GeV. The distributions of costheta(*), where theta(*) is the center-of-mass polar angle of the phi meson or the forward rho(0) meson, suggest production by two-virtual-photon annihilation. We measure cross sections within the range |costheta(*)|<0.8 of sigma(e(+)e(-)-->rho(0)rho(0))=20.7+/-0.7(stat)+/-2.7(syst) fb and sigma(e(+)e(-)-->phirho(0))=5.7+/-0.5(stat)+/-0.8(syst) fb.

  3. Role of Rho and Rho kinase in the activation of volume-regulated anion channels in bovine endothelial cells.

    PubMed

    Nilius, B; Voets, T; Prenen, J; Barth, H; Aktories, K; Kaibuchi, K; Droogmans, G; Eggermont, J

    1999-04-01

    1. We have studied the modulation of volume-regulated anion channels (VRACs) by the small GTPase Rho and by one of its targets, Rho kinase, in calf pulmonary artery endothelial (CPAE) cells. 2. RT-PCR and immunoblot analysis showed that both RhoA and Rho kinase are expressed in CPAE cells. 3. ICl,swell, the chloride current through VRACs, was activated by challenging CPAE cells with a 25 % hypotonic extracellular solution (HTS) or by intracellular perfusion with a pipette solution containing 100 microM GTPgammaS. 4. Pretreatment of CPAE cells with the Clostridium C2IN-C3 fusion toxin, which inactivates Rho by ADP ribosylation, significantly impaired the activation of ICl,swell in response to the HTS. The current density at +100 mV was 49 +/- 13 pA pF-1 (n = 17) in pretreated cells compared with 172 +/- 17 pA pF-1 (n = 21) in control cells. 5. The volume-independent activation of ICl,swell by intracellular perfusion with GTPgammaS was also impaired in C2IN-C3-pretreated cells (31 +/- 7 pA pF-1, n = 11) compared with non-treated cells (132 +/- 21 pA pF-1, n = 15). 6. Activation of ICl,swell was pertussis toxin (PTX) insensitive. 7. Y-27632, a blocker of Rho kinase, inhibited ICl,swell and delayed its activation. 8. Inhibition of Rho and of Rho kinase by the above-described treatments did not affect the extent of cell swelling in response to HTS. 9. These experiments provide strong evidence that the Rho-Rho kinase pathway is involved in the VRAC activation cascade.

  4. Rho GTPase–independent regulation of mitotic progression by the RhoGEF Net1

    PubMed Central

    Menon, Sarita; Oh, Wonkyung; Carr, Heather S.; Frost, Jeffrey A.

    2013-01-01

    Neuroepithelial transforming gene 1 (Net1) is a RhoA-subfamily–specific guanine nucleotide exchange factor that is overexpressed in multiple human cancers and is required for proliferation. Molecular mechanisms underlying its role in cell proliferation are unknown. Here we show that overexpression or knockdown of Net1 causes mitotic defects. Net1 is required for chromosome congression during metaphase and generation of stable kinetochore microtubule attachments. Accordingly, inhibition of Net1 expression results in spindle assembly checkpoint activation. The ability of Net1 to control mitosis is independent of RhoA or RhoB activation, as knockdown of either GTPase does not phenocopy effects of Net1 knockdown on nuclear morphology, and effects of Net1 knockdown are effectively rescued by expression of catalytically inactive Net1. We also observe that Net1 expression is required for centrosomal activation of p21-activated kinase and its downstream kinase Aurora A, which are critical regulators of centrosome maturation and spindle assembly. These results identify Net1 as a novel regulator of mitosis and indicate that altered expression of Net1, as occurs in human cancers, may adversely affect genomic stability. PMID:23864709

  5. The A-kinase anchoring protein (AKAP)-Lbc-signaling complex mediates alpha1 adrenergic receptor-induced cardiomyocyte hypertrophy.

    PubMed

    Appert-Collin, Aline; Cotecchia, Susanna; Nenniger-Tosato, Monique; Pedrazzini, Thierry; Diviani, Dario

    2007-06-12

    In response to various pathological stresses, the heart undergoes a pathological remodeling process that is associated with cardiomyocyte hypertrophy. Because cardiac hypertrophy can progress to heart failure, a major cause of lethality worldwide, the intracellular signaling pathways that control cardiomyocyte growth have been the subject of intensive investigation. It has been known for more than a decade that the small molecular weight GTPase RhoA is involved in the signaling pathways leading to cardiomyocyte hypertrophy. Although some of the hypertrophic pathways activated by RhoA have now been identified, the identity of the exchange factors that modulate its activity in cardiomyocytes is currently unknown. In this study, we show that AKAP-Lbc, an A-kinase anchoring protein (AKAP) with an intrinsic Rho-specific guanine nucleotide exchange factor activity, is critical for activating RhoA and transducing hypertrophic signals downstream of alpha1-adrenergic receptors (ARs). In particular, our results indicate that suppression of AKAP-Lbc expression by infecting rat neonatal ventricular cardiomyocytes with lentiviruses encoding AKAP-Lbc-specific short hairpin RNAs strongly reduces both alpha1-AR-mediated RhoA activation and hypertrophic responses. Interestingly, alpha1-ARs promote AKAP-Lbc activation via a pathway that requires the alpha subunit of the heterotrimeric G protein G12. These findings identify AKAP-Lbc as the first Rho-guanine nucleotide exchange factor (GEF) involved in the signaling pathways leading to cardiomyocytes hypertrophy.

  6. Measurement of the B0 Lifetime with Partial Reconstruction of overline B o right arrow D *+rho-

    NASA Astrophysics Data System (ADS)

    Soffer, A.

    2002-03-01

    A sample of about 5500 overline B0 right arrow D (sup asterisk +) rho - and 700 overline B0 right arrow D(sup asterisk +) alpha -1 events is identified, using the technique of partial reconstruction, among 22.7 million B overline B pairs collected by the BABAR experiment at the PEP-II storage ring. With these events, the B 0 lifetime is measured to be 1.616 plus or minus 0.064 plus or minus 0.075 ps. As the first time-dependent analysis conducted with partial reconstruction of overline B 0 right arrow D (sup asterisk +) rho -, this measurement serves as validation for the procedures required to measure sin(2beta + gamma) with this technique.

  7. The protective effect of Rho-associated kinase inhibitor on aluminum-induced neurotoxicity in rat cortical neurons.

    PubMed

    Chen, Tsan-Ju; Hung, Hui-Shan; Wang, Dean-Chuan; Chen, Shun-Sheng

    2010-07-01

    Aluminum (Al) is a neurotoxicant and is implicated in several neurodegenerative diseases, including Alzheimer's disease (AD). In AD brains, one of the pathological hallmarks is the extracellular deposition of senile plaques, which are mainly composed of aggregated amyloid-beta (Abeta). Endoproteolysis of the amyloid-beta precursor protein (AbetaPP) by the beta-secretase and the gamma-secretase generates Abeta. AbetaPP can also be cleaved by the alpha-secretase within the Abeta region, which releases a soluble fragment sAPPalpha and precludes the formation of Abeta. Al has been reported to increase the level of Abeta, promote Abeta aggregation, and increase Abeta neurotoxicity. In contrast, small G protein Rho and its effector, Rho-associated kinase (ROCK), are known to negatively regulate the amount of Abeta. Inhibition of the Rho-ROCK pathway may underlie the ability of nonsteroidal anti-inflammatory drugs and statins to reduce Abeta production. Whether the Rho-ROCK pathway is involved in Al-induced elevation and aggregation of Abeta is unknown. In the present study, cultured rat cortical neurons were treated with Al(malt)(3) in the absence or presence of ROCK inhibitor Y-27632. After the treatment of Al(malt)(3), the cell viability and the level of sAPPalpha were reduced, whereas the amyloid fibrils in the conditioned media were increased. Treatment with Y-27632 prevented these adverse effects of Al(malt)(3) and thus maintained neuronal survival. These results reveal that the activation of the Rho-ROCK signaling pathway was involved in Al-induced effects in terms of the cell viability, the production of sAPPalpha, and the formation of amyloid fibril, which provides a novel mechanism underlying Al-induced neurotoxicity.

  8. Regulated Localization Is Sufficient for Hormonal Control of Regulator of G Protein Signaling Homology Rho Guanine Nucleotide Exchange Factors (RH-RhoGEFs)*

    PubMed Central

    Carter, Angela M.; Gutowski, Stephen; Sternweis, Paul C.

    2014-01-01

    The regulator of G protein signaling homology (RH) Rho guanine nucleotide exchange factors (RhoGEFs) (p115RhoGEF, leukemia-associated RhoGEF, and PDZ-RhoGEF) contain an RH domain and are specific GEFs for the monomeric GTPase RhoA. The RH domains interact specifically with the α subunits of G12 heterotrimeric GTPases. Activated Gα13 modestly stimulates the exchange activity of both p115RhoGEF and leukemia-associated RhoGEF but not PDZ-RhoGEF. Because all three RH-RhoGEFs can localize to the plasma membrane upon expression of activated Gα13, cellular localization of these RhoGEFs has been proposed as a mechanism for controlling their activity. We use a small molecule-regulated heterodimerization system to rapidly control the localization of RH-RhoGEFs. Acute localization of the proteins to the plasma membrane activates RhoA within minutes and to levels that are comparable with activation of RhoA by hormonal stimulation of G protein-coupled receptors. The catalytic activity of membrane-localized RhoGEFs is not dependent on activated Gα13. We further show that the conserved RH domains can rewire two different RacGEFs to activate Rac1 in response to a traditional activator of RhoA. Thus, RH domains act as independent detectors for activated Gα13 and are sufficient to modulate the activity of RhoGEFs by hormones via mediating their localization to substrate, membrane-associated RhoA. PMID:24855647

  9. eIF5A1/RhoGDIα pathway: a novel therapeutic target for treatment of spinal cord injury identified by a proteomics approach

    PubMed Central

    Liu, Wei; Shang, Fei-Fei; Xu, Yang; Belegu, Visar; Xia, Lei; Zhao, Wei; Liu, Ran; Wang, Wei; Liu, Jin; Li, Chen-Yun; Wang, Ting-Hua

    2015-01-01

    Spinal cord injury (SCI) is frequently accompanied by a degree of spontaneous functional recovery. The underlying mechanisms through which such recovery is generated remain elusive. In this study, we observed a significant spontaneous motor function recovery 14 to 28 days after spinal cord transection (SCT) in rats. Using a comparative proteomics approach, caudal to the injury, we detected difference in 20 proteins. Two of these proteins, are eukaryotic translation initiation factor 5A1 (eIF5A1) that is involved in cell survival and proliferation, and Rho GDP dissociation inhibitor alpha (RhoGDIα), a member of Rho GDI family that is involved in cytoskeletal reorganization. After confirming the changes in expression levels of these two proteins following SCT, we showed that in vivo eIF5A1 up-regulation and down-regulation significantly increased and decreased, respectively, motor function recovery. In vitro, eIF5A1 overexpression in primary neurons increased cell survival and elongated neurite length while eIF5A1 knockdown reversed these results. We found that RhoGDIα up-regulation and down-regulation rescues the effect of eIF5A1 down-regulation and up-regulation both in vivo and in vitro. Therefore, we have identified eIF5A1/RhoGDIα pathway as a new therapeutic target for treatment of spinal cord injured patients. PMID:26593060

  10. Tension on JAM-A activates RhoA via GEF-H1 and p115 RhoGEF.

    PubMed

    Scott, David W; Tolbert, Caitlin E; Burridge, Keith

    2016-05-01

    Junctional adhesion molecule A (JAM-A) is a broadly expressed adhesion molecule that regulates cell-cell contacts and facilitates leukocyte transendothelial migration. The latter occurs through interactions with the integrin LFA-1. Although we understand much about JAM-A, little is known regarding the protein's role in mechanotransduction or as a modulator of RhoA signaling. We found that tension imposed on JAM-A activates RhoA, which leads to increased cell stiffness. Activation of RhoA in this system depends on PI3K-mediated activation of GEF-H1 and p115 RhoGEF. These two GEFs are further regulated by FAK/ERK and Src family kinases, respectively. Finally, we show that phosphorylation of JAM-A at Ser-284 is required for RhoA activation in response to tension. These data demonstrate a direct role of JAM-A in mechanosignaling and control of RhoA and implicate Src family kinases in the regulation of p115 RhoGEF.

  11. Tension on JAM-A activates RhoA via GEF-H1 and p115 RhoGEF

    PubMed Central

    Scott, David W.; Tolbert, Caitlin E.; Burridge, Keith

    2016-01-01

    Junctional adhesion molecule A (JAM-A) is a broadly expressed adhesion molecule that regulates cell–cell contacts and facilitates leukocyte transendothelial migration. The latter occurs through interactions with the integrin LFA-1. Although we understand much about JAM-A, little is known regarding the protein’s role in mechanotransduction or as a modulator of RhoA signaling. We found that tension imposed on JAM-A activates RhoA, which leads to increased cell stiffness. Activation of RhoA in this system depends on PI3K-mediated activation of GEF-H1 and p115 RhoGEF. These two GEFs are further regulated by FAK/ERK and Src family kinases, respectively. Finally, we show that phosphorylation of JAM-A at Ser-284 is required for RhoA activation in response to tension. These data demonstrate a direct role of JAM-A in mechanosignaling and control of RhoA and implicate Src family kinases in the regulation of p115 RhoGEF. PMID:26985018

  12. Measurement of branching fractions and charge asymmetries in B+/--->rho+/-pi0 and B+/--->rho0pi+/- decays, and search for B0-->rho0pi0.

    PubMed

    Aubert, B; Barate, R; Boutigny, D; Couderc, F; Gaillard, J-M; Hicheur, A; Karyotakis, Y; Lees, J P; Robbe, P; Tisserand, V; Zghiche, A; Palano, A; Pompili, A; Chen, J C; Qi, N D; Rong, G; Wang, P; Zhu, Y S; Eigen, G; Ofte, I; Stugu, B; Abrams, G S; Borgland, A W; Breon, A B; Brown, D N; Button-Shafer, J; Cahn, R N; Charles, E; Day, C T; Gill, M S; Gritsan, A V; Groysman, Y; Jacobsen, R G; Kadel, R W; Kadyk, J; Kerth, L T; Kolomensky, Yu G; Kukartsev, G; LeClerc, C; Levi, M E; Lynch, G; Mir, L M; Oddone, P J; Orimoto, T J; Pripstein, M; Roe, N A; Romosan, A; Ronan, M T; Shelkov, V G; Telnov, A V; Wenzel, W A; Ford, K; Harrison, T J; Hawkes, C M; Knowles, D J; Morgan, S E; Penny, R C; Watson, A T; Watson, N K; Goetzen, K; Held, T; Koch, H; Lewandowski, B; Pelizaeus, M; Peters, K; Schmuecker, H; Steinke, M; Boyd, J T; Chevalier, N; Cottingham, W N; Kelly, M P; Latham, T E; Mackay, C; Wilson, F F; Abe, K; Cuhadar-Donszelmann, T; Hearty, C; Mattison, T S; McKenna, J A; Thiessen, D; Kyberd, P; McKemey, A K; Teodorescu, L; Blinov, V E; Bukin, A D; Golubev, V B; Ivanchenko, V N; Kravchenko, E A; Onuchin, A P; Serednyakov, S I; Skovpen, Yu I; Solodov, E P; Yushkov, A N; Best, D; Bruinsma, M; Chao, M; Kirkby, D; Lankford, A J; Mandelkern, M; Mommsen, R K; Roethel, W; Stoker, D P; Buchanan, C; Hartfiel, B L; Gary, J W; Layter, J; Shen, B C; Wang, K; del Re, D; Hadavand, H K; Hill, E J; MacFarlane, D B; Paar, H P; Rahatlou, Sh; Sharma, V; Berryhill, J W; Campagnari, C; Dahmes, B; Levy, S L; Long, O; Lu, A; Mazur, M A; Richman, J D; Verkerke, W; Beck, T W; Beringer, J; Eisner, A M; Heusch, C A; Lockman, W S; Schalk, T; Schmitz, R E; Schumm, B A; Seiden, A; Spradlin, P; Turri, M; Walkowiak, W; Williams, D C; Wilson, M G; Albert, J; Chen, E; Dubois-Felsmann, G P; Dvoretskii, A; Erwin, R J; Hitlin, D G; Narsky, I; Piatenko, T; Porter, F C; Ryd, A; Samuel, A; Yang, S; Jayatilleke, S; Mancinelli, G; Meadows, B T; Sokoloff, M D; Abe, T; Blanc, F; Bloom, P; Chen, S; Clark, P J; Ford, W T; Nauenberg, U; Olivas, A; Rankin, P; Roy, J; Smith, J G; van Hoek, W C; Zhang, L; Harton, J L; Hu, T; Soffer, A; Toki, W H; Wilson, R J; Zhang, J; Altenburg, D; Brandt, T; Brose, J; Colberg, T; Dickopp, M; Dubitzky, R S; Hauke, A; Lacker, H M; Maly, E; Müller-Pfefferkorn, R; Nogowski, R; Otto, S; Schubert, J; Schubert, K R; Schwierz, R; Spaan, B; Wilden, L; Bernard, D; Bonneaud, G R; Brochard, F; Cohen-Tanugi, J; Grenier, P; Thiebaux, Ch; Vasileiadis, G; Verderi, M; Khan, A; Lavin, D; Muheim, F; Playfer, S; Swain, J E; Andreotti, M; Azzolini, V; Bettoni, D; Bozzi, C; Calabrese, R; Cibinetto, G; Luppi, E; Negrini, M; Piemontese, L; Sarti, A; Treadwell, E; Baldini-Ferroli, R; Calcaterra, A; de Sangro, R; Falciai, D; Finocchiaro, G; Patteri, P; Piccolo, M; Zallo, A; Buzzo, A; Capra, R; Contri, R; Crosetti, G; Lo Vetere, M; Macri, M; Monge, M R; Passaggio, S; Patrignani, C; Robutti, E; Santroni, A; Tosi, S; Bailey, S; Morii, M; Won, E; Bhimji, W; Bowerman, D A; Dauncey, P D; Egede, U; Eschrich, I; Gaillard, J R; Morton, G W; Nash, J A; Taylor, G P; Grenier, G J; Lee, S-J; Mallik, U; Cochran, J; Crawley, H B; Lamsa, J; Meyer, W T; Prell, S; Rosenberg, E I; Yi, J; Davier, M; Grosdidier, G; Höcker, A; Laplace, S; Le Diberder, F; Lepeltier, V; Lutz, A M; Petersen, T C; Plaszczynski, S; Schune, M H; Tantot, L; Wormser, G; Brigljević, V; Cheng, C H; Lange, D J; Simani, M C; Wright, D M; Bevan, A J; Coleman, J P; Fry, J R; Gabathuler, E; Gamet, R; Kay, M; Parry, R J; Payne, D J; Sloane, R J; Touramanis, C; Back, J J; Harrison, P F; Shorthouse, H W; Vidal, P B; Brown, C L; Cowan, G; Flack, R L; Flaecher, H U; George, S; Green, M G; Kurup, A; Marker, C E; McMahon, T R; Ricciardi, S; Salvatore, F; Vaitsas, G; Winter, M A; Brown, D; Davis, C L; Allison, J; Barlow, N R; Barlow, R J; Hart, P A; Hodgkinson, M C; Jackson, F; Lafferty, G D; Lyon, A J; Weatherall, J H; Williams, J C; Farbin, A; Jawahery, A; Kovalskyi, D; Lae, C K; Lillard, V; Roberts, D A; Blaylock, G; Dallapiccola, C; Flood, K T; Hertzbach, S S; Kofler, R; Koptchev, V B; Moore, T B; Saremi, S; Staengle, H; Willocq, S; Cowan, R; Sciolla, G; Taylor, F; Yamamoto, R K; Mangeol, D J J; Patel, P M; Robertson, S H; Lazzaro, A; Palombo, F; Bauer, J M; Cremaldi, L; Eschenburg, V; Godang, R; Kroeger, R; Reidy, J; Sanders, D A; Summers, D J; Zhao, H W; Brunet, S; Cote-Ahern, D; Taras, P; Nicholson, H; Cartaro, C; Cavallo, N; De Nardo, G; Fabozzi, F; Gatto, C; Lista, L; Paolucci, P; Piccolo, D; Sciacca, C; Baak, M A; Raven, G; LoSecco, J M; Gabriel, T A; Brau, B; Gan, K K; Honscheid, K; Hufnagel, D; Kagan, H; Kass, R; Pulliam, T; Wong, Q K; Brau, J; Frey, R; Igonkina, O; Potter, C T; Sinev, N B; Strom, D; Torrence, E; Colecchia, F; Dorigo, A; Galeazzi, F; Margoni, M; Morandin, M; Posocco, M; Rotondo, M; Simonetto, F; Stroili, R; Tiozzo, G; Voci, C; Benayoun, M; Briand, H; Chauveau, J; David, P; de la Vaissière, Ch; Del Buono, L; Hamon, O; John, M J J; Leruste, Ph; Ocariz, J; Pivk, M; Roos, L; Stark, J; T'Jampens, S; Therin, G; Manfredi, P F; Re, V; Behera, P K; Gladney, L; Guo, Q H; Panetta, J; Anulli, F; Biasini, M; Peruzzi, I M; Pioppi, M; Angelini, C; Batignani, G; Bettarini, S; Bondioli, M; Bucci, F; Calderini, G; Carpinelli, M; Del Gamba, V; Forti, F; Giorgi, M A; Lusiani, A; Marchiori, G; Martinez-Vidal, F; Morganti, M; Neri, N; Paoloni, E; Rama, M; Rizzo, G; Sandrelli, F; Walsh, J; Haire, M; Judd, D; Paick, K; Wagoner, D E; Danielson, N; Elmer, P; Lu, C; Miftakov, V; Olsen, J; Smith, A J S; Tanaka, H A; Varnes, E W; Bellini, F; Cavoto, G; Faccini, R; Ferrarotto, F; Ferroni, F; Gaspero, M; Mazzoni, M A; Morganti, S; Pierini, M; Piredda, G; Safai Tehrani, F; Voena, C; Christ, S; Wagner, G; Waldi, R; Adye, T; De Groot, N; Franek, B; Geddes, N I; Gopal, G P; Olaiya, E O; Xella, S M; Aleksan, R; Emery, S; Gaidot, A; Ganzhur, S F; Giraud, P-F; Hamel de Monchenault, G; Kozanecki, W; Langer, M; Legendre, M; London, G W; Mayer, B; Schott, G; Vasseur, G; Yeche, Ch; Zito, M; Purohit, M V; Weidemann, A W; Yumiceva, F X; Aston, D; Bartoldus, R; Berger, N; Boyarski, A M; Buchmueller, O L; Convery, M R; Cristinziani, M; Dong, D; Dorfan, J; Dujmic, D; Dunwoodie, W; Elsen, E E; Field, R C; Glanzman, T; Gowdy, S J; Grauges-Pous, E; Hadig, T; Halyo, V; Hast, C; Hryn'ova, T; Innes, W R; Jessop, C P; Kelsey, M H; Kim, P; Kocian, M L; Langenegger, U; Leith, D W G S; Libby, J; Luitz, S; Luth, V; Lynch, H L; Marsiske, H; Messner, R; Muller, D R; O'Grady, C P; Ozcan, V E; Perazzo, A; Perl, M; Petrak, S; Ratcliff, B N; Roodman, A; Salnikov, A A; Schindler, R H; Schwiening, J; Simi, G; Snyder, A; Soha, A; Stelzer, J; Su, D; Sullivan, M K; Va'vra, J; Wagner, S R; Weaver, M; Weinstein, A J R; Wisniewski, W J; Wright, D H; Young, C C; Burchat, P R; Edwards, A J; Meyer, T I; Petersen, B A; Roat, C; Ahmed, M; Ahmed, S; Alam, M S; Ernst, J A; Saeed, M A; Saleem, M; Wappler, F R; Bugg, W; Krishnamurthy, M; Spanier, S M; Eckmann, R; Kim, H; Ritchie, J L; Schwitters, R F; Izen, J M; Kitayama, I; Lou, X C; Ye, S; Bianchi, F; Bona, M; Gallo, F; Gamba, D; Borean, C; Bosisio, L; Della Ricca, G; Dittongo, S; Grancagnolo, S; Lanceri, L; Poropat, P; Vitale, L; Vuagnin, G; Panvini, R S; Banerjee, Sw; Brown, C M; Fortin, D; Jackson, P D; Kowalewski, R; Roney, J M; Band, H R; Dasu, S; Datta, M; Eichenbaum, A M; Johnson, J R; Kutter, P E; Li, H; Liu, R; Di Lodovico, F; Mihalyi, A; Mohapatra, A K; Pan, Y; Prepost, R; Sekula, S J; von Wimmersperg-Toeller, J H; Wu, J; Wu, S L; Yu, Z; Neal, H

    2004-07-30

    We present measurements of branching fractions and charge asymmetries in B-meson decays to rho(+)pi(0), rho(0)pi(+), and rho(0)pi(0). The data sample comprises 89x10(6) Upsilon(4S)-->BBmacr; decays collected with the BABAR detector at the PEP-II asymmetric-energy B Factory at SLAC. We find the charge-averaged branching fractions B(B+-->rho(+)pi(0))=[10.9+/-1.9(stat)+/-1.9(syst)]x10(-6) and B(B+-->rho(0)pi(+))=(9.5+/-1.1+/-0.9)x10(-6), and we set a 90% confidence-level upper limit B(B0-->rho(0)pi(0))<2.9x10(-6). We measure the charge asymmetries ACP(pi(0))(rho(+))=0.24+/-0.16+/-0.06 and ACP(pi(+))(rho(0))=-0.19+/-0.11+/-0.02.

  13. Diacylglycerol kinase ζ regulates RhoA activation via a kinase-independent scaffolding mechanism.

    PubMed

    Ard, Ryan; Mulatz, Kirk; Abramovici, Hanan; Maillet, Jean-Christian; Fottinger, Alexandra; Foley, Tanya; Byham, Michèle-Renée; Iqbal, Tasfia Ahmed; Yoneda, Atsuko; Couchman, John R; Parks, Robin J; Gee, Stephen H

    2012-10-01

    Rho GTPases share a common inhibitor, Rho guanine nucleotide dissociation inhibitor (RhoGDI), which regulates their expression levels, membrane localization, and activation state. The selective dissociation of individual Rho GTPases from RhoGDI ensures appropriate responses to cellular signals, but the underlying mechanisms are unclear. Diacylglycerol kinase ζ (DGKζ), which phosphorylates diacylglycerol to yield phosphatidic acid, selectively dissociates Rac1 by stimulating PAK1-mediated phosphorylation of RhoGDI on Ser-101/174. Similarly, phosphorylation of RhoGDI on Ser-34 by protein kinase Cα (PKCα) selectively releases RhoA. Here we show DGKζ is required for RhoA activation and Ser-34 phosphorylation, which were decreased in DGKζ-deficient fibroblasts and rescued by wild-type DGKζ or a catalytically inactive mutant. DGKζ bound directly to the C-terminus of RhoA and the regulatory arm of RhoGDI and was required for efficient interaction of PKCα and RhoA. DGKζ-null fibroblasts had condensed F-actin bundles and altered focal adhesion distribution, indicative of aberrant RhoA signaling. Two targets of the RhoA effector ROCK showed reduced phosphorylation in DGKζ-null cells. Collectively our findings suggest DGKζ functions as a scaffold to assemble a signaling complex that functions as a RhoA-selective, GDI dissociation factor. As a regulator of Rac1 and RhoA activity, DGKζ is a critical factor linking changes in lipid signaling to actin reorganization.

  14. Constraining |V(td)|/|V(ts)| Using Radiative Penguin B -> V(K*/rho/omega)gamma Decays

    SciTech Connect

    Tan, Ping; /Wisconsin U., Madison

    2006-03-08

    Exclusive radiative penguin B decays, B {yields} (K*{sup 0}/K*{sup +}) and B {yields} ({rho}/{omega}){gamma}, are flavor-changing neutral-current (FCNC) processes. Studies of these decays are of special interest in testing Standard Model (SM) predictions and searching for other beyond-the-SM FCNC interactions. Using 89 x 10{sup 6} B{bar B} pairs from BABAR, we measure the branching fraction ({Beta}), CP-asymmetry ({Alpha}), and isospin asymmetry ({Delta}{sub 0-}) of B {yields} (K*{sup 0}/K*{sup +}){gamma} as follows: {Beta}(B{sup 0} {yields} K*{sup 0}{gamma}) = 3.92 {+-} 0.20(stat.) {+-} 0.24(syst.); {Beta}(B{sup +} {yields} K*{sup +}{gamma}) = 3.87 {+-} 0.28(stat.) {+-} 0.26(syst.); {Alpha}(B {yields} K*{gamma}) = -0.013 {+-} 0.36(stat.) {+-} 0.10(syst.); {Delta}{sub 0-}(B {yields} K*{gamma}) = 0.050 {+-} 0.045(stat.) {+-} 0.028(syst.) {+-} 0.024(R{sup +/0}). The 90% confidence intervals for the CP-asymmetry and the isospin-asymmetry in the B {yields} K*{gamma} decay are given as: -0.074 < {Alpha}(B {yields} K*{gamma}) < 0.049, -0.046 < {Delta}{sub 0-} (B {yields} K*{gamma}) < 0.146. We also search for B {yields} ({rho}/{omega}){gamma} decays using 211 x 10{sup 6} B{bar B} pairs from BABAR. No evidence for these decays is found. We set the upper limits at 90% confidence level for these decays: {Beta}(B{sup 0} {yields} {rho}{sup 0}{gamma}) < 0.4 x 10{sup -6}; {Beta}(B{sup +}{yields} {rho}{sup =}{gamma}) < 1.8 x 10{sup -6}; {Beta}(B{sup 0} {yields} {omega}{gamma}) < 1.0 x 10{sup -6}; {bar {Beta}}(B {yields} ({rho}/{omega}){gamma}) < 1.2 x 10{sup -6}. These results are in good agreement with the SM predictions. The branching fractions of these decays are then used to constrain the ratio |V{sub td}|/|V{sub ts}|.

  15. The Rho Family Member RhoE Interacts with Skp2 and Is Degraded at the Proteasome during Cell Cycle Progression*

    PubMed Central

    Lonjedo, Marta; Poch, Enric; Mocholí, Enric; Hernández-Sánchez, Marta; Ivorra, Carmen; Franke, Thomas F.; Guasch, Rosa M.; Pérez-Roger, Ignacio

    2013-01-01

    RhoE/Rnd3 is an atypical member of the Rho family of small GTPases. In addition to regulating actin cytoskeleton dynamics, RhoE is involved in the regulation of cell proliferation, survival, and metastasis. We examined RhoE expression levels during cell cycle and investigated mechanisms controlling them. We show that RhoE accumulates during G1, in contact-inhibited cells, and when the Akt pathway is inhibited. Conversely, RhoE levels rapidly decrease at the G1/S transition and remain low for most of the cell cycle. We also show that the half-life of RhoE is shorter than that of other Rho proteins and that its expression levels are regulated by proteasomal degradation. The expression patterns of RhoE overlap with that of the cell cycle inhibitor p27. Consistently with an involvement of RhoE in cell cycle regulation, RhoE and p27 levels decrease after overexpression of the F-box protein Skp2. We have identified a region between amino acids 231 and 240 of RhoE as the Skp2-interacting domain and Lys235 as the substrate for ubiquitylation. Based on our results, we propose a mechanism according to which proteasomal degradation of RhoE by Skp2 regulates its protein levels to control cellular proliferation. PMID:24045951

  16. Search for medium modification of the $\\rho$ meson

    SciTech Connect

    R. Nasseripour; M. H. Wood; C. Djalali; D. P. Weygand; C. Tur; U. Mosel; P. Muehlich; CLAS Collaboration

    2007-08-01

    The photoproduction of vector mesons on various nuclei has been studied using the CEBAF Large Acceptance Spectrometer (CLAS) at Jefferson Laboratory. The vector mesons, $\\rho$, $\\omega$, and $\\phi$, are observed via their decay to $e^+e^-$, in order to reduce the effects of final state interactions in the nucleus. Of particular interest are possible in-medium effects on the properties of the $\\rho$ meson. The $\\rho$ spectral function is extracted from the data on various nuclei, carbon, iron, and titanium, and compared to the spectrum from liquid deuterium, which is relatively free of nuclear effects. We observe no significant mass shift for the $\\rho$ meson; however, there is some widening of the resonance in titanium and iron, which is consistent with expected collisional broadening.

  17. Rho family-associated kinases PAK1 and rock.

    PubMed

    Maruta, Hiroshi; Nheu, Thao V; He, Hong; Hirokawa, Yumiko

    2003-01-01

    Rho family GTPases (Rho, Rac and CDC42) share around 30% sequence identity with RAS family GTPases, and are essential for RAS-induced malignant transformation, i.e., aberrant serum/anchorage-independent growth and actin cytoskeleton-linked morphological changes. Oncogenic RAS mutants such as v-Ha-RAS trigger cell cycle entry (G0-G1 transition) mainly by up-regulating cyclin D1, an activator of cyclin-dependent kinases (CDK), and down-regulating p27, a CDK inhibitor. Although both Rac and CDC42 are clearly activated by RAS, there is so far no evidence that RAS activates Rho. In this chapter, we will discuss the role of these Rho family GTPases and their effectors, in particular the Ser/Thr kinases PAK1 and Rock, in RAS-induced serum/anchorage-independent cell cycling, and discuss several potential therapeutics, peptides or chemical compounds, that could block this oncogenic cell cycle signalling pathway.

  18. Underexpression of RhoH in Hairy Cell Leukemia.

    PubMed

    Galiègue-Zouitina, Sylvie; Delestré, Laure; Dupont, Caroline; Troussard, Xavier; Shelley, Carl Simon

    2008-06-15

    The cause of hairy cell leukemia (HCL) is unknown. Current treatments seem effective only for a limited period of time. In addition, a significant proportion of patients remain refractive to all treatment options. These considerations indicate the need to develop alternative therapeutic strategies for HCL. Here, we report that HCL is characterized by underexpression of RhoH. In vitro reconstitution of RhoH expression inhibits the aberrant adhesion and transendothelial migration that drives disease pathogenesis. In an in vivo model of HCL, RhoH reconstitution limits malignant progression and protects against mortality. These findings provide the proof of principle that RhoH reconstitution represents a potential new approach to the treatment of HCL.

  19. Alpha Blockers

    MedlinePlus

    ... conditions such as high blood pressure and benign prostatic hyperplasia. Find out more about this class of medication. ... these conditions: High blood pressure Enlarged prostate (benign prostatic hyperplasia) Though alpha blockers are commonly used to treat ...

  20. Alpha fetoprotein

    MedlinePlus

    ... Alpha fetoprotein - series References Cunningham FG, Leveno KJ, Bloom SL, et al. Prenatal diagnosis and fetal therapy. In: Cunningham FG, Leveno KJ, Bloom SL, et al, eds. Williams Obstetrics . 23rd ed. ...

  1. Alpha Thalassemia

    MedlinePlus

    ... an apparently normal individual has a child with hemoglobin H disease or alpha thalassemia minor. It can ... gene on one chromosome 25% 25% 25% 25% hemoglobin H disease there is a 25% chance with ...

  2. Control of T lymphocyte morphology by the GTPase Rho

    NASA Technical Reports Server (NTRS)

    Woodside, Darren G.; Wooten, David K.; Teague, T. Kent; Miyamoto, Yuko J.; Caudell, Eva G.; Udagawa, Taturo; Andruss, Bernard F.; McIntyre, Bradley W.

    2003-01-01

    BACKGROUND: Rho family GTPase regulation of the actin cytoskeleton governs a variety of cell responses. In this report, we have analyzed the role of the GTPase Rho in maintenance of the T lymphocyte actin cytoskeleton. RESULTS: Inactivation of the GTPase Rho in the human T lymphocytic cell line HPB-ALL does not inhibit constitutively high adhesion to the integrin beta1 substrate fibronectin. It did however result in the aberrant extension of finger-like dendritic processes on the substrates VCAM-1, Fn, and mAb specific to beta1 integrins. Time-lapse video microscopy demonstrated that C3 induced extensions were primarily the result of an altered pseudopod elongation rather than retraction. Once the stellate pseudopodia extended, none retracted, and cells became completely immobile. Filipodial structures were absent and the dendritic-like processes in C3 treated cells were rich in filamentous actin. Immunolocalization of RhoA in untreated HPB-ALL cells spreading on fibronectin demonstrated a diffuse staining pattern within the pseudopodia. In C3 treated cells, clusters of RhoA were pronounced and localized within the altered extensions. CONCLUSIONS: GTPase Rho is actively involved in the regulation of T lymphocyte morphology and motility.

  3. Local translation of RhoA regulates growth cone collapse

    PubMed Central

    Cox, Llewellyn J.; Macosko, Evan Z.; Jeromin, Andreas; Urquhart, Erica R.; Jaffrey, Samie R.

    2005-01-01

    Neuronal development requires highly coordinated regulation of the cytoskeleton within the developing axon. This dynamic regulation manifests itself in axonal branching, turning, and pathfinding, presynaptic differentiation, and growth cone collapse and extension. Semaphorin 3A (Sema3A), a secreted guidance cue that primarily acts to repel axons from inappropriate targets, induces cytoskeletal rearrangements that results in growth cone collapse 1. These effects require intra-axonal mRNA translation. Here we show that transcripts for RhoA, a small GTPase that regulates the actin cytoskeleton, are localized to developing axons and growth cones, and this localization is mediated by an axonal targeting element located in the RhoA 3’UTR. Sema3A induces intra-axonal translation of RhoA mRNA and this local translation of RhoA is necessary and sufficient for Sema3A-mediated growth cone collapse. These studies indicate that local RhoA translation regulates the neuronal cytoskeleton and identify a novel mechanism for the regulation of RhoA signaling. PMID:16107849

  4. Search for radiative penguin decays B(+)-->rho(+)gamma, B(0)-->rho(0)gamma, and B(0)-->omegagamma.

    PubMed

    Aubert, B; Barate, R; Boutigny, D; Couderc, F; Gaillard, J-M; Hicheur, A; Karyotakis, Y; Lees, J P; Tisserand, V; Zghiche, A; Palano, A; Pompili, A; Chen, J C; Qi, N D; Rong, G; Wang, P; Zhu, Y S; Eigen, G; Ofte, I; Stugu, B; Abrams, G S; Borgland, A W; Breon, A B; Brown, D N; Button-Shafer, J; Cahn, R N; Charles, E; Day, C T; Gill, M S; Gritsan, A V; Groysman, Y; Jacobsen, R G; Kadel, R W; Kadyk, J; Kerth, L T; Kolomensky, Yu G; Kukartsev, G; Lynch, G; Mir, L M; Oddone, P J; Orimoto, T J; Pripstein, M; Roe, N A; Ronan, M T; Shelkov, V G; Wenzel, W A; Barrett, M; Ford, K E; Harrison, T J; Hart, A J; Hawkes, C M; Morgan, S E; Watson, A T; Fritsch, M; Goetzen, K; Held, T; Koch, H; Lewandowski, B; Pelizaeus, M; Steinke, M; Boyd, J T; Chevalier, N; Cottingham, W N; Kelly, M P; Latham, T E; Wilson, F F; Cuhadar-Donszelmann, T; Hearty, C; Knecht, N S; Mattison, T S; McKenna, J A; Thiessen, D; Khan, A; Kyberd, P; Teodorescu, L; Blinov, A E; Blinov, V E; Druzhinin, V P; Golubev, V B; Ivanchenko, V N; Kravchenko, E A; Onuchin, A P; Serednyakov, S I; Skovpen, Yu I; Solodov, E P; Yushkov, A N; Best, D; Bruinsma, M; Chao, M; Eschrich, I; Kirkby, D; Lankford, A J; Mandelkern, M; Mommsen, R K; Roethel, W; Stoker, D P; Buchanan, C; Hartfiel, B L; Foulkes, S D; Gary, J W; Shen, B C; Wang, K; Del Re, D; Hadavand, H K; Hill, E J; Macfarlane, D B; Paar, H P; Rahatlou, Sh; Sharma, V; Berryhill, J W; Campagnari, C; Dahmes, B; Long, O; Lu, A; Mazur, M A; Richman, J D; Verkerke, W; Beck, T W; Eisner, A M; Heusch, C A; Kroseberg, J; Lockman, W S; Nesom, G; Schalk, T; Schumm, B A; Seiden, A; Spradlin, P; Williams, D C; Wilson, M G; Albert, J; Chen, E; Dubois-Felsmann, G P; Dvoretskii, A; Hitlin, D G; Narsky, I; Piatenko, T; Porter, F C; Ryd, A; Samuel, A; Yang, S; Jayatilleke, S; Mancinelli, G; Meadows, B T; Sokoloff, M D; Abe, T; Blanc, F; Bloom, P; Chen, S; Ford, W T; Nauenberg, U; Olivas, A; Rankin, P; Smith, J G; Zhang, J; Zhang, L; Chen, A; Harton, J L; Soffer, A; Toki, W H; Wilson, R J; Zeng, Q L; Altenburg, D; Brandt, T; Brose, J; Dickopp, M; Feltresi, E; Hauke, A; Lacker, H M; Müller-Pfefferkorn, R; Nogowski, R; Otto, S; Petzold, A; Schubert, J; Schubert, K R; Schwierz, R; Spaan, B; Sundermann, J E; Bernard, D; Bonneaud, G R; Brochard, F; Grenier, P; Schrenk, S; Thiebaux, Ch; Vasileiadis, G; Verderi, M; Bard, D J; Clark, P J; Lavin, D; Muheim, F; Playfer, S; Xie, Y; Andreotti, M; Azzolini, V; Bettoni, D; Bozzi, C; Calabrese, R; Cibinetto, G; Luppi, E; Negrini, M; Piemontese, L; Sarti, A; Treadwell, E; Anulli, F; Baldini-Ferroli, R; Calcaterra, A; de Sangro, R; Finocchiaro, G; Patteri, P; Peruzzi, I M; Piccolo, M; Zallo, A; Buzzo, A; Capra, R; Contri, R; Crosetti, G; Vetere, M Lo; Macri, M; Monge, M R; Passaggio, S; Patrignani, C; Robutti, E; Santroni, A; Tosi, S; Bailey, S; Brandenburg, G; Chaisanguanthum, K S; Morii, M; Won, E; Dubitzky, R S; Langenegger, U; Bhimji, W; Bowerman, D A; Dauncey, P D; Egede, U; Gaillard, J R; Morton, G W; Nash, J A; Nikolich, M B; Taylor, G P; Charles, M J; Grenier, G J; Mallik, U; Cochran, J; Crawley, H B; Lamsa, J; Meyer, W T; Prell, S; Rosenberg, E I; Rubin, A E; Yi, J; Biasini, M; Covarelli, R; Pioppi, M; Davier, M; Giroux, X; Grosdidier, G; Höcker, A; Laplace, S; Diberder, F Le; Lepeltier, V; Lutz, A M; Petersen, T C; Plaszczynski, S; Schune, M H; Tantot, L; Wormser, G; Cheng, C H; Lange, D J; Simani, M C; Wright, D M; Bevan, A J; Chavez, C A; Coleman, J P; Forster, I J; Fry, J R; Gabathuler, E; Gamet, R; Hutchcroft, D E; Parry, R J; Payne, D J; Sloane, R J; Touramanis, C; Back, J J; Cormack, C M; Harrison, P F; Lodovico, F Di; Mohanty, G B; Brown, C L; Cowan, G; Flack, R L; Flaecher, H U; Green, M G; Jackson, P S; McMahon, T R; Ricciardi, S; Salvatore, F; Winter, M A; Brown, D; Davis, C L; Allison, J; Barlow, N R; Barlow, R J; Hart, P A; Hodgkinson, M C; Lafferty, G D; Lyon, A J; Williams, J C; Farbin, A; Hulsbergen, W D; Jawahery, A; Kovalskyi, D; Lae, C K; Lillard, V; Roberts, D A; Blaylock, G; Dallapiccola, C; Flood, K T; Hertzbach, S S; Kofler, R; Koptchev, V B; Moore, T B; Saremi, S; Staengle, H; Willocq, S; Cowan, R; Sciolla, G; Sekula, S J; Taylor, F; Yamamoto, R K; Mangeol, D J J; Patel, P M; Robertson, S H; Lazzaro, A; Lombardo, V; Palombo, F; Bauer, J M; Cremaldi, L; Eschenburg, V; Godang, R; Kroeger, R; Reidy, J; Sanders, D A; Summers, D J; Zhao, H W; Brunet, S; Côté, D; Taras, P; Nicholson, H; Cavallo, N; Fabozzi, F; Gatto, C; Lista, L; Monorchio, D; Paolucci, P; Piccolo, D; Sciacca, C; Baak, M; Bulten, H; Raven, G; Snoek, H L; Wilden, L; Jessop, C P; Losecco, J M; Allmendinger, T; Gan, K K; Honscheid, K; Hufnagel, D; Kagan, H; Kass, R; Pulliam, T; Rahimi, A M; Ter-Antonyan, R; Wong, Q K; Brau, J; Frey, R; Igonkina, O; Potter, C T; Sinev, N B; Strom, D; Torrence, E; Colecchia, F; Dorigo, A; Galeazzi, F; Margoni, M; Morandin, M; Posocco, M; Rotondo, M; Simonetto, F; Stroili, R; Tiozzo, G; Voci, C; Benayoun, M; Briand, H; Chauveau, J; David, P; de la Vaissière, Ch; Buono, L Del; Hamon, O; John, M J J; Leruste, Ph; Malcles, J; Ocariz, J; Pivk, M; Roos, L; T'jampens, S; Therin, G; Manfredi, P F; Re, V; Behera, P K; Gladney, L; Guo, Q H; Panetta, J; Angelini, C; Batignani, G; Bettarini, S; Bondioli, M; Bucci, F; Calderini, G; Carpinelli, M; Forti, F; Giorgi, M A; Lusiani, A; Marchiori, G; Martinez-Vidal, F; Morganti, M; Neri, N; Paoloni, E; Rama, M; Rizzo, G; Sandrelli, F; Walsh, J; Haire, M; Judd, D; Paick, K; Wagoner, D E; Danielson, N; Elmer, P; Lau, Y P; Lu, C; Miftakov, V; Olsen, J; Smith, A J S; Telnov, A V; Bellini, F; Cavoto, G; Faccini, R; Ferrarotto, F; Ferroni, F; Gaspero, M; Gioi, L Li; Mazzoni, M A; Morganti, S; Pierini, M; Piredda, G; Tehrani, F Safai; Voena, C; Christ, S; Wagner, G; Waldi, R; Adye, T; Groot, N De; Franek, B; Geddes, N I; Gopal, G P; Olaiya, E O; Aleksan, R; Emery, S; Gaidot, A; Ganzhur, S F; Giraud, P-F; de Monchenault, G Hamel; Kozanecki, W; Legendre, M; London, G W; Mayer, B; Schott, G; Vasseur, G; Yèche, Ch; Zito, M; Purohit, M V; Weidemann, A W; Wilson, J R; Yumiceva, F X; Aston, D; Bartoldus, R; Berger, N; Boyarski, A M; Buchmueller, O L; Claus, R; Convery, M R; Cristinziani, M; Nardo, G De; Dong, D; Dorfan, J; Dujmic, D; Dunwoodie, W; Elsen, E E; Fan, S; Field, R C; Glanzman, T; Gowdy, S J; Hadig, T; Halyo, V; Hast, C; Hryn'ova, T; Innes, W R; Kelsey, M H; Kim, P; Kocian, M L; Leith, D W G S; Libby, J; Luitz, S; Luth, V; Lynch, H L; Marsiske, H; Messner, R; Muller, D R; O'grady, C P; Ozcan, V E; Perazzo, A; Perl, M; Petrak, S; Ratcliff, B N; Roodman, A; Salnikov, A A; Schindler, R H; Schwiening, J; Simi, G; Snyder, A; Soha, A; Stelzer, J; Su, D; Sullivan, M K; Va'vra, J; Wagner, S R; Weaver, M; Weinstein, A J R; Wisniewski, W J; Wittgen, M; Wright, D H; Yarritu, A K; Young, C C; Burchat, P R; Edwards, A J; Meyer, T I; Petersen, B A; Roat, C; Ahmed, S; Alam, M S; Ernst, J A; Saeed, M A; Saleem, M; Wappler, F R; Bugg, W; Krishnamurthy, M; Spanier, S M; Eckmann, R; Kim, H; Ritchie, J L; Satpathy, A; Schwitters, R F; Izen, J M; Kitayama, I; Lou, X C; Ye, S; Bianchi, F; Bona, M; Gallo, F; Gamba, D; Bosisio, L; Cartaro, C; Cossutti, F; Ricca, G Della; Dittongo, S; Grancagnolo, S; Lanceri, L; Poropat, P; Vitale, L; Vuagnin, G; Panvini, R S; Banerjee, Sw; Brown, C M; Fortin, D; Jackson, P D; Kowalewski, R; Roney, J M; Sobie, R J; Band, H R; Cheng, B; Dasu, S; Datta, M; Eichenbaum, A M; Graham, M; Hollar, J J; Johnson, J R; Kutter, P E; Li, H; Liu, R; Mihalyi, A; Mohapatra, A K; Pan, Y; Prepost, R; Tan, P; von Wimmersperg-Toeller, J H; Wu, J; Wu, S L; Yu, Z; Greene, M G; Neal, H

    2005-01-14

    A search for the decays B-->rho(770)gamma and B0-->omega(782)gamma is performed on a sample of 211 x 10(6) Upsilon(4S)-->BB events collected by the BABAR detector at the SLAC PEP-II asymmetric-energy e(+)e(-) storage ring. No evidence for the decays is seen. We set the following limits on the individual branching fractions: B(B+-->rho(+)gamma)<1.8 x 10(-6), B(B0-->rho(0)gamma)<0.4 x 10(-6), and B(B0-->omegagamma)<1.0 x 10(-6) at the 90% confidence level. We use the quark model to limit the combined branching fraction B [B-->(rho/omega)gamma]<1.2 x 10(-6), from which we determine a constraint on the ratio of Cabibbo-Kobayashi-Maskawa matrix elements |V(td)|/|V(ts)|.

  5. Ginseng (Panax quinquefolius) attenuates leptin-induced cardiac hypertrophy through inhibition of p115Rho guanine nucleotide exchange factor-RhoA/Rho-associated, coiled-coil containing protein kinase-dependent mitogen-activated protein kinase pathway activation.

    PubMed

    Moey, Melissa; Rajapurohitam, Venkatesh; Zeidan, Asad; Karmazyn, Morris

    2011-12-01

    Leptin is a 16-kDa peptide primarily derived from white adipocytes and is typically elevated in plasma of obese individuals. Although leptin plays a critical role in appetite regulation, leptin receptors have been identified in numerous tissues including the heart and have been shown to directly mediate cardiac hypertrophy through RhoA/ROCK (Ras homolog gene family, member A/Rho-associated, coiled-coil containing protein kinase)-dependent p38 mitogen-activated protein kinase (MAPK) activation; however, the basis for RhoA stimulation is unknown. Rho guanine nucleotide exchange factors (GEFs) catalyze the exchange of GDP for GTP resulting in Rho activation and may be the potential upstream factors mediating leptin-induced RhoA activation and therefore a potential target for inhibition. We investigated the effects of North American ginseng (Panax quinquefolius), reported to reduce cardiac hypertrophy, on RhoA/ROCK and MAPK activation in ventricular cardiomyocytes exposed to leptin (50 ng/ml) and the possible role of p115RhoGEF and p63RhoGEF in these responses. Leptin produced a robust hypertrophic response that was associated with RhoA/ROCK activation resulting in a significant increase in cofilin-2 phosphorylation and actin polymerization, the latter evidenced by a reduction in the G/F actin ratio. These effects were prevented by ginseng (10 μg/ml). The stimulation of RhoA/ROCK by leptin was associated with significantly increased p115RhoGEF gene and protein expression and exchange activity, all of which were completely prevented by ginseng. The ability of ginseng to prevent leptin-induced activation of RhoA/ROCK was further associated with diminished p38 MAPK activation and nuclear translocation. These results demonstrate a potent inhibitory effect of ginseng against leptin-induced cardiac hypertrophy, an effect associated with prevention of p115RhoGEF-RhoA/ROCK-dependent p38 MAPK activation.

  6. Characterization of NH/sub 4/-rho and vacuum-calcined H-rho zeolites by multinuclear NMR spectroscopy

    SciTech Connect

    Vega, A.J.; Luz, Z.

    1987-01-15

    Proton spin counting NMR is used to follow the vacuum calcination of the zeolite NH/sub 4/-rho. Based on these results a two-step vacuum-calcination procedure for the preparation of H-rho is proposed. It consists of heating for 20 h at 80 /sup 0/C followed by 7 h at 450 /sup 0/C. A multinuclear NMR study is then performed in order to characterize the anhydrous and hydrated forms of NH/sub 4/- and H-rho. The techniques include /sup 27/Al, /sup 29/Si, and /sup 1/H magic-angle spinning (MAS), /sup 1/H and /sup 27/Al spin counting, quadrupole-echo deuterium NMR of deuterated samples, as well as /sup 1/H-/sup 29/Si cross polarization. Hydrated NH/sub 4/-rho is characterized by high mobility of the NH/sub 4//sup +/ and H/sub 2/O species and by high framework symmetry and uniformity. In anhydrous NH/sub 4/-rho the NH/sub 4//sup +/ ions are localized at the Al sites, thus slightly distorting the tetrahedral Al symmetry and introducing dispersion between the Si sites. The NH/sub 4//sup +/ ions are distorted from tetrahedral symmetry and rotate rapidly. Anhydrous H-rho has approx.11 hydrogen atoms/uc in Bronsted acid sites, some rigid, some undergoing fast large amplitude librations, but no translational diffusion between different sites is observed. A small fraction of the hydrogens are in terminal hydroxyl groups. All Al sites are severely distorted so that the /sup 27/Al signal is unobservable. Hydration of H-rho restores the framework symmetry and the proton mobility. The /sup 29/Si chemical shifts indicate pronounced structural changes between the various forms of zeolite rho.

  7. Measurements of Branching Fractions for B+ -> rho+ gamma, B0 -> rho0 gamma, and B0 -> omega gamma

    SciTech Connect

    Aubert, B

    2008-08-15

    The authors present branching fraction measurements for the radiative decays B{sup +} {yields} {rho}{sup +}{gamma}, B{sup 0} {yields} {rho}{sup 0}{gamma}, and B{sup 0} {yields} {omega}{gamma}. The analysis is based on a data sample of 465 million B{bar B} events collected with the BABAR detector at the PEP-II asymmetric-energy B Factory located at the Stanford Linear Accelerator Center (SLAC). They find {Beta}(B{sup +} {yields} {rho}{sup +}{gamma}) = (1.20{sub -0.37}{sup +0.42} {+-} 0.20) x 10{sup -6}, {Beta}(B{sup 0} {yields} {rho}{sup 0}{gamma}) = (0.97{sub -0.22}{sup +0.24} {+-} 0.06) x 10{sup -6}, and a 90% C.L. upper limit {Beta}(B{sup 0} {yields} {omega}{gamma}) < 0.9 x 10{sup -6}, where the first error is statistical and the second is systematic. They also measure the isospin-violating quantity {Lambda}(B{sup +} {yields} {rho}{sup +}{gamma})/2{Lambda}(B{sup 0} {yields} {rho}{sup 0}{gamma}) - 1 = -0.43{sub -0.22}{sup +0.25} {+-} 0.10.

  8. Activating Transcription Factor 4 (ATF4) modulates Rho GTPase levels and function via regulation of RhoGDIα

    PubMed Central

    Pasini, Silvia; Liu, Jin; Corona, Carlo; Peze-Heidsieck, Eugenie; Shelanski, Michael; Greene, Lloyd A.

    2016-01-01

    In earlier studies, we showed that ATF4 down-regulation affects post-synaptic development and dendritic spine morphology in neurons through increased turnover of the Rho GTPase Cell Division Cycle 42 (Cdc42) protein. Here, we find that ATF4 down-regulation in both hippocampal and cortical neuron cultures reduces protein and message levels of RhoGDIα, a stabilizer of the Rho GTPases including Cdc42. This effect is rescued by an shATF4-resistant active form of ATF4, but not by a mutant that lacks transcriptional activity. This is, at least in part, due to the fact that Arhgdia, the gene encoding RhoGDIα, is a direct transcriptional target of ATF4 as is shown in ChIP assays. This pathway is not restricted to neurons. This is seen in an impairment of cell migration on ATF4 reduction in non-neuronal cells. In conclusion, we have identified a new cellular pathway in which ATF4 regulates the expression of RhoGDIα that in turn affects Rho GTPase protein levels, and thereby, controls cellular functions as diverse as memory and cell motility. PMID:27841340

  9. Activated RhoA Binds to the Pleckstrin Homology (PH) Domain of PDZ-RhoGEF, a Potential Site for Autoregulation

    SciTech Connect

    Chen, Zhe; Medina, Frank; Liu, Mu-ya; Thomas, Celestine; Sprang, Stephen R.; Sternweis, Paul C.

    2010-07-19

    Guanine nucleotide exchange factors (GEFs) catalyze exchange of GDP for GTP by stabilizing the nucleotide-free state of the small GTPases through their Dbl homology/pleckstrin homology (DH {center_dot} PH) domains. Unconventionally, PDZ-RhoGEF (PRG), a member of the RGS-RhoGEFs, binds tightly to both nucleotide-free and activated RhoA (RhoA {center_dot} GTP). We have characterized the interaction between PRG and activated RhoA and determined the structure of the PRG-DH {center_dot} PH-RhoA {center_dot} GTP{gamma}S (guanosine 5{prime}-O-[{gamma}-thio]triphosphate) complex. The interface bears striking similarity to a GTPase-effector interface and involves the switch regions in RhoA and a hydrophobic patch in PRG-PH that is conserved among all Lbc RhoGEFs. The two surfaces that bind activated and nucleotide-free RhoA on PRG-DH {center_dot} PH do not overlap, and a ternary complex of PRG-DH {center_dot} PH bound to both forms of RhoA can be isolated by size-exclusion chromatography. This novel interaction between activated RhoA and PH could play a key role in regulation of RhoGEF activity in vivo.

  10. Genetic Analysis of the Saccharomyces Cerevisiae Rho3 Gene, Encoding a Rho-Type Small Gtpase, Provides Evidence for a Role in Bud Formation

    PubMed Central

    Imai, J.; Toh-e, A.; Matsui, Y.

    1996-01-01

    RHO3 encodes a Rho-type small GTPase of the yeast Saccharomyces cerevisiae. We isolated temperature-sensitive alleles and a dominant active allele of RHO3. Ts(-) rho3 cells lost cell polarity during bud formation and grew more isotropically than wild-type cells at nonpermissive temperatures. In contrast, cells carrying a dominant active mutant RHO3 displayed cold sensitivity, and the cells became elongated and bent, often at the position where actin patches were concentrated. These phenotypes of the rho3 mutants strongly suggest that RHO3 is involved in directing the growing points during bud formation. In addition, we found that SRO6, previously isolated as a multicopy suppressor of rho3, is the same as SEC4. The sec4-2 mutation was synthetic lethal with temperature-sensitive rho3 mutations and suppressed the cold sensitivity caused by a dominant active mutant RHO3. The genetic interactions between RHO3 and SEC4, taken together with the fact that the Rab-type GTPase Sec4p is required to fuse secretory vesicles together with plasma membrane for exocytosis, support a model in which the Rho3p pathway modulates morphogenesis during bud growth via directing organization of the actin cytoskeleton and the position of the secretory machinery for exocytosis. PMID:8852836

  11. Foam-stabilizing effects and cling formation patterns of iso-alpha-acids and reduced iso-alpha-acids in lager beer.

    PubMed

    Kunimune, Takeshi; Shellhammer, Thomas H

    2008-09-24

    Foam-stabilizing properties and cling formation patterns of iso-alpha-acids and reduced iso-alpha-acids were investigated using an unhopped lager beer. Unhopped beer was dosed with iso-alpha-acid (Iso), rho-iso-alpha-acid (Rho), tetrahydro-iso-alpha-acid (Tetra), and hexahydro-iso-alpha-acid (Hexa), separately, over a range of concentrations from 2 to 10 ppm. A uniform foam was created by Inpack 2000 Flasher Head and was measured by a NIBEM Foam Stability Tester (NIBEM-TPH) followed by a NIBEM Cling Meter (NIBEM-CLM) to determine the relationship between the concentration and NIBEM-30 and the cling formation ability of each compound. The foam-stabilizing power was determined to be Tetra, Hexa, Iso, and Rho from the strongest to weakest. Linear regression models were created using the NIBEM-TPH data set, and on the basis of 95% confidence intervals, the foam stability of Tetra or Hexa became significantly larger than that of Iso when 2.4 or 4.2 ppm of Tetra or Hexa was used as a replacement for Iso, respectively. Cling formation patterns could be categorized into three groups: "ring", "mesh", and "powdery". The control beer had the lowest foam stability and did not yield any foam cling.

  12. RhoA GTPase oxidation stimulates cell proliferation via nuclear factor-κB activation.

    PubMed

    Kim, Jae-Gyu; Kwon, Hyung-Joo; Wu, Guang; Park, Yohan; Lee, Jae-Yong; Kim, Jaebong; Kim, Sung-Chan; Choe, Myoen; Kang, Seung Goo; Seo, Goo-Young; Kim, Pyeung-Hyeun; Park, Jae-Bong

    2017-02-01

    Reactive oxygen species (ROS) produced by many kinds of stimuli are essential for cellular signaling including cell proliferation. The dysregulation of ROS, therefore, is related to a variety of diseases including cancer. However, it was not clearly elucidated how ROS regulate cell proliferation and tumorigenesis. In this study, we investigated a mechanism by which the oxidation of RhoA GTPase regulates nuclear factor-κB (NF-κB) and cell proliferation. Hydrogen peroxide activated NF-κB and RhoA GTPase, but did not activate RhoA C16/20A mutant, an oxidation-resistant form. Remarkably, the oxidation of RhoA reduced its affinity towards RhoGDI, leading to the dissociation of RhoA-RhoGDI complex. Si-Vav2, a guanine nucleotide exchange factor (GEF), inhibited RhoA activation upon hydrogen peroxide. The oxidized RhoA (oxRhoA)-GTP was readily bound to IκB kinase γ (IKKγ), whereas oxidized RhoGDI did not bind to IKKγ. The oxRhoA-GTP bound to IKKγ activated IKKβ, leading to IκB phosphorylation and degradation, consequently NF-κB activation. Hydrogen peroxide induced cell proliferation, but RhoA C16/20A mutant suppressed cell proliferation and tumorigenesis. Conclusively, RhoA oxidation at Cys16/20 is critically involved in cell proliferation and tumorigenesis through NF-κB activation in response to ROS.

  13. The rho-guanine nucleotide exchange factor domain of obscurin regulates assembly of titin at the Z-disk through interactions with Ran binding protein 9.

    PubMed

    Bowman, Amber L; Catino, Dawn H; Strong, John C; Randall, William R; Kontrogianni-Konstantopoulos, Aikaterini; Bloch, Robert J

    2008-09-01

    Obscurin is an approximately 800-kDa protein composed of structural and signaling domains that organizes contractile structures in striated muscle. We have studied the Rho-GEF domain of obscurin to understand its roles in morphogenesis and signaling. We used adenoviral overexpression of this domain, together with ultrastructural and immunofluorescence methods, to examine its effect on maturing myofibrils. We report that overexpression of the Rho-GEF domain specifically inhibits the incorporation of titin into developing Z-disks and disrupts the structure of the Z-disk and Z/I junction, and alters features of the A/I junction. The organization of other sarcomeric markers, including alpha-actinin, was not affected. We identified Ran binding protein 9 (RanBP9) as a novel ligand of the Rho-GEF domain and showed that binding is specific, with an apparent binding affinity of 1.9 microM. Overexpression of the binding region of RanBP9 also disrupted the incorporation of titin into developing Z-disks. Immunofluorescence localization during myofibrillogenesis indicated that the Rho-GEF domain assembles into sarcomeres before RanBP9, which first occurs in myonuclei and later in development translocates to the myoplasm, where it colocalizes with obscurin. Both the Rho-GEF domain and its binding region on RanBP9 bind directly to the N-terminal Ig domains of titin, which flank the Z-disk. Our results suggest that the Rho-GEF domain interacts with RanBP9 and that both can interact with the N-terminal region of titin to influence the formation of the Z-disk and A/I junction.

  14. Unique Structural and Nucleotide Exchange Features of the Rho1 GTPase of Entamoeba histolytica

    SciTech Connect

    Bosch, Dustin E.; Wittchen, Erika S.; Qiu, Connie; Burridge, Keith; Siderovski, David P.

    2012-08-10

    The single-celled human parasite Entamoeba histolytica possesses a dynamic actin cytoskeleton vital for its intestinal and systemic pathogenicity. The E. histolytica genome encodes several Rho family GTPases known to regulate cytoskeletal dynamics. EhRho1, the first family member identified, was reported to be insensitive to the Rho GTPase-specific Clostridium botulinum C3 exoenzyme, raising the possibility that it may be a misclassified Ras family member. Here, we report the crystal structures of EhRho1 in both active and inactive states. EhRho1 is activated by a conserved switch mechanism, but diverges from mammalian Rho GTPases in lacking a signature Rho insert helix. EhRho1 engages a homolog of mDia, EhFormin1, suggesting a role in mediating serum-stimulated actin reorganization and microtubule formation during mitosis. EhRho1, but not a constitutively active mutant, interacts with a newly identified EhRhoGDI in a prenylation-dependent manner. Furthermore, constitutively active EhRho1 induces actin stress fiber formation in mammalian fibroblasts, thereby identifying it as a functional Rho family GTPase. EhRho1 exhibits a fast rate of nucleotide exchange relative to mammalian Rho GTPases due to a distinctive switch one isoleucine residue reminiscent of the constitutively active F28L mutation in human Cdc42, which for the latter protein, is sufficient for cellular transformation. Nonconserved, nucleotide-interacting residues within EhRho1, revealed by the crystal structure models, were observed to contribute a moderating influence on fast spontaneous nucleotide exchange. Collectively, these observations indicate that EhRho1 is a bona fide member of the Rho GTPase family, albeit with unique structural and functional aspects compared with mammalian Rho GTPases.

  15. Phosphorylation and Activation of RhoA by ERK in Response to Epidermal Growth Factor Stimulation

    PubMed Central

    Tong, Junfeng; Li, Laiji; Ballermann, Barbara; Wang, Zhixiang

    2016-01-01

    The small GTPase RhoA has been implicated in various cellular activities, including the formation of stress fibers, cell motility, and cytokinesis. In addition to the canonical GTPase cycle, recent findings have suggested that phosphorylation further contributes to the tight regulation of Rho GTPases. Indeed, RhoA is phosphorylated on serine 188 (188S) by a number of protein kinases. We have recently reported that Rac1 is phosphorylated on threonine 108 (108T) by extracellular signal-regulated kinases (ERK) in response to epidermal growth factor (EGF) stimulation. Here, we provide evidence that RhoA is phosphorylated by ERK on 88S and 100T in response to EGF stimulation. We show that ERK interacts with RhoA and that this interaction is dependent on the ERK docking site (D-site) at the C-terminus of RhoA. EGF stimulation enhanced the activation of the endogenous RhoA. The phosphomimetic mutant, GFP-RhoA S88E/T100E, when transiently expressed in COS-7 cells, displayed higher GTP-binding than wild type RhoA. Moreover, the expression of GFP-RhoA S88E/T100E increased actin stress fiber formation in COS-7 cells, which is consistent with its higher activity. In contrast to Rac1, phosphorylation of RhoA by ERK does not target RhoA to the nucleus. Finally, we show that regardless of the phosphorylation status of RhoA and Rac1, substitution of the RhoA PBR with the Rac1 PBR targets RhoA to the nucleus and substitution of Rac1 PBR with RhoA PBR significantly reduces the nuclear localization of Rac1. In conclusion, ERK phosphorylates RhoA on 88S and 100T in response to EGF, which upregulates RhoA activity. PMID:26816343

  16. Rho-kinase in sea urchin eggs and embryos.

    PubMed

    Aguirre-Armenta, Beatriz; López-Godínez, Juana; Martínez-Cadena, Guadalupe; García-Soto, Jesús

    2011-06-01

    The activation of sea urchin eggs at fertilization provides an ideal system for studying the molecular events involved in cellular activation. Rho GTPases, which are key signaling enzymes in eukaryotes, are involved in sustaining the activation of sea urchin eggs; however, their downstream effectors have not yet been characterized. In somatic cells, RhoA regulates a serine/threonine kinase known as Rho-kinase (ROCK). The activity of ROCK in early sea urchin development has been inferred, but not tested directly. A ROCK gene was identified in the sea urchin (Strongylocentrotus purpuratus) genome and the sequence of its cDNA determined. The sea urchin ROCK (SpROCK) sequence predicts a protein of 158 kDa with >72% and 45% identities with different protein orthologues of the kinase catalytic domain and the complete protein sequence, respectively. SpROCK mRNA levels are high in unfertilized eggs and decrease to 35% after 15 min postfertilization and remain low up to the 4 cell stage. Antibodies to the human ROCK-I kinase domain revealed SpROCK to be concentrated in the cortex of eggs and early embryos. Co-immunoprecipitation assays indicate that RhoA and SpROCK are physically associated. This association is destroyed by treatment with the C3 exoenzyme and with the ROCK antagonist H-1152. H-1152 also inhibited DNA synthesis in embryos. We conclude that the Rho-dependent signaling pathway, via SpROCK, is essential for early embryonic development.

  17. Measurements of Charmless B Decays Related to alpha at BaBar

    SciTech Connect

    Lombardo, Vincenzo; /INFN, Milan

    2009-12-09

    We report recent measurements of the CKM angle {alpha} using data collected by the BABAR detector at the PEP-II asymmetric-energy e{sup +}e{sup -} collider at the SLAC National Accelerator Laboratory. In addition to improved constraints on {alpha} from the decays B{sup {+-}} {yields} {rho}{sup {+-}}{rho}{sup 0}, we also present preliminary results of neutral and charged B meson decays to K{sub 1}(1270){pi} and K{sub 1}(1400){pi} and its impact on the estimate for the CKM angle {alpha} based on time-dependent analysis of CP-violating asymmetries in B{sup 0} {yields} a{sub 1}(1260){sup {+-}} {pi}{sup {-+}}. Moreover we report the first observation of the decay B {yields} a{sub 1}(1260){sup {+-}}a{sub 1}(1260){sup {-+}}; this mode can be used, in principle, to provide an independent measurement of {alpha}.

  18. Leucine zipper-mediated homo-oligomerization regulates the Rho-GEF activity of AKAP-Lbc.

    PubMed

    Baisamy, Laurent; Jurisch, Nathalie; Diviani, Dario

    2005-04-15

    AKAP-Lbc is a novel member of the A-kinase anchoring protein (AKAPs) family, which functions as a cAMP-dependent protein kinase (PKA)-targeting protein as well as a guanine nucleotide exchange factor (GEF) for RhoA. We recently demonstrated that AKAP-Lbc Rho-GEF activity is stimulated by the alpha-subunit of the heterotrimeric G protein G(12), whereas phosphorylation of AKAP-Lbc by the anchored PKA induces the recruitment of 14-3-3, which inhibits its GEF function. In the present report, using co-immunoprecipitation approaches, we demonstrated that AKAP-Lbc can form homo-oligomers inside cells. Mutagenesis studies revealed that oligomerization is mediated by two adjacent leucine zipper motifs located in the C-terminal region of the anchoring protein. Most interestingly, disruption of oligomerization resulted in a drastic increase in the ability of AKAP-Lbc to stimulate the formation of Rho-GTP in cells under basal conditions, suggesting that oligomerization maintains AKAP-Lbc in a basal-inactive state. Based on these results and on our previous findings showing that AKAP-Lbc is inactivated through the association with 14-3-3, we investigated the hypothesis that AKAP-Lbc oligomerization might be required for the regulatory action of 14-3-3. Most interestingly, we found that mutants of AKAP-Lbc impaired in their ability to undergo oligomerization were completely resistant to the inhibitory effect of PKA and 14-3-3. This suggests that 14-3-3 can negatively regulate the Rho-GEF activity of AKAP-Lbc only when the anchoring protein is in an oligomeric state. Altogether, these findings provide a novel mechanistic explanation of how oligomerization can regulate the activity of exchange factors of the Dbl family.

  19. The Oncogenic Role of RhoGAPs in Basal-Like Breast Cancer

    DTIC Science & Technology

    2015-02-01

    was also found to control cell spreading and migration . RacGAP1 depletion significantly retarded BLBC growth but, in contrast to ArhGAP11A, this was...ArhGAP11A, RacGAP1, basal-like breast cancer, RhoGAPs, Rho GTPases, RhoA, Rac1, Cdc42, proliferation, migration 16. SECURITY CLASSIFICATION OF: 17...like breast cancer, RhoGAPs, Rho GTPases, RhoA, Rac1, Cdc42, proliferation, migration 3. ACCOMPLISHMENTS: What were the major goals of the project? The

  20. Isoprenylation of RhoB is necessary for its degradation. A novel determinant in the complex regulation of RhoB expression by the mevalonate pathway.

    PubMed

    Stamatakis, Konstantinos; Cernuda-Morollón, Eva; Hernández-Perera, Octavio; Pérez-Sala, Dolores

    2002-12-20

    Statins improve vascular functions by mechanisms independent from their cholesterol-lowering effect. Rho GTPases are emerging as key targets for the vascular effects of statins. RhoB is a short-lived, early-response inducible protein involved in receptor endocytosis, apoptosis, and gene expression. Here we show that statins regulate RhoB expression by acting at multiple levels. Simvastatin increased RhoB protein levels by 8- to 10-fold. This effect was related to a depletion of isoprenoid intermediates, as deduced from the observation that several metabolites of the cholesterol biosynthetic pathway, namely, mevalonate and geranylgeranyl-pyrophosphate, attenuated simvastatin-induced RhoB up-regulation. Moreover, prenyltransferase inhibitors mimicked simvastatin effect. Cholesterol supplementation did not prevent simvastatin-elicited up-regulation but increased RhoB levels per se. Simvastatin moderately augmented RhoB transcript levels, but markedly impaired the degradation of RhoB protein, which accumulated in the cytosol in its non-isoprenylated form. Inhibition of RhoB isoprenylation was apparently required for simvastatin-induced up-regulation, because levels of an isoprenylation-deficient RhoB mutant were not affected by simvastatin. Moreover, this mutant was found to be markedly more stable than the wild-type protein. These results show that RhoB isoprenylation is necessary for rapid turnover of this protein and identify a novel link between the cholesterol biosynthetic pathway and the regulation of G-protein expression.

  1. Mixed antagonistic effects of bilobalide at rho1 GABAC receptor.

    PubMed

    Huang, S H; Duke, R K; Chebib, M; Sasaki, K; Wada, K; Johnston, G A R

    2006-01-01

    Bilobalide was found to be a moderately potent antagonist with a weak use-dependent effect at recombinant human rho(1) GABA(C) receptors expressed in Xenopus oocytes using two-electrode voltage clamp methodology. Antagonism of bilobalide at homomeric rho(1) GABA(C) receptors appeared to be mixed. At low concentration, bilobalide (3 microM) caused a parallel right shift and surmountable GABA maximal response of the GABA dose-response curve characteristic of a competitive antagonist. At high concentrations, bilobalide (10-100 microM) caused nonparallel right shifts and reduced maximal GABA responses of GABA dose-response curves characteristic of a noncompetitive antagonist. The potency of bilobalide appears to be dependent on the concentrations of GABA and was more potent at lower GABA concentrations. The mechanism of action of bilobalide at rho(1) GABA(C) receptors appears to be similar to that of the chloride channel blocker picrotoxinin.

  2. Coherent rho 0 photoproduction in bulk matter at high energies

    SciTech Connect

    Couderc, Elsa; Klein, Spencer

    2009-01-09

    The momentum transfer {Delta}k required for a photon to scatter from a target and emerge as a {rho}{sup 0} decreases as the photon energy k rises. For k > 3 x 10{sup 14} eV, {Delta}k is small enough that the interaction cannot be localized to a single nucleus. At still higher energies, photons may coherently scatter elastically from bulk matter and emerge as a {rho}{sup 0}, in a manner akin to kaon regeneration. Constructive interference from the different nuclei coherently raises the cross section and the interaction probability rises linearly with energy. At energies above 10{sup 23} eV, coherent conversion is the dominant process; photons interact predominantly as {rho}{sup 0}. We compute the coherent scattering probabilities in slabs of lead, water and rock, and discuss the implications of the increased hadronic interaction probabilities for photons on ultra-high energy shower development.

  3. Signaling through rho gtpases in microgravity (rho signaling) on iss (soyuz tma-1) belgian soyuz mission "odissea"

    NASA Astrophysics Data System (ADS)

    Nusgens, B.; Lambert, Ch.; Lapière, C. M.

    2007-09-01

    Rho GTPases, RhoA, Rac1 and Cdc42, are molecular switches in the intracellular signaling pathways, that relay the information collected by receptors to soluble mediators and insoluble extracellular matrix environment. The objective of this experiment was to investigate the impact of microgravity on cellular processes depending on Rho GTPases activity, i.e.cytoskeleton and focal adhesions organization, GTPases translocation to the membrane, nuclear translocation of signaling molecules and gene expression. WI26 fibroblasts were stably transfected by the constitutively active form of each of the Rho GTPase. Selected clone of the engineered cells and the wild-type cells were used during the belgian ODISSEA Soyuz mission to investigate the alterations of the mechanical and phenotypic expression of the cells induced by microgravity and their rescue by the engineered Rho GTPases. A failure in the time schedule, a disconnection of the experiment containers before the automatic activation of the fixation procedure, was responsible for the loss of the biological samples.

  4. Impacts of Rho kinase inhibitor Fasudil on Rho/ROCK signaling pathway in rabbits with optic nerve injury

    PubMed Central

    Yu, Jianglong; Lin, Lin; Luan, Xinping; Jing, Xiepan; Maierab

    2015-01-01

    Objective: The aim of this study was to study the impacts of Rho kinase inhibitor Fasudil on expressions of Rho/ROCK signaling pathway associated genes in rabbits with optic nerve injury (ONI), and to explore the therapeutic mechanisms towards ONI. Methods: The rabbit ONI model was established, then the rabbits were divided into model group (treated with saline), control group (treated with dexamethasone, Dex), and intervention group (treated with Fasudil, Fas). The eyeball and optic nerve were sampled at 3, 7, 14 and 21 days after injury. The morphological changes of retina and optic nerve were observed. The expressions of RhoA, Caspase-3, Rock 2 and Nogo-A gene were determined by immunohistochemistry and real-time polymerase chain reaction (RT-PCR) methods. Results: At different time after injury, there were significant differences of RhoA, Caspase-3, Rock 2 and Nogo-A gene expression among three groups (P < 0.05). Conclusions: After ONI, Fas can decrease the expression of Caspase-3 gene, and down-regulate the expressions of Nogo-A and Rock 2 gene. Therefore, it can treat ONI through affecting the Rho/ROCK signaling pathway. PMID:26823796

  5. RhoA Proteolysis Regulates the Actin Cytoskeleton in Response to Oxidative Stress

    PubMed Central

    Girouard, Marie-Pier; Pool, Madeline; Alchini, Ricardo; Rambaldi, Isabel

    2016-01-01

    The small GTPase RhoA regulates the actin cytoskeleton to affect multiple cellular processes including endocytosis, migration and adhesion. RhoA activity is tightly regulated through several mechanisms including GDP/GTP cycling, phosphorylation, glycosylation and prenylation. Previous reports have also reported that cleavage of the carboxy-terminus inactivates RhoA. Here, we describe a novel mechanism of RhoA proteolysis that generates a stable amino-terminal RhoA fragment (RhoA-NTF). RhoA-NTF is detectable in healthy cells and tissues and is upregulated following cell stress. Overexpression of either RhoA-NTF or the carboxy-terminal RhoA cleavage fragment (RhoA-CTF) induces the formation of disorganized actin stress fibres. RhoA-CTF also promotes the formation of disorganized actin stress fibres and nuclear actin rods. Both fragments disrupt the organization of actin stress fibres formed by endogenous RhoA. Together, our findings describe a novel RhoA regulatory mechanism. PMID:27992599

  6. The Role of RhoJ in Endothelial Cell Biology and Tumor Pathology

    PubMed Central

    Shi, Ting-Ting; Li, Gang

    2016-01-01

    Background. RhoJ, an endothelially expressed member of Cdc42 (cell division cycle 42) subfamily of Rho GTPase, plays an important role in endocytic pathway, adipocyte differentiation, endothelial motility, tube formation, and focal adhesion. RhoJ is a selective and effective therapeutic target in tumor tissues or retinopathy. Methods. A systematic review was related to “small Rho GTPase” or “RhoJ” with “endothelial motility, tube formation and focal adhesion” and “tumor therapy”. This led to many cross-references involving RhoJ and these data have been incorporated into the following study. Results. We have grouped the role of RhoJ according to three main effects: RhoJ regulates endocytic pathway and adipocyte differentiation in early studies, and RhoJ shows an important role in endothelial cell biology; furthermore, RhoJ blockade serves as a target in tumor vasculature and enhances the effects of anticancer drug. Conclusions. More research is necessary to understand the role of RhoJ in many aspects, on the basis of current knowledge of the role of RhoJ blockade in tumor vessels, there are opportunities for the therapy of tumor, and RhoJ is expressed outside tumour vasculature and is involved in wound healing. Taking advantage of the opportunities could result in a development in tumor therapy. PMID:27556037

  7. Anchoring of both PKA and 14-3-3 inhibits the Rho-GEF activity of the AKAP-Lbc signaling complex.

    PubMed

    Diviani, Dario; Abuin, Liliane; Cotecchia, Susanna; Pansier, Laetitia

    2004-07-21

    A-kinase anchoring proteins (AKAPs) target the cAMP-regulated protein kinase (PKA) to its physiological substrates. We recently identified a novel anchoring protein, called AKAP-Lbc, which functions as a PKA-targeting protein as well as a guanine nucleotide exchange factor (GEF) for RhoA. We demonstrated that AKAP-Lbc Rho-GEF activity is stimulated by the alpha subunit of the heterotrimeric G protein G12. Here, we identified 14-3-3 as a novel regulatory protein interacting with AKAP-Lbc. Elevation of the cellular concentration of cAMP activates the PKA holoenzyme anchored to AKAP-Lbc, which phosphorylates the anchoring protein on the serine 1565. This phosphorylation event induces the recruitment of 14-3-3, which inhibits the Rho-GEF activity of AKAP-Lbc. AKAP-Lbc mutants that fail to interact with PKA or with 14-3-3 show a higher basal Rho-GEF activity as compared to the wild-type protein. This suggests that, under basal conditions, 14-3-3 maintains AKAP-Lbc in an inactive state. Therefore, while it is known that AKAP-Lbc activity can be stimulated by Galpha12, in this study we demonstrated that it is inhibited by the anchoring of both PKA and 14-3-3.

  8. The 'invisible hand': regulation of RHO GTPases by RHOGDIs.

    PubMed

    Garcia-Mata, Rafael; Boulter, Etienne; Burridge, Keith

    2011-07-22

    The 'invisible hand' is a term originally coined by Adam Smith in The Theory of Moral Sentiments to describe the forces of self-interest, competition and supply and demand that regulate the resources in society. This metaphor continues to be used by economists to describe the self-regulating nature of a market economy. The same metaphor can be used to describe the RHO-specific guanine nucleotide dissociation inhibitor (RHOGDI) family, which operates in the background, as an invisible hand, using similar forces to regulate the RHO GTPase cycle.

  9. The interstellar carbon abundance. II - Rho Ophiuchi and Beta Scorpii

    NASA Technical Reports Server (NTRS)

    Welty, D. E.; York, D. G.; Hobbs, L. M.

    1986-01-01

    A procedure designed to obtain increased sensitivity from high-dispersion IUE spectra by using a flat-field spectrum to remove nonrandom noise due to the response pattern of the SEC vidicon detector is described. Application of this procedure to spectra of Rho Oph and Beta(1) Sco near the spin-forbidden interstellar 2325 line of C II yields 2 sigma upper limits on absorption of W (lambda) not greater than about 4 mA. The resulting depletion of carbon from the interstellar gas toward Rho Oph exceeds a factor of 1.4.

  10. Signalling through the RhoGEF Pebble in Drosophila.

    PubMed

    Gregory, Stephen L; Lorensuhewa, Nirmal; Saint, Robert

    2010-04-01

    Small GTPase pathways of the Ras superfamily are implicated in a wide range of signalling processes in animal cells. Small GTPases control pathways by acting as molecular switches. They are converted from an inactive GDP-bound form to an active GTP-bound form by GTP exchange factors (GEFs). The spatial and temporal regulation of GEFs is a major component of the regulation of small GTPases. Here we review the role of the Drosophila RhoGEF, Pebble (the Drosophila ortholog of mammalian ECT2). We discuss its roles in cytokinesis and cell migration, highlighting the diversity with which Rho family signalling pathways operate in biological systems.

  11. RhoC and ROCKs regulate cancer cell interactions with endothelial cells.

    PubMed

    Reymond, Nicolas; Im, Jae Hong; Garg, Ritu; Cox, Susan; Soyer, Magali; Riou, Philippe; Colomba, Audrey; Muschel, Ruth J; Ridley, Anne J

    2015-06-01

    RhoC is a member of the Rho GTPase family that is implicated in cancer progression by stimulating cancer cell invasiveness. Here we report that RhoC regulates the interaction of cancer cells with vascular endothelial cells (ECs), a crucial step in the metastatic process. RhoC depletion by RNAi reduces PC3 prostate cancer cell adhesion to ECs, intercalation between ECs as well as transendothelial migration in vitro. Depletion of the kinases ROCK1 and ROCK2, two known RhoC downstream effectors, similarly decreases cancer interaction with ECs. RhoC also regulates the extension of protrusions made by cancer cells on vascular ECs in vivo. Transient RhoC depletion is sufficient to reduce both early PC3 cell retention in the lungs and experimental metastasis formation in vivo. Our results indicate RhoC plays a central role in cancer cell interaction with vascular ECs, which is a critical event for cancer progression.

  12. Dental enamel structure is altered by expression of dominant negative RhoA in ameloblasts.

    PubMed

    Li, Yong; Pugach, Megan K; Kuehl, Melissa A; Peng, Li; Bouchard, Jessica; Hwang, Soon Y; Gibson, Carolyn W

    2011-01-01

    Using in vitrotooth germ cultures and analysis by confocal microscopy, ameloblasts treated with sodium fluoride were found to have elevated amounts of filamentous actin. Because this response is reduced by inhibitors of the Rho/ROCK signaling pathway, we generated mice that express dominant negative RhoA (RhoA(DN)) in ameloblasts for in vivo analysis. Expression of the EGFP-RhoA(DN) fusion protein was evaluated by RT-PCR and immunohistochemistry, and teeth were analyzed by scanning electron microscopy. The 3 strains expressed at either low (TgEGFP-RhoA(DN)-8), intermediate (TgEGFP-RhoA(DN)-2), or high (TgEGFP-RhoA(DN)-13) levels, and the molar teeth from the 3 strains had enamel hypoplasia and surface defects. We conclude that RhoA(DN) expressed in ameloblasts interferes with normal enamel development through the pathway that is induced by sodium fluoride.

  13. RhoE is required for contact inhibition and negatively regulates tumor initiation and progression

    PubMed Central

    Hernández-Sánchez, Marta; Poch, Enric; Guasch, Rosa M.; Ortega, Joaquín; López-Almela, Inmaculada; Palmero, Ignacio; Pérez-Roger, Ignacio

    2015-01-01

    RhoE is a small GTPase involved in the regulation of actin cytoskeleton dynamics, cell cycle and apoptosis. The role of RhoE in cancer is currently controversial, with reports of both oncogenic and tumor-suppressive functions for RhoE. Using RhoE-deficient mice, we show here that the absence of RhoE blunts contact-inhibition of growth by inhibiting p27Kip1 nuclear translocation and cooperates in oncogenic transformation of mouse primary fibroblasts. Heterozygous RhoE+/gt mice are more susceptible to chemically induced skin tumors and RhoE knock-down results in increased metastatic potential of cancer cells. These results indicate that RhoE plays a role in suppressing tumor initiation and progression. PMID:26036260

  14. RhoGDIα suppresses self-renewal and tumorigenesis of glioma stem cells

    PubMed Central

    Wu, Fan; Hu, Peishan; Li, Dengke; Hu, Yan; Qi, Yingjiao; Yin, Bin; Jiang, Tao; Yuan, Jiangang; Han, Wei; Peng, Xiaozhong

    2016-01-01

    Glioma stem cells (GSCs) are a subset of tumor cells that drive glioma initiation and progression. The molecular mechanisms underlying the maintenance of GSCs are still poorly understood. Here we investigated the role of Rho GDP dissociation inhibitor α (RhoGDIα) in GSCs. RhoGDIα was down-regulated in glioma stem cells. Over-expression of RhoGDIα suppressed the self-renewal and tumorigenesis of GSCs. Further data showed that RhoGDIα inhibited the transcription activity of stem cell marker Oct4. Moreover, inactivation of ROCK1, a downstream effector of RhoGDIα, also decreased the self-renewal and Oct4 transcription activity, and rescued the effects caused by RhoGDIα knockdown. Our results indicate that RhoGDIα is involved in the maintenance of GSCs. PMID:27557508

  15. RhoE is required for contact inhibition and negatively regulates tumor initiation and progression.

    PubMed

    Hernández-Sánchez, Marta; Poch, Enric; Guasch, Rosa M; Ortega, Joaquín; López-Almela, Inmaculada; Palmero, Ignacio; Pérez-Roger, Ignacio

    2015-07-10

    RhoE is a small GTPase involved in the regulation of actin cytoskeleton dynamics, cell cycle and apoptosis. The role of RhoE in cancer is currently controversial, with reports of both oncogenic and tumor-suppressive functions for RhoE. Using RhoE-deficient mice, we show here that the absence of RhoE blunts contact-inhibition of growth by inhibiting p27Kip1 nuclear translocation and cooperates in oncogenic transformation of mouse primary fibroblasts. Heterozygous RhoE+/gt mice are more susceptible to chemically induced skin tumors and RhoE knock-down results in increased metastatic potential of cancer cells. These results indicate that RhoE plays a role in suppressing tumor initiation and progression.

  16. RhoGAP18B Isoforms Act on Distinct Rho-Family GTPases and Regulate Behavioral Responses to Alcohol via Cofilin

    PubMed Central

    Kalahasti, Geetha; Rodan, Aylin R.; Rothenfluh, Adrian

    2015-01-01

    Responses to the effects of ethanol are highly conserved across organisms, with reduced responses to the sedating effects of ethanol being predictive of increased risk for human alcohol dependence. Previously, we described that regulators of actin dynamics, such as the Rho-family GTPases Rac1, Rho1, and Cdc42, alter Drosophila’s sensitivity to ethanol-induced sedation. The GTPase activating protein RhoGAP18B also affects sensitivity to ethanol. To better understand how different RhoGAP18B isoforms affect ethanol sedation, we examined them for their effects on cell shape, GTP-loading of Rho-family GTPase, activation of the actin-severing cofilin, and actin filamentation. Our results suggest that the RhoGAP18B-PA isoform acts on Cdc42, while PC and PD act via Rac1 and Rho1 to activate cofilin. In vivo, a loss-of-function mutation in the cofilin-encoding gene twinstar leads to reduced ethanol-sensitivity and acts in concert with RhoGAP18B. Different RhoGAP18B isoforms, therefore, act on distinct subsets of Rho-family GTPases to modulate cofilin activity, actin dynamics, and ethanol-induced behaviors. PMID:26366560

  17. Use of lycorine and DAPI staining in Saccharomyces cerevisiae to differentiate between rho0 and rho- cells in a cce1/delta cce1 nuclear background.

    PubMed

    Massardo, D R; Zweifel, S G; Gunge, N; Miyakawa, I; Sando, N; Del Giudice, A; Wolf, K; Del Giudice, L

    2000-11-01

    In the yeast Saccharomyces cerevisiae, mutants are viable with large deletions (rho-), or even complete loss of the mitochondrial genome (rho0). One class of rho- mutants, which is called hypersuppressive, is characterised by a high transmission of the mutated mitochondrial genome to the diploid progeny when mated to a wild-type (rho+) haploid. The nuclear gene CCE1 encodes a cruciform cutting endonuclease, which is located in the mitochondrion and is responsible for the highly biased transmission of the hypersuppressive rho- genome. CCE1 is a Holliday junction specific endonuclease that resolves recombination intermediates in mitochondrial DNA. The cleavage activity shows a strong preference for cutting after a 5'-CT dinucleotide. In the absence of the CCE1 gene product, the mitochondrial genomes remain interconnected and have difficulty segregating to the daughter cells. As a consequence, there is an increase in the fraction of daughter cells that are rho0. In this paper we demonstrate the usefulness of lycorine, together with staining by 4',6-diamidino-2-phenylindole (DAPI), to assay for the mitotic stability of a variety of mitochondrial genomes. We have found that rho+ and rho- strains that contain CT sequences produce a large fraction of rho0 progeny in the absence of CCE1 activity. Only those rho- mitochondrial genomes lacking the CT recognition sequence are unaffected by the cce1 allele.

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

    PubMed Central

    1993-01-01

    Evidence is accumulating that the rho family, a member of the ras p21- related small GTP-binding protein superfamily, regulates cell morphology, cell motility, and smooth muscle contraction through the actomyosin system. The actomyosin system is also known to be essential for cytoplasmic division of cells (cytokinesis). In this study, we examined the action of rho p21, its inhibitory GDP/GTP exchange protein, named rho GDI, its stimulatory GDP/GTP exchange protein, named smg GDS, and botulinum ADP-ribosyltransferase C3, known to selectively ADP-ribosylate rho p21 and to impair its function, in the cytoplasmic division using Xenopus embryos. The sperm-induced cytoplasmic division of Xenopus embryos was not affected by microinjection into the embryos of either smg GDS or the guanosine-5'-(3-O-thio)triphosphate (GTP gamma S)-bound form of rhoA p21, one member of the rho family, but completely inhibited by microinjection of rho GDI or C3. Under these conditions, nuclear division occurred normally but the furrow formation, which was induced by the contractile ring consisting of actomyosin just beneath the plasma membrane, was impaired. Comicroinjection of rho GDI with the GTP gamma S-bound form of rhoA p21 prevented the rho GDI action. Moreover, the sperm-induced cytoplasmic division of Xenopus embryos was inhibited by microinjection into the embryos of the rhoA p21 pre-ADP- ribosylated by C3 which might serve as a dominant negative inhibitor of endogenous rho p21. These results indicate that rho p21 together with its regulatory proteins regulates the cytoplasmic division through the actomyosin system. PMID:8436590

  19. Evolution of the Rho family of ras-like GTPases in eukaryotes.

    PubMed

    Boureux, Anthony; Vignal, Emmanuel; Faure, Sandrine; Fort, Philippe

    2007-01-01

    GTPases of the Rho family are molecular switches that play important roles in converting and amplifying external signals into cellular effects. Originally demonstrated to control the dynamics of the F-actin cytoskeleton, Rho GTPases have been implicated in many basic cellular processes that influence cell proliferation, differentiation, motility, adhesion, survival, or secretion. To elucidate the evolutionary history of the Rho family, we have analyzed over 20 species covering major eukaryotic clades from unicellular organisms to mammals, including platypus and opossum, and have reconstructed the ontogeny and the chronology of emergence of the different subfamilies. Our data establish that the 20 mammalian Rho members are structured into 8 subfamilies, among which Rac is the founder of the whole family. Rho, Cdc42, RhoUV, and RhoBTB subfamilies appeared before Coelomates and RhoJQ, Cdc42 isoforms, RhoDF, and Rnd emerged in chordates. In vertebrates, gene duplications and retrotranspositions increased the size of each chordate Rho subfamily, whereas RhoH, the last subfamily, arose probably by horizontal gene transfer. Rac1b, a Rac1 isoform generated by alternative splicing, emerged in amniotes, and RhoD, only in therians. Analysis of Rho mRNA expression patterns in mouse tissues shows that recent subfamilies have tissue-specific and low-level expression that supports their implication only in narrow time windows or in differentiated metabolic functions. These findings give a comprehensive view of the evolutionary canvas of the Rho family and provide guides for future structure and evolution studies of other components of Rho signaling pathways, in particular regulators of the RhoGEF family.

  20. Evolution of the Rho family of ras-like GTPases in eukaryotes

    PubMed Central

    Boureux, Anthony; Vignal, Emmanuel; Faure, Sandrine; Fort, Philippe

    2007-01-01

    GTPases of the Rho family are molecular switches that play important roles in converting and amplifying external signals into cellular effects. Originally demonstrated to control the dynamics of the F-actin cytoskeleton, Rho GTPases have been implicated in many basic cellular processes that influence cell proliferation, differentiation, motility, adhesion, survival or secretion. To elucidate the evolutionary history of the Rho family, we have analyzed over twenty species covering major eukaryotic clades from unicellular organisms to mammals, including platypus and opossum, and have reconstructed the ontogeny and the chronology of emergence of the different subfamilies. Our data establish that the 20 mammalian Rho members are structured into eight subfamilies, among which Rac is the founder of the whole family. Rho, Cdc42, RhoUV and RhoBTB subfamilies appeared before Coelomates, and RhoJQ, RhoDF and Rnd emerged in Chordates. In Vertebrates, gene duplications and retrotranspositions increased the size of each chordate Rho subfamily, while RhoH, the last subfamily, arose probably by horizontal gene transfer. Rac1b, a Rac1 isoform generated by alternative splicing, emerged in amniotes, and RhoD, only in therians. Analysis of Rho mRNA expression patterns in mouse tissues shows that recent subfamilies have tissue-specific specific and low level expression, which supports their implication only in narrow time windows or in differentiated metabolic functions. These findings give a comprehensive view of the evolutionary canvas of the Rho family and provide guides for future structure and evolution studies of other components of Rho signaling pathways, in particular regulators of the RhoGEF family. PMID:17035353

  1. RhoA promotes epidermal stem cell proliferation via PKN1-cyclin D1 signaling

    PubMed Central

    Wang, Fan; Zhan, Rixing; Chen, Liang; Dai, Xia; Wang, Wenping; Guo, Rui; Li, Xiaoge; Li, Zhe; Wang, Liang; Huang, Shupeng; Shen, Jie

    2017-01-01

    Objective Epidermal stem cells (ESCs) play a critical role in wound healing, but the mechanism underlying ESC proliferation is not well defined. Here, we explore the effects of RhoA on ESC proliferation and the possible underlying mechanism. Methods Human ESCs were enriched by rapid adhesion to collagen IV. RhoA(+/+)(G14V), RhoA(-/-)(T19N) and pGFP control plasmids were transfected into human ESCs. The effect of RhoA on cell proliferation was detected by cell proliferation and DNA synthesis assays. Induction of PKN1 activity by RhoA was determined by immunoblot analysis, and the effects of PKN1 on RhoA in terms of inducing cell proliferation and cyclin D1 expression were detected using specific siRNA targeting PKN1. The effects of U-46619 (a RhoA agonist) and C3 transferase (a RhoA antagonist) on ESC proliferation were observed in vivo. Results RhoA had a positive effect on ESC proliferation, and PKN1 activity was up-regulated by the active RhoA mutant (G14V) and suppressed by RhoA T19N. Moreover, the ability of RhoA to promote ESC proliferation and DNA synthesis was interrupted by PKN1 siRNA. Additionally, cyclin D1 protein and mRNA expression levels were up-regulated by RhoA G14V, and these effects were inhibited by siRNA-mediated knock-down of PKN1. RhoA also promoted ESC proliferation via PKN in vivo. Conclusion This study shows that the effect of RhoA on ESC proliferation is mediated by activation of the PKN1-cyclin D1 pathway in vitro, suggesting that RhoA may serve as a new therapeutic target for wound healing. PMID:28222172

  2. Problem-Solving Test: The Mechanism of Transcription Termination by the Rho Factor

    ERIC Educational Resources Information Center

    Szeberenyi, Jozsef

    2012-01-01

    Transcription termination comes in two forms in "E. coli" cells. Rho-dependent termination requires the binding of a termination protein called Rho factor to the transcriptional machinery at the terminator region, whereas Rho-independent termination is achieved by conformational changes in the transcript itself. This article presents a test…

  3. Structural Mechanisms and Drug Discovery Prospects of Rho GTPases

    PubMed Central

    Smithers, Cameron C.; Overduin, Michael

    2016-01-01

    Rho GTPases regulate cellular morphology and dynamics, and some are key drivers of cancer progression. This superfamily offers attractive potential targets for therapeutic intervention, with RhoA, Rac1 and Cdc42 being prime examples. The challenges in developing agents that act on these signaling enzymes include the lack of obvious druggable pockets and their membrane-bound activities. However, progress in targeting the similar Ras protein is illuminating new strategies for specifically inhibiting oncogenic GTPases. The structures of multiple signaling and regulatory states of Rho proteins have been determined, and the post-translational modifications including acylation and phosphorylation points have been mapped and their functional effects examined. The development of inhibitors to probe the significance of overexpression and mutational hyperactivation of these GTPases underscores their importance in cancer progression. The ability to integrate in silico, in vitro, and in vivo investigations of drug-like molecules indicates the growing tractability of GTPase systems for lead optimization. Although no Rho-targeted drug molecules have yet been clinically approved, this family is clearly showing increasing promise for the development of precision medicine and combination cancer therapies. PMID:27304967

  4. Exclusive electroproduction of the rho+ meson on the proton @ CLAS

    SciTech Connect

    Ahmed Fradi

    2011-10-01

    We present preliminary results of the exclusive electroproduction of {rho}{sup +} on the proton at CLAS. We discuss the interpretation of the cross sections in terms of t-channel Reggeon exchanges and in terms of Generalized Parton Distributions (GPDs) formalism.

  5. Epithelial junctions and Rho family GTPases: the zonular signalosome

    PubMed Central

    Citi, Sandra; Guerrera, Diego; Spadaro, Domenica; Shah, Jimit

    2014-01-01

    The establishment and maintenance of epithelial cell-cell junctions is crucially important to regulate adhesion, apico-basal polarity and motility of epithelial cells, and ultimately controls the architecture and physiology of epithelial organs. Junctions are supported, shaped and regulated by cytoskeletal filaments, whose dynamic organization and contractility are finely tuned by GTPases of the Rho family, primarily RhoA, Rac1 and Cdc42. Recent research has identified new molecular mechanisms underlying the cross-talk between these GTPases and epithelial junctions. Here we briefly summarize the current knowledge about the organization, molecular evolution and cytoskeletal anchoring of cell-cell junctions, and we comment on the most recent advances in the characterization of the interactions between Rho GTPases and junctional proteins, and their consequences with regards to junction assembly and regulation of cell behavior in vertebrate model systems. The concept of “zonular signalosome” is proposed, which highlights the close functional relationship between proteins of zonular junctions (zonulae occludentes and adhaerentes) and the control of cytoskeletal organization and signaling through Rho GTPases, transcription factors, and their effectors. PMID:25483301

  6. {rho} meson decay in 2+1 flavor lattice QCD

    SciTech Connect

    Aoki, S.; Ishizuka, N.; Taniguchi, Y.; Ukawa, A.; Yoshie, T.; Ishikawa, K-I.; Okawa, M.; Kanaya, K.; Kuramashi, Y.; Namekawa, Y.; Ukita, N.; Yamazaki, T.

    2011-11-01

    We perform a lattice QCD study of the {rho} meson decay from the N{sub f}=2+1 full QCD configurations generated with a renormalization group improved gauge action and a nonperturbatively O(a)-improved Wilson fermion action. The resonance parameters, the effective {rho}{yields}{pi}{pi} coupling constant and the resonance mass, are estimated from the P-wave scattering phase shift for the isospin I=1 two-pion system. The finite size formulas are employed to calculate the phase shift from the energy on the lattice. Our calculations are carried out at two quark masses, m{sub {pi}=}410 MeV (m{sub {pi}/}m{sub {rho}=}0.46) and m{sub {pi}=}300 MeV (m{sub {pi}/}m{sub {rho}=}0.35), on a 32{sup 3}x64 (La=2.9 fm) lattice at the lattice spacing a=0.091 fm. We compare our results at these two quark masses with those given in the previous works using N{sub f}=2 full QCD configurations and the experiment.

  7. IQGAP1 Interaction with RHO Family Proteins Revisited

    PubMed Central

    Nouri, Kazem; Fansa, Eyad K.; Amin, Ehsan; Dvorsky, Radovan; Gremer, Lothar; Willbold, Dieter; Schmitt, Lutz; Timson, David J.; Ahmadian, Mohammad R.

    2016-01-01

    IQ motif-containing GTPase activating protein 1 (IQGAP1) plays a central role in the physical assembly of relevant signaling networks that are responsible for various cellular processes, including cell adhesion, polarity, and transmigration. The RHO family proteins CDC42 and RAC1 have been shown to mainly interact with the GAP-related domain (GRD) of IQGAP1. However, the role of its RASGAP C-terminal (RGCT) and C-terminal domains in the interactions with RHO proteins has remained obscure. Here, we demonstrate that IQGAP1 interactions with RHO proteins underlie a multiple-step binding mechanism: (i) a high affinity, GTP-dependent binding of RGCT to the switch regions of CDC42 or RAC1 and (ii) a very low affinity binding of GRD and a C terminus adjacent to the switch regions. These data were confirmed by phosphomimetic mutation of serine 1443 to glutamate within RGCT, which led to a significant reduction of IQGAP1 affinity for CDC42 and RAC1, clearly disclosing the critical role of RGCT for these interactions. Unlike CDC42, an extremely low affinity was determined for the RAC1-GRD interaction, suggesting that the molecular nature of IQGAP1 interaction with CDC42 partially differs from that of RAC1. Our study provides new insights into the interaction characteristics of IQGAP1 with RHO family proteins and highlights the complementary importance of kinetic and equilibrium analyses. We propose that the ability of IQGAP1 to interact with RHO proteins is based on a multiple-step binding process, which is a prerequisite for the dynamic functions of IQGAP1 as a scaffolding protein and a critical mechanism in temporal regulation and integration of IQGAP1-mediated cellular responses. PMID:27815503

  8. Basal and Activated Calcium Sensitization Mediated by RhoA/Rho Kinase Pathway in Rats with Genetic and Salt Hypertension

    PubMed Central

    Bencze, Michal; Vaněčková, Ivana; Kuneš, Jaroslav; Zicha, Josef

    2017-01-01

    Calcium sensitization mediated by RhoA/Rho kinase pathway can be evaluated either in the absence (basal calcium sensitization) or in the presence of endogenous vasoconstrictor systems (activated calcium sensitization). Our aim was to compare basal and activated calcium sensitization in three forms of experimental hypertension with increased sympathetic tone and enhanced calcium entry—spontaneously hypertensive rats (SHR), heterozygous Ren-2 transgenic rats (TGR), and salt hypertensive Dahl rats. Activated calcium sensitization was determined as blood pressure reduction induced by acute administration of Rho kinase inhibitor fasudil in conscious rats with intact sympathetic nervous system (SNS) and renin-angiotensin system (RAS). Basal calcium sensitization was studied as fasudil-dependent difference in blood pressure response to calcium channel opener BAY K8644 in rats subjected to RAS and SNS blockade. Calcium sensitization was also estimated from reduced development of isolated artery contraction by Rho kinase inhibitor Y-27632. Activated calcium sensitization was enhanced in all three hypertensive models (due to the hyperactivity of vasoconstrictor systems). In contrast, basal calcium sensitization was reduced in SHR and TGR relative to their controls, whereas it was augmented in salt-sensitive Dahl rats relative to their salt-resistant controls. Similar differences in calcium sensitization were seen in femoral arteries of SHR and Dahl rats. PMID:28197417

  9. Observation of e^+e^- to \\rho^+\\rho^- near \\sqrt{s}=10.58\\gev

    SciTech Connect

    Aubert, B.; Bona, M.; Karyotakis, Y.; Lees, J.P.; Poireau, V.; Prencipe, E.; Prudent, X.; Tisserand, V.; Garra Tico, J.; Grauges, E.; Lopez, L.; Palano, A.; Pappagallo, M.; Eigen, G.; Stugu, B.; Sun, L.; Abrams, G.S.; Battaglia, M.; Brown, David Nathan; Cahn, R.N.; Jacobsen, R.G.; /LBL, Berkeley /UC, Berkeley /Birmingham U. /Ruhr U., Bochum /Bristol U. /British Columbia U. /Brunel U. /Novosibirsk, IYF /UC, Irvine /UCLA /UC, Riverside /UC, San Diego /UC, Santa Barbara /UC, Santa Cruz /Caltech /Cincinnati U. /Colorado U. /Colorado State U. /Dortmund U. /Dresden, Tech. U. /Ecole Polytechnique /Edinburgh U. /INFN, Ferrara /Ferrara U. /Frascati /INFN, Genoa /Genoa U. /Harvard U. /Heidelberg U. /Humboldt U., Berlin /Imperial Coll., London /Iowa U. /Iowa State U. /Johns Hopkins U. /Karlsruhe U. /Orsay, LAL /LLNL, Livermore /Liverpool U. /Queen Mary, U. of London /Royal Holloway, U. of London /Louisville U. /Manchester U. /Maryland U. /Massachusetts U., Amherst /MIT, LNS /McGill U. /INFN, Milan /Milan U. /Mississippi U. /Montreal U. /Mt. Holyoke Coll. /INFN, Naples /Naples U. /NIKHEF, Amsterdam /Notre Dame U. /Ohio State U. /Oregon U. /INFN, Padua /Padua U. /Paris U., VI-VII /Pennsylvania U. /INFN, Perugia /Perugia U. /INFN, Pisa /Pisa U. /Princeton U. /INFN, Rome /Rome U. /Rostock U. /Rutherford /DSM, DAPNIA, Saclay /South Carolina U. /SLAC /Stanford U., Phys. Dept. /SUNY, Albany /Tennessee U. /Texas U. /Texas U., Dallas /INFN, Turin /Turin U. /INFN, Trieste /Trieste U. /Valencia U., IFIC /Victoria U. /Warwick U. /Wisconsin U., Madison

    2008-06-26

    The authors report the first observation of e{sup +}e{sup -} {yields} {rho}{sup +}{rho}{sup -}, in a data sample of 379 fb{sup -1} collected with the BABAR detector at the PEP-II e{sup +}e{sup -} storage ring at center-of-mass energies near {radical}s = 10.58 GeV. The authors measure a cross section of {sigma}(e{sup +}e{sup -} {yields} {rho}{sup +}{rho}{sup -}) = 19.5 {+-} 1.6(stat) {+-} 3.2(syst) fb. Assuming production through single-photon annihilation, there are three independent helicity amplitudes. They measure the ratios of their squared moduli to be |F{sub 00}|{sup 2} : |F{sub 10}|{sup 2} : |F{sub 11}|{sup 2} = 0.51 {+-} 0.14(stat) {+-} 0.07(syst) : 0.10 {+-} 0.04(stat) {+-} 0.01(syst) : 0.04 {+-} 0.03(stat) {+-} 0.01(syst). The |F{sub 00}|{sup 2} result is inconsistent with the prediction of 1.0 made by QCD models with a significance of 3.1 standard deviations including systematic uncertainties.

  10. Basal and Activated Calcium Sensitization Mediated by RhoA/Rho Kinase Pathway in Rats with Genetic and Salt Hypertension.

    PubMed

    Behuliak, Michal; Bencze, Michal; Vaněčková, Ivana; Kuneš, Jaroslav; Zicha, Josef

    2017-01-01

    Calcium sensitization mediated by RhoA/Rho kinase pathway can be evaluated either in the absence (basal calcium sensitization) or in the presence of endogenous vasoconstrictor systems (activated calcium sensitization). Our aim was to compare basal and activated calcium sensitization in three forms of experimental hypertension with increased sympathetic tone and enhanced calcium entry-spontaneously hypertensive rats (SHR), heterozygous Ren-2 transgenic rats (TGR), and salt hypertensive Dahl rats. Activated calcium sensitization was determined as blood pressure reduction induced by acute administration of Rho kinase inhibitor fasudil in conscious rats with intact sympathetic nervous system (SNS) and renin-angiotensin system (RAS). Basal calcium sensitization was studied as fasudil-dependent difference in blood pressure response to calcium channel opener BAY K8644 in rats subjected to RAS and SNS blockade. Calcium sensitization was also estimated from reduced development of isolated artery contraction by Rho kinase inhibitor Y-27632. Activated calcium sensitization was enhanced in all three hypertensive models (due to the hyperactivity of vasoconstrictor systems). In contrast, basal calcium sensitization was reduced in SHR and TGR relative to their controls, whereas it was augmented in salt-sensitive Dahl rats relative to their salt-resistant controls. Similar differences in calcium sensitization were seen in femoral arteries of SHR and Dahl rats.

  11. Small interfering RNAs as a tool to assign Rho GTPase exchange-factor function in vivo.

    PubMed Central

    Gampel, Alexandra; Mellor, Harry

    2002-01-01

    Rho GTPases control a complex network of intracellular signalling pathways. Whereas progress has been made in identifying downstream signalling partners for these proteins, the characterization of Rho upstream regulatory guanine-nucleotide exchange factors (GEFs) has been hampered by a lack of suitable research tools. Here we use small interfering RNAs (siRNAs) to examine the cellular regulation of the RhoB GTPase, and show that RhoB is activated downstream of the epidermal-growth-factor receptor through the Vav2 exchange factor. These studies demonstrate that siRNAs are an ideal research tool for the assignment of Rho GEF function in vivo. PMID:12113653

  12. Polo-like kinase Cdc5 controls the local activation of Rho1 to promote cytokinesis.

    PubMed

    Yoshida, Satoshi; Kono, Keiko; Lowery, Drew M; Bartolini, Sara; Yaffe, Michael B; Ohya, Yoshikazu; Pellman, David

    2006-07-07

    The links between the cell cycle machinery and the cytoskeletal proteins controlling cytokinesis are poorly understood. The small guanine nucleotide triphosphate (GTP)-binding protein RhoA stimulates type II myosin contractility and formin-dependent assembly of the cytokinetic actin contractile ring. We found that budding yeast Polo-like kinase Cdc5 controls the targeting and activation of Rho1 (RhoA) at the division site via Rho1 guanine nucleotide exchange factors. This role of Cdc5 (Polo-like kinase) in regulating Rho1 is likely to be relevant to cytokinesis and asymmetric cell division in other organisms.

  13. Assessment of roles for the Rho-specific guanine nucleotide dissociation inhibitor (RhoGDI) Ly-GDI in platelet function: a spatial systems approach.

    PubMed

    Ngo, Anh T P; Thierheimer, Marisa L D; Babur, Özgün; Rocheleau, Anne D; Huang, Tao; Pang, Jiaqing; Rigg, Rachel A; Mitrugno, Annachiara; Theodorescu, Dan; Burchard, Julja; Nan, Xiaolin; Demir, Emek; McCarty, Owen J T; Aslan, Joseph E

    2017-02-01

    Upon activation at sites of vascular injury, platelets undergo morphological alterations essential to hemostasis via cytoskeletal reorganizations driven by the Rho GTPases Rac1, Cdc42 and RhoA. Here we investigate roles for Rho-specific guanine nucleotide dissociation inhibitor proteins (RhoGDIs) in platelet function. We find that platelets express two RhoGDI family members, RhoGDI and Ly-GDI. While RhoGDI localizes throughout platelets in a granule-like manner, Ly-GDI shows an asymmetric, polarized localization that largely overlaps with Rac1 and Cdc42 as well as microtubules and protein kinase C (PKC) in platelets adherent to fibrinogen. Antibody interference and platelet spreading experiments suggest a specific role for Ly-GDI in platelet function. Intracellular signaling studies based on interactome and pathways analyses also support a regulatory role for Ly-GDI, which is phosphorylated at PKC substrate motifs in a PKC-dependent manner in response to the platelet collagen receptor glycoprotein (GP)VI-specific agonist collagen-related peptide. Additionally, PKC inhibition diffuses the polarized organization of Ly-GDI in spread platelets relative to its colocalization with Rac1 and Cdc42. Together our results suggest a role for Ly-GDI in the localized regulation of Rho GTPases in platelets and hypothesize a link between the PKC and Rho GTPase signaling systems in platelet function.

  14. The crystal structure of the RhoA–AKAP-Lbc DH–PH domain complex

    PubMed Central

    Abdul Azeez, Kamal R.; Knapp, Stefan; Fernandes, João M. P.; Klussmann, Enno; Elkins, Jonathan M.

    2014-01-01

    The RhoGEF (Rho GTPase guanine-nucleotide-exchange factor) domain of AKAP-Lbc (A-kinase-anchoring protein-Lbc, also known as AKAP13) catalyses nucleotide exchange on RhoA and is involved in the development of cardiac hypertrophy. The RhoGEF activity of AKAP-Lbc has also been implicated in cancer. We have determined the X-ray crystal structure of the complex between RhoA–GDP and the AKAP-Lbc RhoGEF [DH (Dbl-homologous)–PH (pleckstrin homology)] domain to 2.1 Å (1 Å=0.1 nm) resolution. The structure reveals important differences compared with related RhoGEF proteins such as leukaemia-associated RhoGEF. Nucleotide-exchange assays comparing the activity of the DH–PH domain to the DH domain alone showed no role for the PH domain in nucleotide exchange, which is explained by the RhoA–AKAP-Lbc structure. Comparison with a structure of the isolated AKAP-Lbc DH domain revealed a change in conformation of the N-terminal ‘GEF switch’ region upon binding to RhoA. Isothermal titration calorimetry showed that AKAP-Lbc has only micromolar affinity for RhoA, which combined with the presence of potential binding pockets for small molecules on AKAP-Lbc, raises the possibility of targeting AKAP-Lbc with GEF inhibitors. PMID:25186459

  15. ATP-dependent motor activity of the transcription termination factor Rho from Mycobacterium tuberculosis.

    PubMed

    D'Heygère, François; Schwartz, Annie; Coste, Franck; Castaing, Bertrand; Boudvillain, Marc

    2015-07-13

    The bacterial transcription termination factor Rho-a ring-shaped molecular motor displaying directional, ATP-dependent RNA helicase/translocase activity-is an interesting therapeutic target. Recently, Rho from Mycobacterium tuberculosis (MtbRho) has been proposed to operate by a mechanism uncoupled from molecular motor action, suggesting that the manner used by Rho to dissociate transcriptional complexes is not conserved throughout the bacterial kingdom. Here, however, we demonstrate that MtbRho is a bona fide molecular motor and directional helicase which requires a catalytic site competent for ATP hydrolysis to disrupt RNA duplexes or transcription elongation complexes. Moreover, we show that idiosyncratic features of the MtbRho enzyme are conferred by a large, hydrophilic insertion in its N-terminal 'RNA binding' domain and by a non-canonical R-loop residue in its C-terminal 'motor' domain. We also show that the 'motor' domain of MtbRho has a low apparent affinity for the Rho inhibitor bicyclomycin, thereby contributing to explain why M. tuberculosis is resistant to this drug. Overall, our findings support that, in spite of adjustments of the Rho motor to specific traits of its hosting bacterium, the basic principles of Rho action are conserved across species and could thus constitute pertinent screening criteria in high-throughput searches of new Rho inhibitors.

  16. Platelet Rho GTPases–a focus on novel players, roles and relationships

    PubMed Central

    Goggs, Robert; Williams, Christopher M.; Mellor, Harry; Poole, Alastair W.

    2015-01-01

    Rho GTPases are critical for platelet function. Although the roles of RhoA, Rac and Cdc42 are characterized, platelets express other Rho GTPases, whose activities are less well understood. This review summarizes our understanding of the roles of platelet Rho GTPases and focuses particularly on the functions of Rif and RhoG. In human platelets, Rif interacts with cytoskeleton regulators including formins mDia1 and mDia3, whereas RhoG binds SNARE-complex proteins and cytoskeletal regulators ELMO and DOCK1. Knockout mouse studies suggest that Rif plays no critical functions in platelets, likely due to functional overlap with other Rho GTPases. In contrast, RhoG is essential for normal granule secretion downstream of the collagen receptor GPVI. The central defect in RhoG−/− platelets is reduced dense granule secretion, which impedes integrin activation and aggregation and limits platelet recruitment to growing thrombi under shear, translating into reduced thrombus formation in vivo. Potential avenues for future work on Rho GTPases in platelets are also highlighted, including identification of the key regulator for platelet filopodia formation and investigation of the role of the many Rho GTPase regulators in platelet function in both health and disease. PMID:25748676

  17. The crystal structure of the RhoA-AKAP-Lbc DH-PH domain complex.

    PubMed

    Abdul Azeez, Kamal R; Knapp, Stefan; Fernandes, João M P; Klussmann, Enno; Elkins, Jonathan M

    2014-12-01

    The RhoGEF (Rho GTPase guanine-nucleotide-exchange factor) domain of AKAP-Lbc (A-kinase-anchoring protein-Lbc, also known as AKAP13) catalyses nucleotide exchange on RhoA and is involved in the development of cardiac hypertrophy. The RhoGEF activity of AKAP-Lbc has also been implicated in cancer. We have determined the X-ray crystal structure of the complex between RhoA-GDP and the AKAP-Lbc RhoGEF [DH (Dbl-homologous)-PH (pleckstrin homology)] domain to 2.1 Å (1 Å = 0.1 nm) resolution. The structure reveals important differences compared with related RhoGEF proteins such as leukaemia-associated RhoGEF. Nucleotide-exchange assays comparing the activity of the DH-PH domain to the DH domain alone showed no role for the PH domain in nucleotide exchange, which is explained by the RhoA-AKAP-Lbc structure. Comparison with a structure of the isolated AKAP-Lbc DH domain revealed a change in conformation of the N-terminal 'GEF switch' region upon binding to RhoA. Isothermal titration calorimetry showed that AKAP-Lbc has only micromolar affinity for RhoA, which combined with the presence of potential binding pockets for small molecules on AKAP-Lbc, raises the possibility of targeting AKAP-Lbc with GEF inhibitors.

  18. Inhibition of phospholipase D activation by CYL-26z in formyl peptide-stimulated neutrophils involves the blockade of RhoA activation.

    PubMed

    Kuan, Yu-Hsiang; Lin, Ruey-Hseng; Tsao, Lo-Ti; Chen, Yeh-Long; Tzeng, Cherng-Chyi; Wang, Jih-Pyang

    2005-09-15

    5-[4-Acridin-9-ylamino]phenyl]-5-methyl-3-methylenedihydrofuran-2-one (CYL-26z) inhibited the formyl-Met-Leu-Phe (fMLP)-stimulated phospholipase D (PLD) activity, which was assessed by the production of phosphatidylethanol (PEt) in the presence of ethanol, in rat neutrophils (IC50 1.2+/-0.2 microM). CYL-26z caused a slight but significant attenuation of the global protein tyrosine phosphorylation stimulated by fMLP only at concentrations of CYL-26z up to 30 microM. CYL-26z blocked the membrane recruitment of protein kinase C-alpha (PKC-alpha) at concentrations of CYL-26z > or =3 microM, but failed to affect the membrane association of PKC-betaI and -betaII. The translocation of RhoA to the membrane was attenuated by CYL-26z (IC50 3.8+/-0.8 microM) in fMLP-stimulated neutrophils, whereas CYL-26z caused no significant inhibition of the membrane recruitment of ADP-ribosylation factor (Arf). CYL-26z inhibited the activation of RhoA and dissociation of the RhoA-Rho guanine nucleotide dissociation inhibitor (GDI) complex in fMLP-stimulated neutrophils (IC50 1.8+/-1.0 microM and 1.8+/-0.9 microM, respectively). In a cell-free system, CYL-26z effectively attenuated the membrane association of RhoA in response to GTPgammaS (IC50 1.3+/-0.5 microM). In contrast, the GTPgammaS-stimulated translocation of Arf to membrane was suppressed only at concentrations of CYL-26z up to 30 microM. CYL-26z inhibited the fMLP-stimulated membrane expression of CD11b, CD45 and CD63, and the release of lysozyme and beta-glucuronidase. These results indicate that CYL-26z inhibited the fMLP-stimulated PLD activity, mainly through the blockade of RhoA activation, and degranulation in rat neutrophils.

  19. The Sm-like RNA chaperone Hfq mediates transcription antitermination at Rho-dependent terminators

    PubMed Central

    Rabhi, Makhlouf; Espéli, Olivier; Schwartz, Annie; Cayrol, Bastien; Rahmouni, A Rachid; Arluison, Véronique; Boudvillain, Marc

    2011-01-01

    In Escherichia coli, the essential motor protein Rho promotes transcription termination in a tightly controlled manner that is not fully understood. Here, we show that the general post-transcriptional regulatory protein Hfq associates with Rho to regulate Rho function. The Hfq:Rho complex can be further stabilized by RNA bridging both factors in a configuration that inhibits the ATP hydrolysis and duplex unwinding activities of Rho and that mediates transcription antitermination at Rho-dependent terminators in vitro and in vivo. Antitermination at a prototypical terminator (λtR1) requires Hfq binding to an A/U-rich transcript region directly upstream from the terminator. Antitermination is modulated by trans-acting factors (NusG or nucleic acid competitors) that affect Hfq association with Rho or RNA. These data unveil a new Hfq function and a novel transcription regulatory mechanism with potentially important implications for bacterial RNA metabolism, gene silencing, and pathogenicity. PMID:21673658

  20. Mechanisms for spatiotemporal regulation of Rho-GTPase signaling at synapses

    PubMed Central

    Duman, Joseph G.; Mulherkar, Shalaka; Tu, Yen-Kuei; Cheng, Jinxuan; Tolias, Kimberley F.

    2015-01-01

    Synapses mediate information flow between neurons and undergo plastic changes in response to experience, which is critical for learning and memory. Conversely, synaptic defects impair information processing and underlie many brain pathologies. Rho-family GTPases control synaptogenesis by transducing signals from extracellular stimuli to the cytoskeleton and nucleus. The Rho-GTPases Rac1 and Cdc42 promote synapse development and the growth of axons and dendrites, while RhoA antagonizes these processes. Despite its significance, many aspects of Rho-GTPase signaling remain relatively unknown. Rho-GTPases are activated by guanine nucleotide exchange factors (GEFs) and inhibited by GTPase-activating proteins (GAPs). Though the number of both GEFs and GAPs greatly exceeds that of Rho-GTPases, loss of even a single GEF or GAP often has profound effects on cognition and behavior. Here, we explore how the actions of specific GEFs and GAPs give rise to the precise spatiotemporal activation patterns of Rho-GTPases in neurons. We consider the effects of coupling GEFs and GAPs targeting the same Rho-GTPase and the modular pathways that connect specific cellular stimuli with a given Rho-GTPase via different GEFs. We discuss how the creation of sharp borders between Rho-GTPase activation zones is achieved by pairing a GEF for one Rho-GTPase with a GAP for another and the extensive crosstalk between different Rho-GTPases. Given the importance of synapses for cognition and the fundamental roles that Rho-GTPases play in regulating them, a detailed understanding of Rho-GTPase signaling is essential to the progress of neuroscience. PMID:26003445

  1. Functional consequences of Gα13 mutations that disrupt interaction with p115RhoGEF

    PubMed Central

    Grabocka, Elda; Wedegaertner, Philip B.

    2006-01-01

    The G-protein α subunit, α13, regulates cell growth and differentiation through the monomeric Rho GTPase. α13 activates Rho through direct stimulation of the guanine nucleotide exchange factor p115RhoGEF, which contains a regulator of G-protein signaling homology domain (RH) in its N-terminus. Through its RH domain p115RhoGEF also functions as a GAP for Gα13. The mechanism for the Gα13/p115RhoGEF interaction is not well understood. Here, we determined specific α13 residues important for its interaction with p115RhoGEF. GST-pull downs and co-immunoprecipitation assays revealed that individually mutating α13 residues Lys204, Glu229, or Arg232 to opposite charge residues disrupts the interaction of activated α13 with the RH domain of p115RhoGEF or full-length p115RhoGEF. We further demonstrate that mutation of Glu229, and to a lesser extent Lys204 or Arg232, disrupts the ability of activated α13 to induce the recruitment of p115RhoGEF to the plasma membrane (PM) and to activate Rho-mediated SRE-luciferase gene transcription. Interestingly, an α13 mutant where a conserved Gly was mutated to a Ser (G205S) retained its ability to bind to p115RhoGEF, induce p115RhoGEF recruitment to the PM, and activate Rho-dependent signaling, even though identical Gly to Ser mutations in other α disrupt their interaction with RGS proteins. These results demonstrate that whereas several features of a typical α/RGS interaction are preserved in the α13/p115RhoGEF interaction, there are also significant differences. PMID:15735747

  2. RhoA GTPase regulates radiation-induced alterations in endothelial cell adhesion and migration

    SciTech Connect

    Rousseau, Matthieu; Gaugler, Marie-Helene; Rodallec, Audrey; Bonnaud, Stephanie; Paris, Francois; Corre, Isabelle

    2011-11-04

    Highlights: Black-Right-Pointing-Pointer We explore the role of RhoA in endothelial cell response to ionizing radiation. Black-Right-Pointing-Pointer RhoA is rapidly activated by single high-dose of radiation. Black-Right-Pointing-Pointer Radiation leads to RhoA/ROCK-dependent actin cytoskeleton remodeling. Black-Right-Pointing-Pointer Radiation-induced apoptosis does not require the RhoA/ROCK pathway. Black-Right-Pointing-Pointer Radiation-induced alteration of endothelial adhesion and migration requires RhoA/ROCK. -- Abstract: Endothelial cells of the microvasculature are major target of ionizing radiation, responsible of the radiation-induced vascular early dysfunctions. Molecular signaling pathways involved in endothelial responses to ionizing radiation, despite being increasingly investigated, still need precise characterization. Small GTPase RhoA and its effector ROCK are crucial signaling molecules involved in many endothelial cellular functions. Recent studies identified implication of RhoA/ROCK in radiation-induced increase in endothelial permeability but other endothelial functions altered by radiation might also require RhoA proteins. Human microvascular endothelial cells HMEC-1, either treated with Y-27632 (inhibitor of ROCK) or invalidated for RhoA by RNA interference were exposed to 15 Gy. We showed a rapid radiation-induced activation of RhoA, leading to a deep reorganisation of actin cytoskeleton with rapid formation of stress fibers. Endothelial early apoptosis induced by ionizing radiation was not affected by Y-27632 pre-treatment or RhoA depletion. Endothelial adhesion to fibronectin and formation of focal adhesions increased in response to radiation in a RhoA/ROCK-dependent manner. Consistent with its pro-adhesive role, ionizing radiation also decreased endothelial cells migration and RhoA was required for this inhibition. These results highlight the role of RhoA GTPase in ionizing radiation-induced deregulation of essential endothelial

  3. Concordance and interaction of guanine nucleotide dissociation inhibitor (RhoGDI) with RhoA in oogenesis and early development of the sea urchin

    PubMed Central

    Zazueta-Novoa, Vanesa; Martínez-Cadena, Guadalupe; Wessel, Gary M.; Zazueta-Sandoval, Roberto; Castellano, Laura; García-Soto, Jesús

    2016-01-01

    Rho GTPases are Ras-related GTPases that regulate a variety of cellular processes. In the sea urchin Strongylocentrotus purpuratus, RhoA in the oocyte associates with the membrane of the cortical granules and directs their movement from the cytoplasm to the cell cortex during maturation to an egg. RhoA also plays an important role regulating the Na+-H+ exchanger activity which determines the internal pH of the cell during the first minutes of embryogenesis. We investigated how this activity may be regulated by a Guanine-nucleotide Dissociation Inhibitor (RhoGDI). The sequence of this RhoA regulatory protein was identified in the genome on the basis of its similarity to other RhoGDI species, especially for key segments in the formation of the isoprenyl-binding pocket and in interactions with the Rho GTPase. We examined the expression and the subcellular localization of RhoGDI during oogenesis and in different developmental stages. We found that RhoGDI mRNA levels were high in eggs and during cleavage divisions until blastula, when it disappeared, only to reappear in gastrula stage. RhoGDI localization overlaps the presence of RhoA during oogenesis and in embryonic development, reinforcing the regulatory premise of the interaction. By use of recombinant protein interactions in vitro, we also find that these two proteins selectively interact. These results support the hypothesis of a functional relationship in vivo and now enable mechanistic insight for the cellular and organellar rearrangements that occur during oogenesis and embryonic development. PMID:21492154

  4. Concordance and interaction of guanine nucleotide dissociation inhibitor (RhoGDI) with RhoA in oogenesis and early development of the sea urchin.

    PubMed

    Zazueta-Novoa, Vanesa; Martínez-Cadena, Guadalupe; Wessel, Gary M; Zazueta-Sandoval, Roberto; Castellano, Laura; García-Soto, Jesús

    2011-04-01

    Rho GTPases are Ras-related GTPases that regulate a variety of cellular processes. In the sea urchin Strongylocentrotus purpuratus, RhoA in the oocyte associates with the membrane of the cortical granules and directs their movement from the cytoplasm to the cell cortex during maturation to an egg. RhoA also plays an important role regulating the Na(+) -H(+) exchanger activity, which determines the internal pH of the cell during the first minutes of embryogenesis. We investigated how this activity may be regulated by a guanine-nucleotide dissociation inhibitor (RhoGDI). The sequence of this RhoA regulatory protein was identified in the genome on the basis of its similarity to other RhoGDI species, especially for key segments in the formation of the isoprenyl-binding pocket and in interactions with the Rho GTPase. We examined the expression and the subcellular localization of RhoGDI during oogenesis and in different developmental stages. We found that RhoGDI mRNA levels were high in eggs and during cleavage divisions until blastula, when it disappeared, only to reappear in gastrula stage. RhoGDI localization overlaps the presence of RhoA during oogenesis and in embryonic development, reinforcing the regulatory premise of the interaction. By use of recombinant protein interactions in vitro, we also find that these two proteins selectively interact. These results support the hypothesis of a functional relationship in vivo and now enable mechanistic insight for the cellular and organelle rearrangements that occur during oogenesis and embryonic development.

  5. Resonance Parameters of the Rho-Meson from Lattice QCD

    SciTech Connect

    Xu Feng, Karl Jansen, Dru Renner

    2011-05-01

    We perform a non-perturbative lattice calculation of the P-wave pion-pion scattering phase in the rho-meson decay channel using two flavors of maximally twisted mass fermions at pion masses ranging from 480 MeV to 290 MeV. Making use of finite-size methods, we evaluate the pion-pion scattering phase in the center-of-mass frame and two moving frames. Applying an effective range formula, we find a good description of our results for the scattering phase as a function of the energy covering the resonance region. This allows us to extract the rho-meson mass and decay width and to study their quark mass dependence.

  6. Angular momentum content of the rho meson in lattice QCD.

    PubMed

    Glozman, Leonid Ya; Lang, C B; Limmer, Markus

    2009-09-18

    The variational method allows one to study the mixing of interpolators with different chiral transformation properties in the nonperturbatively determined physical state. It is then possible to define and calculate in a gauge-invariant manner the chiral as well as the partial wave content of the quark-antiquark component of a meson in the infrared, where mass is generated. Using a unitary transformation from the chiral basis to the ;{2S+1}L_{J} basis one may extract a partial wave content of a meson. We present results for the ground state of the rho meson using quenched simulations as well as simulations with n_{f} = 2 dynamical quarks, all for lattice spacings close to 0.15 fm. We point out that these results indicate a simple ;{3}S_{1}-wave composition of the rho meson in the infrared, like in the SU(6) flavor-spin quark model.

  7. Role of Rho GTPases in desmosomal adhesion and pemphigus pathogenesis.

    PubMed

    Spindler, Volker; Waschke, Jens

    2011-05-01

    Desmosomes are distinct intercellular contacts essential to the integrity of epithelial tissues and the heart muscle. This function is impaired in the disease pemphigus, in which patients develop autoantibodies against the cadherin-type desmosomal core proteins desmogleins. Autoantibody binding induces loss of cell-cell adhesion leading to blisters within the epidermis and mucous membranes. Despite the relevance of desmosomes for integrity of such essential organs as the skin, data on the regulation of desmosome assembly and maintenance and desmosome-mediated adhesion are only slowly emerging. Small guanosine triphosphatases (GTPases) of the Rho family have long been established as regulators of other cell junctions such as adherens junctions, but also have been implicated in participating in the formation of desmosomes. In this short review we summarize two papers from our group dealing with the role of Rho family GTPases for desmosomal adhesion and pemphigus and discuss these data integrating novel work recently published.

  8. Measurement of the Michel parameter {rho} in normal muon decay

    SciTech Connect

    Tu, X.; Amann, J.F.; Bolton, R.D.; Chen, Y.; Cooper, M.D.; Cooper, P.S.; Dzemidzic, M.; Foreman, W.; Gagliardi, C.A.; Haim, D.; Harrison, R.; Hart, G.; Hogan, G.E.; Hungerford, E.V. III; Jui, C.C.H.; Knott, J.E.; Koetke, D.D.; Kozlowski, T.; Kroupa, M.A.; Lan, K.; Liu, F.; Manweiler, R.; Mayes, B.W. II; Mischke, R.E.; Otis, J.N.; Piilonen, L.E.

    1995-07-10

    A new measurement of the Michel parameter {rho} in normal muon decay has been performed using the MEGA positron spectrometer. Over 500 million triggers were recorded and the data are currently being analyzed. The previous result has a precision on the value of {rho}{plus_minus}0.0026. The present experiment expects to improve the precision to {plus_minus}0.0008 or better. The improved result will be a precise test of the standard model of electroweak interactions for a purely leptonic process. It also will provide a better constraint on the {ital W}{sub {ital R}}{minus}{ital W}{sub {ital L}} mixing angle in the left-right symmetric models. {copyright} {ital 1995} {ital American} {ital Institute} {ital of} {ital Physics}.

  9. Rho-GTPase-regulated vesicle trafficking in plant cell polarity.

    PubMed

    Chen, Xu; Friml, Jiří

    2014-02-01

    ROPs (Rho of plants) belong to a large family of plant-specific Rho-like small GTPases that function as essential molecular switches to control diverse cellular processes including cytoskeleton organization, cell polarization, cytokinesis, cell differentiation and vesicle trafficking. Although the machineries of vesicle trafficking and cell polarity in plants have been individually well addressed, how ROPs co-ordinate those processes is still largely unclear. Recent progress has been made towards an understanding of the co-ordination of ROP signalling and trafficking of PIN (PINFORMED) transporters for the plant hormone auxin in both root and leaf pavement cells. PIN transporters constantly shuttle between the endosomal compartments and the polar plasma membrane domains, therefore the modulation of PIN-dependent auxin transport between cells is a main developmental output of ROP-regulated vesicle trafficking. The present review focuses on these cellular mechanisms, especially the integration of ROP-based vesicle trafficking and plant cell polarity.

  10. Rho GTPases in insulin-stimulated glucose uptake

    PubMed Central

    Satoh, Takaya

    2014-01-01

    Insulin is secreted into blood vessels from β cells of pancreatic islets in response to high blood glucose levels. Insulin stimulates an array of physiological responses in target tissues, including liver, skeletal muscle, and adipose tissue, thereby reducing the blood glucose level. Insulin-dependent glucose uptake in skeletal muscle and adipose tissue is primarily mediated by the redistribution of the glucose transporter type 4 from intracellular storage sites to the plasma membrane. Evidence for the participation of the Rho family GTPase Rac1 in glucose uptake signaling in skeletal muscle has emerged from studies using cell cultures and genetically engineered mice. Herein, recent progress in understanding the function and regulation of Rac1, especially the cross-talk with the protein kinase Akt2, is highlighted. In addition, the role for another Rho family member TC10 and its regulatory mechanism in adipocyte insulin signaling are described. PMID:24613967

  11. Color transparency in incoherent electroproduction of {rho} mesons off nuclei

    SciTech Connect

    Nemchik, J.; Kopeliovich, B. Z.; Potashnikova, I. K.

    2013-04-15

    Color transparency (CT) phenomena in elastic electroproduction of vector mesons off nuclei are usually infected by the onset of coherence length (CL) effects. However, at low energies corresponding to the CLAS experiment at Jefferson Lab (JLab), one can study practically the net CT effects, since CL is much shorter than the nuclear radius. We investigate various manifestations of CT effects using rigorous quantum mechanical approach based on the path integral technique. We include also the effects of {rho} meson decay inside the nucleus leading to a rise of the nuclear suppression towards small values of Q{sup 2}. Motivated by the last CLAS data we predict the A, Q{sup 2} and l{sub c} dependence of nuclear transparency for {rho}{sup 0} mesons produced incoherently off nuclei. We also perform predictions for expected signal of CT corresponding to the planned JLab upgrade to 12 GeV electron beam.

  12. Lattice QCD calculation of the {rho} meson decay width

    SciTech Connect

    Aoki, S.; Fukugita, M.; Ishikawa, K-I.; Okawa, M.; Ishizuka, N.; Kuramashi, Y.; Ukawa, A.; Yoshie, T.; Kanaya, K.; Namekawa, Y.; Sasaki, K.

    2007-11-01

    We present a lattice QCD calculation of the {rho} meson decay width via the P-wave scattering phase shift for the I=1 two-pion system. Our calculation uses full QCD gauge configurations for N{sub f}=2 flavors generated using a renormalization group improved gauge action and an improved Wilson fermion action on a 12{sup 3}x24 lattice at m{sub {pi}}/m{sub {rho}}=0.41 and the lattice spacing 1/a=0.92 GeV. The phase shift calculated with the use of the finite size formula for the two-pion system in the moving frame shows a behavior consistent with the existence of a resonance at a mass close to the vector meson mass obtained in spectroscopy. The decay width estimated from the phase shift is consistent with the experiment, when the quark mass is scaled to the realistic value.

  13. Modelling Rho GTPase biochemistry to predict collective cell migration

    NASA Astrophysics Data System (ADS)

    Merchant, Brian; Feng, James

    The collective migration of cells, due to individual cell polarization and intercellular contact inhibition of locomotion, features prominently in embryogenesis and metastatic cancers. Existing methods for modelling collectively migrating cells tend to rely either on highly abstracted agent-based models, or on continuum approximations of the group. Both of these frameworks represent intercellular interactions such as contact inhibition of locomotion as hard-coded rules defining model cells. In contrast, we present a vertex-dynamics framework which predicts polarization and contact inhibition of locomotion naturally from an underlying model of Rho GTPase biochemistry and cortical mechanics. We simulate the interaction between many such model cells, and study how modulating Rho GTPases affects migratory characteristics of the group, in the context of long-distance collective migration of neural crest cells during embryogenesis.

  14. Rho kinase activation and ROS production contributes to the cooling enhanced contraction in cutaneous equine digital veins.

    PubMed

    Zerpa, H; Berhane, Y; Woodcock, H; Elliott, J; Bailey, S R

    2010-07-01

    A decrease in environmental temperature can directly affect the contractility of cutaneous vasculature, mediated in part by alpha(2)-adrenoceptors. Most of the cellular mechanisms underlying the cooling-enhanced contractility to alpha(2)-adrenoceptor agonists have been reported in cutaneous arteries but little information is available on cutaneous veins. To investigate the cellular mechanisms associated with the cooling-enhanced contraction to UK-14304 (alpha(2)-adrenoceptor agonist), isolated equine digital veins (EDVs) were studied at 30 degrees C and 22 degrees C. The effects of inhibitors were studied on the contractile response to UK-14304 (0.1 microM). The cooling-enhanced responses were inhibited by Rho kinase inhibitors [maximum response to UK-14304 95.2 +/- 8% of response to depolarizing Krebs solution (DKS) in control vessels cooled to 22 degrees C, compared with 31.4 +/- 6% in the presence of fasudil 1 microM and 75.8 +/- 6% with Y-27632 0.1 microM] and the effects of these inhibitors were considerably less at 30 degrees C (control response 56.4 +/- 5% of DKS; 34.9 +/- 6% with fasudil 1 microM and 50.6 +/- 9% with Y-27632 0.1 microM). Furthermore, Western blotting showed that one of the downstream targets for Rho kinase activity, ezrin/radixin/moesin, was phosphorylated after cooling and reduced by fasudil (1 microM) only at 22 degrees C. The activation of protein kinase C contributed to the contractile response, but predominantly at 30 degrees C (maximum response 82.3 +/- 9% of DKS for control; 57.7 +/- 10% in the presence of chelerythrine 10 microM) with no significant effect at 22 degrees C. The reduction of the response at 22 degrees C by antioxidants, rotenone (14% reduction), and tempol (21% reduction) suggested the contribution of reactive oxygen species (ROS). No evidence was obtained to support the participation of tyrosine kinase. These data demonstrate that Rho kinase activation and the production of ROS contributes to the cooling

  15. Salidroside ameliorates arthritis-induced brain cognition deficits by regulating Rho/ROCK/NF-κB pathway.

    PubMed

    Zhu, Lingpeng; Chen, Tong; Chang, Xiayun; Zhou, Rui; Luo, Fen; Liu, Jingyan; Zhang, Kai; Wang, Yue; Yang, Ying; Long, Hongyan; Liu, Yu; Yan, Tianhua; Ma, Chunhua

    2016-04-01

    The prevalence of cognitive impairment in rheumatoid arthritis (RA) patients was increasingly serious nowadays. The purpose of the current study was to explore whether salidroside (Sal) could alleviate arthritis-induced cognition deficits and examine the relationship between the impairment and Rho/ROCK/NF-κB pathway. Collagen-induced arthritis (CIA) was established by the injection of chicken type II collagen (CII), complete Freund's adjuvant (CFA) and incomplete Freund's adjuvant (IFA). Arthritic lesions of CIA rats were assessed by arthritis index score, swelling of paws and histological analysis. Cognitive deficits symptoms of CIA rats were monitored through Morris water maze test. The contents of pro-inflammatory cytokines tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β) and interleukin-6 (IL-6) in hippocampus and serum were significantly reduced with salidroside (20 mg/kg, 40 mg/kg) treatment compared with those in the CIA group. In parallel, we demonstrated that the expressions of RhoA, ROCK1, ROCK2, p-NF-κBp65, p-IκBα, p-IKKα and p-IKKβ were enhanced accompanying the investigation arthritis-induced cognition deficits, which were remarkably down-regulated by salidroside and confirmed by the results obtained from western blot and immunohistochemistry. LC-MS/MS results ascertained that Sal could enter into the blood and brain tissues to exhibit the protective effect on arthritis-induced cognitive dysfunction. Therefore, it was assumed that Sal might be a potential therapeutic candidate to treat arthritis-induced brain cognition deficits through the regulation of Rho/ROCK/NF-κB signaling.

  16. Precision measurement of the muon decay parameters {rho} and {delta}

    SciTech Connect

    MacDonald, R. P.; Gaponenko, A.; Quraan, M. A.; Bayes, R.; Davydov, Yu. I.; Faszer, W.; Fujiwara, M. C.; Gill, D. R.; Grossheim, A.; Gumplinger, P.; Henderson, R. S.; Hillairet, A.; Hu, J.; Kitching, P.; Marshall, G. M.; Mischke, R. E.; Nozar, M.; Olchanski, K.; Olin, A.; Openshaw, R.

    2008-08-01

    The TWIST Collaboration has performed new measurements of two of the parameters that describe muon decay: {rho}, which governs the shape of the overall momentum spectrum, and {delta}, which governs the momentum dependence of the parity-violating decay asymmetry. This analysis gives the results {rho}=0.750 14{+-}0.000 17(stat){+-}0.000 44(syst){+-}0.000 11({eta}), where the last uncertainty arises from the correlation between {rho} and the decay parameter {eta}, and {delta}=0.750 67{+-}0.000 30(stat){+-}0.000 67(syst). These are consistent with the value of 3/4 given for both parameters in the standard model of particle physics, and are a factor of two more precise than the measurements previously published by TWIST. A new global analysis of all available muon decay data incorporating these results is presented. Improved lower and upper limits on the decay parameter P{sub {mu}}{sup {pi}}{xi} of 0.995 24

  17. Pathophysiological Functions of Rnd3/RhoE

    PubMed Central

    Jie, Wei; Andrade, Kelsey C.; Lin, Xi; Yang, Xiangsheng; Yue, Xiaojing; Chang, Jiang

    2016-01-01

    Rnd3, also known as RhoE, belongs to the Rnd subclass of the Rho family of small GTP-binding proteins. Rnd proteins are unique due to their inability to switch from a GTP-bound to GDP-bound conformation. Even though studies of the biological function of Rnd3 are far from being concluded, information is available regarding its expression pattern, cellular localization, and its activity, which can be altered depending on the conditions. The compiled data from these studies implies that Rnd3 may not be a traditional small GTPase. The basic role of Rnd3 is to report as an endogenous antagonist of RhoA signaling-mediated actin cytoskeleton dynamics, which specifically contributes to cell migration and neuron polarity. In addition, Rnd3 also plays a critical role in arresting cell cycle distribution, inhibiting cell growth, and inducing apoptosis and differentiation. Increasing data have shown that aberrant Rnd3 expression may be the leading cause of some systemic diseases; particularly highlighted in apoptotic cardiomyopathy, developmental arrhythmogenesis and heart failure, hydrocephalus, as well as tumor metastasis and chemotherapy resistance. Therefore, a better understanding of the function of Rnd3 under different physiological and pathological conditions, through the use of suitable models, would provide a novel insight into the origin and treatment of multiple human diseases. PMID:26756630

  18. ARHGDIA mutations cause nephrotic syndrome via defective RHO GTPase signaling

    PubMed Central

    Gee, Heon Yung; Saisawat, Pawaree; Ashraf, Shazia; Hurd, Toby W.; Vega-Warner, Virginia; Fang, Humphrey; Beck, Bodo B.; Gribouval, Olivier; Zhou, Weibin; Diaz, Katrina A.; Natarajan, Sivakumar; Wiggins, Roger C.; Lovric, Svjetlana; Chernin, Gil; Schoeb, Dominik S.; Ovunc, Bugsu; Frishberg, Yaacov; Soliman, Neveen A.; Fathy, Hanan M.; Goebel, Heike; Hoefele, Julia; Weber, Lutz T.; Innis, Jeffrey W.; Faul, Christian; Han, Zhe; Washburn, Joseph; Antignac, Corinne; Levy, Shawn; Otto, Edgar A.; Hildebrandt, Friedhelm

    2013-01-01

    Nephrotic syndrome (NS) is divided into steroid-sensitive (SSNS) and -resistant (SRNS) variants. SRNS causes end-stage kidney disease, which cannot be cured. While the disease mechanisms of NS are not well understood, genetic mapping studies suggest a multitude of unknown single-gene causes. We combined homozygosity mapping with whole-exome resequencing and identified an ARHGDIA mutation that causes SRNS. We demonstrated that ARHGDIA is in a complex with RHO GTPases and is prominently expressed in podocytes of rat glomeruli. ARHGDIA mutations (R120X and G173V) from individuals with SRNS abrogated interaction with RHO GTPases and increased active GTP-bound RAC1 and CDC42, but not RHOA, indicating that RAC1 and CDC42 are more relevant to the pathogenesis of this SRNS variant than RHOA. Moreover, the mutations enhanced migration of cultured human podocytes; however, enhanced migration was reversed by treatment with RAC1 inhibitors. The nephrotic phenotype was recapitulated in arhgdia-deficient zebrafish. RAC1 inhibitors were partially effective in ameliorating arhgdia-associated defects. These findings identify a single-gene cause of NS and reveal that RHO GTPase signaling is a pathogenic mediator of SRNS. PMID:23867502

  19. The small GTPase RhoH is an atypical regulator of haematopoietic cells

    PubMed Central

    Fueller, Florian; Kubatzky, Katharina F

    2008-01-01

    Rho GTPases are a distinct subfamily of the superfamily of Ras GTPases. The best-characterised members are RhoA, Rac and Cdc42 that regulate many diverse actions such as actin cytoskeleton reorganisation, adhesion, motility as well as cell proliferation, differentiation and gene transcription. Among the 20 members of that family, only Rac2 and RhoH show an expression restricted to the haematopoietic lineage. RhoH was first discovered in 1995 as a fusion transcript with the transcriptional repressor LAZ3/BCL6. It was therefore initially named translation three four (TTF) but later on renamed RhoH due to its close relationship to the Ras/Rho family of GTPases. Since then, RhoH has been implicated in human cancer as the gene is subject to somatic hypermutation and by the detection of RHOH as a translocation partner for LAZ3/BCL6 or other genes in human lymphomas. Underexpression of RhoH is found in hairy cell leukaemia and acute myeloid leukaemia. Some of the amino acids that are crucial for GTPase activity are mutated in RhoH so that the protein is a GTPase-deficient, so-called atypical Rho GTPase. Therefore other mechanisms of regulating RhoH activity have been described. These include regulation at the mRNA level and tyrosine phosphorylation of the protein's unique ITAM-like motif. The C-terminal CaaX box of RhoH is mainly a target for farnesyl-transferase but can also be modified by geranylgeranyl-transferase. Isoprenylation of RhoH and changes in subcellular localisation may be an additional factor to fine-tune signalling. Little is currently known about its signalling, regulation or interaction partners. Recent studies have shown that RhoH negatively influences the proliferation and homing of murine haematopoietic progenitor cells, presumably by acting as an antagonist for Rac1. In leukocytes, RhoH is needed to keep the cells in a resting, non-adhesive state, but the exact mechanism has yet to be elucidated. RhoH has also been implicated as a regulatory molecule

  20. Rho family GTPases: key players in neuronal development, neuronal survival, and neurodegeneration

    PubMed Central

    Stankiewicz, Trisha R.; Linseman, Daniel A.

    2014-01-01

    The Rho family of GTPases belongs to the Ras superfamily of low molecular weight (∼21 kDa) guanine nucleotide binding proteins. The most extensively studied members are RhoA, Rac1, and Cdc42. In the last few decades, studies have demonstrated that Rho family GTPases are important regulatory molecules that link surface receptors to the organization of the actin and microtubule cytoskeletons. Indeed, Rho GTPases mediate many diverse critical cellular processes, such as gene transcription, cell–cell adhesion, and cell cycle progression. However, Rho GTPases also play an essential role in regulating neuronal morphology. In particular, Rho GTPases regulate dendritic arborization, spine morphogenesis, growth cone development, and axon guidance. In addition, more recent efforts have underscored an important function for Rho GTPases in regulating neuronal survival and death. Interestingly, Rho GTPases can exert either a pro-survival or pro-death signal in neurons depending upon both the cell type and neurotoxic insult involved. This review summarizes key findings delineating the involvement of Rho GTPases and their effectors in the regulation of neuronal survival and death. Collectively, these results suggest that dysregulation of Rho family GTPases may potentially underscore the etiology of some forms of neurodegenerative disease such as amyotrophic lateral sclerosis. PMID:25339865

  1. Axon growth inhibition by RhoA/ROCK in the central nervous system.

    PubMed

    Fujita, Yuki; Yamashita, Toshihide

    2014-01-01

    Rho kinase (ROCK) is a serine/threonine kinase and a downstream target of the small GTPase Rho. The RhoA/ROCK pathway is associated with various neuronal functions such as migration, dendrite development, and axonal extension. Evidence from animal studies reveals that RhoA/ROCK signaling is involved in various central nervous system (CNS) diseases, including optic nerve and spinal cord injuries, stroke, and neurodegenerative diseases. Given that RhoA/ROCK plays a critical role in the pathophysiology of CNS diseases, the development of therapeutic agents targeting this pathway is expected to contribute to the treatment of CNS diseases. The RhoA/ROCK pathway mediates the effects of myelin-associated axon growth inhibitors-Nogo, myelin-associated glycoprotein (MAG), oligodendrocyte-myelin glycoprotein (OMgp), and repulsive guidance molecule (RGM). Blocking RhoA/ROCK signaling can reverse the inhibitory effects of these molecules on axon outgrowth, and promotes axonal sprouting and functional recovery in animal models of CNS injury. To date, several RhoA/ROCK inhibitors have been under development or in clinical trials as therapeutic agents for neurological disorders. In this review, we focus on the RhoA/ROCK signaling pathway in neurological disorders. We also discuss the potential therapeutic approaches of RhoA/ROCK inhibitors for various neurological disorders.

  2. ATP-dependent motor activity of the transcription termination factor Rho from Mycobacterium tuberculosis

    PubMed Central

    D'Heygère, François; Schwartz, Annie; Coste, Franck; Castaing, Bertrand; Boudvillain, Marc

    2015-01-01

    The bacterial transcription termination factor Rho—a ring-shaped molecular motor displaying directional, ATP-dependent RNA helicase/translocase activity—is an interesting therapeutic target. Recently, Rho from Mycobacterium tuberculosis (MtbRho) has been proposed to operate by a mechanism uncoupled from molecular motor action, suggesting that the manner used by Rho to dissociate transcriptional complexes is not conserved throughout the bacterial kingdom. Here, however, we demonstrate that MtbRho is a bona fide molecular motor and directional helicase which requires a catalytic site competent for ATP hydrolysis to disrupt RNA duplexes or transcription elongation complexes. Moreover, we show that idiosyncratic features of the MtbRho enzyme are conferred by a large, hydrophilic insertion in its N-terminal ‘RNA binding’ domain and by a non-canonical R-loop residue in its C-terminal ‘motor’ domain. We also show that the ‘motor’ domain of MtbRho has a low apparent affinity for the Rho inhibitor bicyclomycin, thereby contributing to explain why M. tuberculosis is resistant to this drug. Overall, our findings support that, in spite of adjustments of the Rho motor to specific traits of its hosting bacterium, the basic principles of Rho action are conserved across species and could thus constitute pertinent screening criteria in high-throughput searches of new Rho inhibitors. PMID:25999346

  3. Thiol-modifying phenylarsine oxide inhibits guanine nucleotide binding of Rho but not of Rac GTPases.

    PubMed

    Gerhard, Ralf; John, Harald; Aktories, Klaus; Just, Ingo

    2003-06-01

    Phenylarsine oxide (PAO) is a phosphotyrosine phosphatase inhibitor that cross-links vicinal thiol groups, thereby inactivating phosphatases possessing XCysXXCysX motifs. The RhoA-GTPase, but not the Rac1-GTPase, also possesses vicinal cysteines within the guanine nucleotide-binding region (aa 13-20) and the phosphohydrolase activity site. Treatment of Caco-2 cells with PAO showed a dose-dependent reorganization of the actin cytoskeleton, indicating involvement of Rho GTPases. As tested by pull-down experiments, RhoA, but not Rac1, from cell lysates was inactivated by PAO in a concentration-dependent manner. Modification of RhoA by PAO resulted in altered mobility on SDS-polyacrylamide gel electrophoresis, and PAO-modified RhoA was no longer substrate for C3-catalyzed ADP-ribosylation. Furthermore, RhoA treated with PAO, but not Rac1 treated with PAO, lost its property to bind to guanine nucleotides. Matrix-assisted laser desorption ionization-mass analysis of PAO-modified RhoA showed a mass shift according to an adduction of a single PAO molecule per molecule RhoA. Further analysis of Glu-C-generated RhoA peptides confirmed binding of PAO to a peptide harboring the guanine nucleotide binding region. Thus, PAO does not exclusively inhibit phosphotyrosine phosphatases but also inactivates RhoA by alteration of nucleotide binding.

  4. Axon growth inhibition by RhoA/ROCK in the central nervous system

    PubMed Central

    Fujita, Yuki; Yamashita, Toshihide

    2014-01-01

    Rho kinase (ROCK) is a serine/threonine kinase and a downstream target of the small GTPase Rho. The RhoA/ROCK pathway is associated with various neuronal functions such as migration, dendrite development, and axonal extension. Evidence from animal studies reveals that RhoA/ROCK signaling is involved in various central nervous system (CNS) diseases, including optic nerve and spinal cord injuries, stroke, and neurodegenerative diseases. Given that RhoA/ROCK plays a critical role in the pathophysiology of CNS diseases, the development of therapeutic agents targeting this pathway is expected to contribute to the treatment of CNS diseases. The RhoA/ROCK pathway mediates the effects of myelin-associated axon growth inhibitors—Nogo, myelin-associated glycoprotein (MAG), oligodendrocyte-myelin glycoprotein (OMgp), and repulsive guidance molecule (RGM). Blocking RhoA/ROCK signaling can reverse the inhibitory effects of these molecules on axon outgrowth, and promotes axonal sprouting and functional recovery in animal models of CNS injury. To date, several RhoA/ROCK inhibitors have been under development or in clinical trials as therapeutic agents for neurological disorders. In this review, we focus on the RhoA/ROCK signaling pathway in neurological disorders. We also discuss the potential therapeutic approaches of RhoA/ROCK inhibitors for various neurological disorders. PMID:25374504

  5. Ethanol increases p190RhoGAP activity, leading to actin cytoskeleton rearrangements.

    PubMed

    Selva, Javier; Egea, Gustavo

    2011-12-01

    We previously reported that cells chronically exposed to ethanol show alterations in actin cytoskeleton organization and dynamics in primary cultures of newborn rat astrocytes, a well-established in vitro model for foetal alcohol spectrum disorders. These alterations were attributed to a decrease in the cellular levels of active RhoA (RhoA-GTP), which in turn was produced by an increase in the total RhoGAP activity. We here provide evidence that p190RhoGAPs are the main factors responsible for such increase. Thus, in astrocytes chronically exposed to ethanol we observe: (i) an increase in p190A- and p190B-associated RhoGAP activity; (ii) a higher binding of p190A and p190B to RhoA-GTP; (iii) a higher p120RasGAP-p190A RhoGAP complex formation; and (iv) the recruitment of both p190RhoGAPs to the plasma membrane. The simultaneous silencing of both p190 isoforms prevents the actin rearrangements and the total RhoGAP activity increase triggered both by ethanol. Therefore, our data directly points p190RhoGAPs as ethanol-exposure molecular targets on glial cells of the CNS.

  6. RhoB controls endothelial barrier recovery by inhibiting Rac1 trafficking to the cell border

    PubMed Central

    Marcos-Ramiro, Beatriz; García-Weber, Diego; Barroso, Susana; Feito, Jorge; Ortega, María C.; Cernuda-Morollón, Eva; Reglero-Real, Natalia; Fernández-Martín, Laura; Durán, Maria C.; Alonso, Miguel A.; Correas, Isabel; Cox, Susan; Ridley, Anne J.

    2016-01-01

    Endothelial barrier dysfunction underlies chronic inflammatory diseases. In searching for new proteins essential to the human endothelial inflammatory response, we have found that the endosomal GTPase RhoB is up-regulated in response to inflammatory cytokines and expressed in the endothelium of some chronically inflamed tissues. We show that although RhoB and the related RhoA and RhoC play additive and redundant roles in various aspects of endothelial barrier function, RhoB specifically inhibits barrier restoration after acute cell contraction by preventing plasma membrane extension. During barrier restoration, RhoB trafficking is induced between vesicles containing RhoB nanoclusters and plasma membrane protrusions. The Rho GTPase Rac1 controls membrane spreading and stabilizes endothelial barriers. We show that RhoB colocalizes with Rac1 in endosomes and inhibits Rac1 activity and trafficking to the cell border during barrier recovery. Inhibition of endosomal trafficking impairs barrier reformation, whereas induction of Rac1 translocation to the plasma membrane accelerates it. Therefore, RhoB-specific regulation of Rac1 trafficking controls endothelial barrier integrity during inflammation. PMID:27138256

  7. Tech: a RhoA GEF selectively expressed in hippocampal and cortical neurons.

    PubMed

    Marx, Ruth; Henderson, Jennifer; Wang, James; Baraban, Jay M

    2005-02-01

    Recent studies implicating the Rho family of small G proteins in the regulation of neuronal morphology have focused attention on identifying key components of Rho signaling pathways in neurons. To this end, we have conducted studies aimed at defining the localization and function of Tech, a Rho guanine nucleotide exchange factor (GEF) family member that is highly enriched in brain. We have found that Tech is selectively expressed in cortical and hippocampal neurons with prominent Tech immunostaining apparent in the cell bodies and dendrites of these cells. In vitro studies with prototypical members of the major Rho subfamilies, RhoA, Rac1 and Cdc42, indicate that Tech binds selectively to and activates RhoA. To assess whether Tech may be involved in the regulation of neuronal morphology, we examined the effects of Tech constructs on the morphology of cortical neurons grown in primary culture. We found that a constitutively active Tech construct, Tech 245DeltaC, decreases the number of dendritic processes present on these neurons. This reduction appears to be mediated by activation of RhoA as it is blocked by insertion of a point mutation into the DH domain of Tech which blocks its ability to activate RhoA or coexpression of a dominant negative RhoA construct. As Tech protein levels increase during post-natal development and remain at peak levels into adulthood, these results indicate that Tech regulates RhoA signaling pathways in developing and mature forebrain neurons.

  8. Modulation of RhoA-Rho kinase-mediated Ca2+ sensitization of rabbit myometrium during pregnancy - role of Rnd3.

    PubMed

    Cario-Toumaniantz, C; Reillaudoux, G; Sauzeau, V; Heutte, F; Vaillant, N; Finet, M; Chardin, P; Loirand, G; Pacaud, P

    2003-10-15

    During pregnancy, the uterus undergoes major functional and structural remodelling. It is well known that during the major part of pregnancy, the myometrium normally remains relatively quiescent but is able to generate powerful contractions at the time of parturition. However, the intracellular molecular events regulating myometrial contractility during pregnancy still remain poorly understood. We applied differential gene expression screening using cDNA array technology to probe myometrium samples from non-pregnant and mid-pregnant (15 days) rabbits. Among the differentially expressed genes, the farnesylated small G-protein of the Rho family, Rnd3, was found to be upregulated (3.6-fold) at mid-pregnancy. Upregulation of Rnd3 was confirmed at the protein level by a 3.4-fold increase in Rnd3 expression in mid-pregnant myometrium. Measurements of contractile properties of beta-escin permeabilized smooth muscle strips revealed that the upregulation of Rnd3 correlated with an inhibition of RhoA-Rho kinase-mediated Ca2+ sensitization at mid-pregnancy. Treatment of muscle strips from mid-pregnant myometrium with the farnesyl-transferase inhibitor manumycin A (10 muM) led to the recovery of RhoA-Rho kinase-dependent Ca2+ sensitization. At late pregnancy (31 days), upregulation of RhoA and Rho kinase expression was associated with an increase in Ca2+ sensitivity of contractile proteins that was inhibited by the Rho kinase inhibitor Y-27632 (10 muM). These data thus demonstrate the time-dependent regulation of the RhoA-Rho kinase-mediated Ca2+ sensitization during the course of pregnancy. The depression of this mechanism at mid-pregnancy followed by its constitutive activation near term is associated with a co-ordinated modulation of Rnd3, RhoA and Rho kinase expression. The RhoA-Rho kinase signalling pathway and its regulators might thus represent potential targets for the development of new treatments for pre-term labour.

  9. Characterization and expression of the human rhoH12 gene product

    SciTech Connect

    Avraham, H.; Weinberg, R.A.

    1989-05-01

    The rho genes constitute an evolutionarily conserved family having significant homology to the ras oncogene family. These genes have been found in Saccharomyces cerevisiae, Drosophila melanogaster, rat, and human; their 21,000-dalton products show strong conservation of structure. In humans, three classes of rho cDNA clones have been identified which differ by virtue of the presence of variable C-terminal domains: rhoH12, rhoH6, and rhoH9. The predicted 193 amino acids of human rhoH12 protein show 88% similarity with those of the human rhoH6 clone, 96.8% similarity with those of the Aplysia rho product, and 81.8% similarity with those of the yeast RHO1 protein. Rat-1 and NIH 3T3 mouse fibroblasts were transfected with clones containing the normal human rhoH12 allele as well as the variants encoding valine in the place of the glycine and leucine in place of the gutamine normally found at residues 14 and 64, respectively. These replacements mirror the changes responsible for oncogenic activation of the related ras-encoded p21 proteins. These mutant rhoH12 clone alleles did not cause focus formation in monolayers or growth in soft agar. However, amplification of normal rhoH12 via contransfection with a dihydrofolate reductase gene resulted in colonies that displayed reduced dependence on serum for growth, grew to higher saturation densities, and were tumorigenic when inoculated into nude mice. Normal p21rho proteins was detected in the transfected cell lines as well as in normal cell lines by Western immunoblot and immunoprecipitation analysis with rabbit antibodies raised against the peptide corresponding to amino acids 122 to 135.

  10. Diagnosing fuel {rho}R and {rho}R asymmetries in cryogenic deuterium-tritium implosions using charged-particle spectrometry at OMEGA

    SciTech Connect

    Frenje, J. A.; Li, C. K.; Seguin, F. H.; Casey, D. T.; Petrasso, R. D.; Sangster, T. C.; Betti, R.; Glebov, V. Yu.; Meyerhofer, D. D.

    2009-04-15

    Determining fuel areal density ({rho}R) in moderate-{rho}R (100-200 mg/cm{sup 2}) cryogenic deuterium-tritium (DT) implosions is challenging as it requires new spectrometry techniques and analysis methods to be developed. In this paper, we describe a new method for analyzing the spectrum of knock-on deuterons (KO-Ds), elastically scattered by primary DT neutrons, from which a fuel {rho}R can be inferred for values up to {approx}200 mg/cm{sup 2}. This new analysis method, which uses Monte Carlo modeling of a cryogenic DT implosion, improves significantly the previous analysis method in two fundamental ways. First, it is not affected by significant spatial-yield variations, which degrade the diagnosis of the fuel {rho}R (spatial yield variations of about {+-}20% are typically observed), and second, it does not break down when the fuel {rho}R exceeds {approx}70 mg/cm{sup 2}.

  11. Memo-RhoA-mDia1 signaling controls microtubules, the actin network, and adhesion site formation in migrating cells.

    PubMed

    Zaoui, Kossay; Honoré, Stéphane; Isnardon, Daniel; Braguer, Diane; Badache, Ali

    2008-11-03

    Actin assembly at the cell front drives membrane protrusion and initiates the cell migration cycle. Microtubules (MTs) extend within forward protrusions to sustain cell polarity and promote adhesion site turnover. Memo is an effector of the ErbB2 receptor tyrosine kinase involved in breast carcinoma cell migration. However, its mechanism of action remained unknown. We report in this study that Memo controls ErbB2-regulated MT dynamics by altering the transition frequency between MT growth and shortening phases. Moreover, although Memo-depleted cells can assemble the Rac1-dependent actin meshwork and form lamellipodia, they show defective localization of lamellipodial markers such as alpha-actinin-1 and a reduced number of short-lived adhesion sites underlying the advancing edge of migrating cells. Finally, we demonstrate that Memo is required for the localization of the RhoA guanosine triphosphatase and its effector mDia1 to the plasma membrane and that Memo-RhoA-mDia1 signaling coordinates the organization of the lamellipodial actin network, adhesion site formation, and MT outgrowth within the cell leading edge to sustain cell motility.

  12. Nitric oxide relaxes circular smooth muscle of rat distal colon through RhoA/Rho-kinase independent Ca2+ desensitisation

    PubMed Central

    Colpaert, Erwin E; Levent, Adnan; Lefebvre, Romain A

    2005-01-01

    The aim of this study in circular smooth muscle of rat distal colon was to determine whether Ca2+ desensitisation, in addition to mechanisms lowering cytosolic free Ca2+ concentration ([Ca2+]cyt), was involved in the relaxation elicited by nitric oxide (NO). Changes in isometric tension and [Ca2+]cyt were recorded simultaneously in fura-2-loaded strips. In methacholine (10−5 M)-precontracted preparations, exogenous NO (10−4 M), adenosine 5′-triphosphate (ATP; 10−3 M) and electrical field stimulation (EFS; 1 ms, 40 V, 4 Hz, 1 min) induced a decrease in smooth muscle tension, which was accompanied by a fall in [Ca2+]cyt. The sarcoplasmic/endoplasmic reticulum Ca2+ ATP-ase (SERCA) inhibitor thapsigargin (10−6 M) did not exert an influence on the decrease in tension produced by exogenous NO, but significantly attenuated the fall in [Ca2+]cyt. Both the relaxation and the fall in [Ca2+]cyt to ATP and EFS were unaffected by thapsigargin. Calyculin-A (10−6 M), a myosin light chain phosphatase (MLCP) inhibitor, significantly reduced the decrease in tension elicited by exogenous NO, but did not alter the fall in [Ca2+]cyt to exogenous NO. Inactivating RhoA by exoenzyme C3 (2 μg ml−1) or inhibiting Rho-kinase with (+)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl) cyclohexanecarboxamide dihydrochloride monohydrate (Y-27632; 10−5 M) had no effect on the decrease of both tension and [Ca2+]cyt generated by exogenous NO. This paper demonstrates that a RhoA/Rho-kinase independent Ca2+ desensitisation pathway contributes to the relaxation by NO in circular smooth muscle strips of rat distal colon. PMID:15655498

  13. RhoA/rho kinase signaling reduces connexin43 expression in high glucose-treated glomerular mesangial cells with zonula occludens-1 involvement

    SciTech Connect

    Xie, Xi; Chen, Cheng; Huang, Kaipeng; Wang, Shaogui; Hao, Jie; Huang, Junying; Huang, Heqing

    2014-10-01

    RhoA/Rho kinase (ROCK) signaling has been suggested to be involved in diabetic nephropathy (DN) pathogenesis. Altered expression of connexin43 (Cx43) has been found in kidneys of diabetic animals. Both of them have been found to regulate nuclear factor kappa-B (NF-κB) activation in high glucose-treated glomerular mesangial cells (GMCs). The aim of this study was to investigate the relationship between RhoA/ROCK signaling and Cx43 in the DN pathogenesis. We found that upregulation of Cx43 expression inhibited NF-κB p65 nuclear translocation induced by RhoA/ROCK signaling in GMCs. Inhibition of RhoA/ROCK signaling attenuated the high glucose-induced decrease in Cx43. F-actin accumulation and an enhanced interaction between zonula occludens-1 (ZO-1) and Cx43 were observed in high glucose-treated GMCs. ZO-1 depletion or disruption of F-actin formation also inhibited the reduction in Cx43 protein levels induced by high glucose. In conclusion, activated RhoA/ROCK signaling induces Cx43 degradation in GMCs cultured in high glucose, depending on F-actin regulation. Increased F-actin induced by RhoA/ROCK signaling promotes the association between ZO-1 and Cx43, which possibly triggered Cx43 endocytosis, a mechanism of NF-κB activation in high glucose-treated GMCs. - Highlights: • RhoA/ROCK signaling induces Cx43 degradation in GMCs. • F-actin and ZO-1 have functions in the regulation of Cx43 by RhoA/ROCK signaling. • We reveal the relationship between RhoA/ROCK and Cx43 in the activation of NF-κB.

  14. The effect of sub-chronic systemic ethanol treatment on corpus cavernosal smooth muscle contraction: the contribution of RhoA/Rho-kinase.

    PubMed

    Kumcu, Eda Karabal; Aydinoglu, Fatma; Astarci, Erhan; Ogulener, Nuran

    2016-03-01

    The aim of this study was to evaluate whether the sub-chronic systemic ethanol exposure has direct effect on cavernosal smooth muscle contractions induced by KCl (depolarizing) and phenylephrine (α1-receptor agonist), and the possible involvement of RhoA/Rho-kinase pathway. Sub-chronic systemic ethanol was applied to mice with inhalation route for 14 days. The blood levels in ethanol-treated mice averaged 121.2 ± 9.1 mg/dl. KCl (80 mM) and phenylephrine (10 nM-100 μM) induced sustained contractions in corpus corporal strips from sham-treated mice. Sub-chronic ethanol treatment reduced the contractions to KCl. However, phenylephrine-induced contractions were not affected by ethanol treatment. Rho-kinase inhibitor fasudil (50 μM) and Y-27632 (50 μM) inhibited contractions to KCl and phenylephrine in sham-treated mice. Ethanol treatment increased the inhibitory effect of Rho-kinase inhibitors on contractions to phenylephrine. The relaxations induced by fasudil (100 μM) and Y-27632 (500 μM) did not change in ethanol treatment group. In ethanol-treated group, the expression of RhoA decreased compared to sham-treated group. Also, ROCK1 expression was reduced by ethanol but not statically significant to sham-treated group; however, the expression of ROCK2 increased in ethanol group. From these findings, it seems that phenylephrine and KCl-induced contractions depends on RhoA/Rho-kinase-mediated Ca(2+) sensitization. Also, these results suggest that the ethanol treatment decreased the expression of RhoA, and the inhibitory effect of ethanol on KCl-induced contractions may be due to, at least in part, the inhibition of a RhoA/Rho-kinase in mouse corpus cavernosum.

  15. Rho-associated coiled-coil containing kinases (ROCK): structure, regulation, and functions.

    PubMed

    Julian, Linda; Olson, Michael F

    2014-01-01

    Rho-associated coiled-coil containing kinases (ROCK) were originally identified as effectors of the RhoA small GTPase. (1)(-) (5) They belong to the AGC family of serine/threonine kinases (6) and play vital roles in facilitating actomyosin cytoskeleton contractility downstream of RhoA and RhoC activation. Since their discovery, ROCK kinases have been extensively studied, unveiling their manifold functions in processes including cell contraction, migration, apoptosis, survival, and proliferation. Two mammalian ROCK homologs have been identified, ROCK1 (also called ROCK I, ROKβ, Rho-kinase β, or p160ROCK) and ROCK2 (also known as ROCK II, ROKα, or Rho kinase), hereafter collectively referred to as ROCK. In this review, we will focus on the structure, regulation, and functions of ROCK.

  16. Nondeletional alpha-thalassemia: first description of alpha Hph alpha and alpha Nco alpha mutations in a Spanish population.

    PubMed

    Ayala, S; Colomer, D; Aymerich, M; Pujades, A; Vives-Corrons, J L

    1996-07-01

    Several different deletions underlie the molecular basis of alpha-thalassemia. The most common alpha-thalassemia determinant in Spain is the rightward deletion (-alpha 3.7). To our knowledge, however, no cases of alpha-thalassemia due to nondeletional mutations have so far been described in this particular Mediterranean area. Here, we report the existence of nondeletional forms of alpha-thalassemia in ten Spanish families. The alpha 2-globin gene was characterized in ten unrelated patients and their relatives only when the presence of deletional alpha-thalassemia was ruled out. The alpha 2-globin gene analysis was performed using the polymerase chain reaction (PCR) followed by restriction enzyme analysis or by allelespecific priming. This allowed the identification of a 5-base pair (bp) deletion at the donor site of IVS I (alpha Hph alpha) in 9 cases and the alpha 2 initiation codon mutation (alpha Nco alpha) in one case. Although these alpha 2-globin gene mutations are found in other mediterranean areas, our results demonstrate their presence in the Spanish population and suggest that the alpha Hph alpha/alpha alpha genotype is probably the most common nondeletional form of alpha-thalassemia in Spain.

  17. Clarifications to the limitations of the s-{alpha} equilibrium model for gyrokinetic computations of turbulence

    SciTech Connect

    Lapillonne, X.; Brunner, S.; Dannert, T.; Jolliet, S.; Marinoni, A.; Villard, L.; Goerler, T.; Jenko, F.; Merz, F.

    2009-03-15

    In the context of gyrokinetic flux-tube simulations of microturbulence in magnetized toroidal plasmas, different treatments of the magnetic equilibrium are examined. Considering the Cyclone DIII-D base case parameter set [Dimits et al., Phys. Plasmas 7, 969 (2000)], significant differences in the linear growth rates, the linear and nonlinear critical temperature gradients, and the nonlinear ion heat diffusivities are observed between results obtained using either an s-{alpha} or a magnetohydrodynamic (MHD) equilibrium. Similar disagreements have been reported previously [Redd et al., Phys. Plasmas 6, 1162 (1999)]. In this paper it is shown that these differences result primarily from the approximation made in the standard implementation of the s-{alpha} model, in which the straight field line angle is identified to the poloidal angle, leading to inconsistencies of order {epsilon} ({epsilon}=a/R is the inverse aspect ratio, a the minor radius and R the major radius). An equilibrium model with concentric, circular flux surfaces and a correct treatment of the straight field line angle gives results very close to those using a finite {epsilon}, low {beta} MHD equilibrium. Such detailed investigation of the equilibrium implementation is of particular interest when comparing flux tube and global codes. It is indeed shown here that previously reported agreements between local and global simulations in fact result from the order {epsilon} inconsistencies in the s-{alpha} model, coincidentally compensating finite {rho}{sup *} effects in the global calculations, where {rho}{sup *}={rho}{sub s}/a with {rho}{sub s} the ion sound Larmor radius. True convergence between local and global simulations is finally obtained by correct treatment of the geometry in both cases, and considering the appropriate {rho}{sup *}{yields}0 limit in the latter case.

  18. Excitation of high-n toroidicity-induced shear Alfven eigenmodes by energetic particles and fusion alpha particles in tokamaks

    SciTech Connect

    Fu, G.Y.; Cheng, C.Z.

    1992-07-01

    The stability of high-n toroidicity-induced shear Alfven eigenmodes (TAE) in the presence of fusion alpha particles or energetic ions in tokamaks is investigated. The TAE modes are discrete in nature and thus can easily tap the free energy associated with energetic particle pressure gradient through wave particle resonant interaction. A quadratic form is derived for the high-n TAE modes using gyro-kinetic equation. The kinetic effects of energetic particles are calculated perturbatively using the ideal MHD solution as the lowest order eigenfunction. The finite Larmor radius (FLR) effects and the finite drift orbit width (FDW) effects are included for both circulating and trapped energetic particles. It is shown that, for circulating particles, FLR and FDW effects have two opposite influences on the stability of the high-n TAE modes. First, they have the usual stabilizing effects by reducing the wave particle interaction strength. Second, they also have destabilizing effects by allowing more particles to resonate with the TAE modes. It is found that the growth rate induced by the circulating alpha particles increase linearly with toroidal mode number n for small {kappa}{sub {theta}}{rho}{sub {alpha}}, and decreases as 1/n for {kappa}{sub {theta}}{rho}{sub {alpha}} {much_gt} 1. The maximum growth rate is obtained at {kappa}{sub {theta}}{rho}{sub {alpha}} on the order of unity and is nearly constant for the range of 0.7 < {upsilon}{sub {alpha}}/{upsilon}{sub A} < 2.5. On the other hand, the trapped particle response is dominated by the precessional drift resonance. The bounce resonant contribution is negligible. The growth rate peaks sharply at the value of {kappa}{sub {theta}}{rho}{sub {alpha}} such that the precessional drift resonance occurs for the most energetic trapped particles. The maximum growth rate due to the energetic trapped particles is comparable to that of circulating particles.

  19. Excitation of high-n toroidicity-induced shear Alfven eigenmodes by energetic particles and fusion alpha particles in tokamaks

    SciTech Connect

    Fu, G.Y.; Cheng, C.Z.

    1992-07-01

    The stability of high-n toroidicity-induced shear Alfven eigenmodes (TAE) in the presence of fusion alpha particles or energetic ions in tokamaks is investigated. The TAE modes are discrete in nature and thus can easily tap the free energy associated with energetic particle pressure gradient through wave particle resonant interaction. A quadratic form is derived for the high-n TAE modes using gyro-kinetic equation. The kinetic effects of energetic particles are calculated perturbatively using the ideal MHD solution as the lowest order eigenfunction. The finite Larmor radius (FLR) effects and the finite drift orbit width (FDW) effects are included for both circulating and trapped energetic particles. It is shown that, for circulating particles, FLR and FDW effects have two opposite influences on the stability of the high-n TAE modes. First, they have the usual stabilizing effects by reducing the wave particle interaction strength. Second, they also have destabilizing effects by allowing more particles to resonate with the TAE modes. It is found that the growth rate induced by the circulating alpha particles increase linearly with toroidal mode number n for small {kappa}{sub {theta}}{rho}{sub {alpha}}, and decreases as 1/n for {kappa}{sub {theta}}{rho}{sub {alpha}} {much gt} 1. The maximum growth rate is obtained at {kappa}{sub {theta}}{rho}{sub {alpha}} on the order of unity and is nearly constant for the range of 0.7 < {upsilon}{sub {alpha}}/{upsilon}{sub A} < 2.5. On the other hand, the trapped particle response is dominated by the precessional drift resonance. The bounce resonant contribution is negligible. The growth rate peaks sharply at the value of {kappa}{sub {theta}}{rho}{sub {alpha}} such that the precessional drift resonance occurs for the most energetic trapped particles. The maximum growth rate due to the energetic trapped particles is comparable to that of circulating particles.

  20. RhoE Deficiency Produces Postnatal Lethality, Profound Motor Deficits and Neurodevelopmental Delay in Mice

    PubMed Central

    Arqué, Gloria; Poch, Enric; Peris, Blanca; Guerri, Consuelo; Dierssen, Mara; Guasch, Rosa M.; Terrado, José; Pérez-Roger, Ignacio

    2011-01-01

    Rnd proteins are a subfamily of Rho GTPases involved in the control of actin cytoskeleton dynamics and other cell functions such as motility, proliferation and survival. Unlike other members of the Rho family, Rnd proteins lack GTPase activity and therefore remain constitutively active. We have recently described that RhoE/Rnd3 is expressed in the Central Nervous System and that it has a role in promoting neurite formation. Despite their possible relevance during development, the role of Rnd proteins in vivo is not known. To get insight into the in vivo function of RhoE we have generated mice lacking RhoE expression by an exon trapping cassette. RhoE null mice (RhoE gt/gt) are smaller at birth, display growth retardation and early postnatal death since only half of RhoE gt/gt mice survive beyond postnatal day (PD) 15 and 100% are dead by PD 29. RhoE gt/gt mice show an abnormal body position with profound motor impairment and impaired performance in most neurobehavioral tests. Null mutant mice are hypoactive, show an immature locomotor pattern and display a significant delay in the appearance of the hindlimb mature responses. Moreover, they perform worse than the control littermates in the wire suspension, vertical climbing and clinging, righting reflex and negative geotaxis tests. Also, RhoE ablation results in a delay of neuromuscular maturation and in a reduction in the number of spinal motor neurons. Finally, RhoE gt/gt mice lack the common peroneal nerve and, consequently, show a complete atrophy of the target muscles. This is the first model to study the in vivo functions of a member of the Rnd subfamily of proteins, revealing the important role of Rnd3/RhoE in the normal development and suggesting the possible involvement of this protein in neurological disorders. PMID:21552537

  1. Phosphorylation of myosin-binding subunit (MBS) of myosin phosphatase by Rho-kinase in vivo.

    PubMed

    Kawano, Y; Fukata, Y; Oshiro, N; Amano, M; Nakamura, T; Ito, M; Matsumura, F; Inagaki, M; Kaibuchi, K

    1999-11-29

    Rho-associated kinase (Rho-kinase), which is activated by the small GTPase Rho, phosphorylates myosin-binding subunit (MBS) of myosin phosphatase and thereby inactivates the phosphatase activity in vitro. Rho-kinase is thought to regulate the phosphorylation state of the substrates including myosin light chain (MLC), ERM (ezrin/radixin/moesin) family proteins and adducin by their direct phosphorylation and by the inactivation of myosin phosphatase. Here we identified the sites of phosphorylation of MBS by Rho-kinase as Thr-697, Ser-854 and several residues, and prepared antibody that specifically recognized MBS phosphorylated at Ser-854. We found by use of this antibody that the stimulation of MDCK epithelial cells with tetradecanoylphorbol-13-acetate (TPA) or hepatocyte growth factor (HGF) induced the phosphorylation of MBS at Ser-854 under the conditions in which membrane ruffling and cell migration were induced. Pretreatment of the cells with Botulinum C3 ADP-ribosyltransferase (C3), which is thought to interfere with Rho functions, or Rho-kinase inhibitors inhibited the TPA- or HGF-induced MBS phosphorylation. The TPA stimulation enhanced the immunoreactivity of phosphorylated MBS in the cytoplasm and membrane ruffling area of MDCK cells. In migrating MDCK cells, phosphorylated MBS as well as phosphorylated MLC at Ser-19 were localized in the leading edge and posterior region. Phosphorylated MBS was localized on actin stress fibers in REF52 fibroblasts. The microinjection of C3 or dominant negative Rho-kinase disrupted stress fibers and weakened the accumulation of phosphorylated MBS in REF52 cells. During cytokinesis, phosphorylated MBS, MLC and ERM family proteins accumulated at the cleavage furrow, and the phosphorylation level of MBS at Ser-854 was increased. Taken together, these results indicate that MBS is phosphorylated by Rho-kinase downstream of Rho in vivo, and suggest that myosin phosphatase and Rho-kinase spatiotemporally regulate the

  2. Measurements of the CKM Angle Alpha at BaBar

    SciTech Connect

    Stracka, Simone; /Milan U. /INFN, Milan

    2012-04-04

    The authors present improved measurements of the branching fractions and CP-asymmetries fin the B{sup 0} {yields} {pi}{sup +}{pi}{sup -}, B{sup 0} {yields} {pi}{sup 0}{pi}{sup 0}, and B{sup +} {yields} {rho}{sup +}{rho}{sup 0} decays, which impact the determination of {alpha}. The combined branching fractions of B {yields} K{sub 1}(1270){pi} and B {yields} K{sub 1}(1400){pi} decays are measured for the first time and allow a novel determination of {alpha} in the B{sup 0} {yields} {alpha}{sub 1}(1260){sup {+-}}{pi}{sup {-+}} decay channel. These measurements are performed using the final dataset collected by the BaBar detector at the PEP-II B-factory. The primary goal of the experiments based at the B factories is to test the Cabibbo-Kobayashi-Maskawa (CKM) picture of CP violation in the standard model of electroweak interactions. This can be achieved by measuring the angles and sides of the Unitarity Triangle in a redundant way.

  3. A mutant form of the rho protein can restore stress fibers and adhesion plaques in v-src transformed fibroblasts.

    PubMed

    Mayer, T; Meyer, M; Janning, A; Schiedel, A C; Barnekow, A

    1999-03-25

    The organization of polymerized actin in the mammalian cell is regulated by several members of the rho family. Three rho proteins, cdc42, rac and rho act in a cascade to organize the intracellular actin cytoskeleton. Rho proteins are involved in the formation of actin stress fibers and adhesion plaques in fibroblasts. During transformation of mammalian cells by oncogenes the cytoskeleton is rearranged and stress fibers and adhesion plaques are disintegrated. In this paper we investigate the function of the rho protein in RR1022 rat fibroblasts transformed by the Rous sarcoma virus. Two activated mutants of the rho protein, rho G14V and rho Q63L, and a dominant negative mutant, rho N1171, were stably transfected into RR1022 cells. The resulting cell lines were analysed for the organization of polymerized actin and adhesion plaques. Cells expressing rho Q63L, but not rho wt, rho G14V or rho N1171, showed an altered morphology. These cells displayed a flat, fibroblast like shape when compared with the RR1022 ancestor cells. Immunofluorescence analyses revealed that actin stress fibers and adhesion plaques were rearranged in these cells. We conclude from these data that an active rho protein can restore elements of the actin cytoskeleton in transformed cells by overriding the tyrosine kinase phosphorylation induced by the pp60(v-src).

  4. RNAi screens for Rho GTPase regulators of cell shape and YAP/TAZ localisation in triple negative breast cancer

    PubMed Central

    Pascual-Vargas, Patricia; Cooper, Samuel; Sero, Julia; Bousgouni, Vicky; Arias-Garcia, Mar; Bakal, Chris

    2017-01-01

    In order to metastasise, triple negative breast cancer (TNBC) must make dynamic changes in cell shape. The shape of all eukaryotic cells is regulated by Rho Guanine Nucleotide Exchange Factors (RhoGEFs), which activate Rho-family GTPases in response to mechanical and informational cues. In contrast, Rho GTPase-activating proteins (RhoGAPs) inhibit Rho GTPases. However, which RhoGEFs and RhoGAPS couple TNBC cell shape to changes in their environment is very poorly understood. Moreover, whether the activity of particular RhoGEFs and RhoGAPs become dysregulated as cells evolve the ability to metastasise is not clear. Towards the ultimate goal of identifying RhoGEFs and RhoGAPs that are essential for TNBC metastasis, we performed an RNAi screen to isolate RhoGEFs and RhoGAPs that contribute to the morphogenesis of the highly metastatic TNBC cell line LM2, and its less-metastatic parental cell line MDA-MB-231. For ~6 million cells from each cell line, we measured 127 different features following the depletion of 142 genes. Using a linear classifier scheme we also describe the morphological heterogeneity of each gene-depleted population. PMID:28248929

  5. RNAi screens for Rho GTPase regulators of cell shape and YAP/TAZ localisation in triple negative breast cancer.

    PubMed

    Pascual-Vargas, Patricia; Cooper, Samuel; Sero, Julia; Bousgouni, Vicky; Arias-Garcia, Mar; Bakal, Chris

    2017-03-01

    In order to metastasise, triple negative breast cancer (TNBC) must make dynamic changes in cell shape. The shape of all eukaryotic cells is regulated by Rho Guanine Nucleotide Exchange Factors (RhoGEFs), which activate Rho-family GTPases in response to mechanical and informational cues. In contrast, Rho GTPase-activating proteins (RhoGAPs) inhibit Rho GTPases. However, which RhoGEFs and RhoGAPS couple TNBC cell shape to changes in their environment is very poorly understood. Moreover, whether the activity of particular RhoGEFs and RhoGAPs become dysregulated as cells evolve the ability to metastasise is not clear. Towards the ultimate goal of identifying RhoGEFs and RhoGAPs that are essential for TNBC metastasis, we performed an RNAi screen to isolate RhoGEFs and RhoGAPs that contribute to the morphogenesis of the highly metastatic TNBC cell line LM2, and its less-metastatic parental cell line MDA-MB-231. For ~6 million cells from each cell line, we measured 127 different features following the depletion of 142 genes. Using a linear classifier scheme we also describe the morphological heterogeneity of each gene-depleted population.

  6. Keeping up to speed with the transcription termination factor Rho motor.

    PubMed

    Boudvillain, Marc; Nollmann, Marcello; Margeat, Emmanuel

    2010-01-01

    In bacteria, a subset of transcription termination events requires the participation of the transcription termination factor Rho. Rho is a homo-hexameric, ring-shaped, motor protein that uses the energy derived from its RNA-dependent ATPase activity to directionally unwind RNA and RNA-DNA helices and to dissociate transcription elongation complexes. Despite a wealth of structural, biochemical and genetic data, the molecular mechanisms used by Rho to carry out its biological functions remain poorly understood. Here, we briefly discuss the most recent findings on Rho mechanisms and function and highlight important questions that remain to be addressed.

  7. An RNA motif advances transcription by preventing Rho-dependent termination

    PubMed Central

    Sevostyanova, Anastasia; Groisman, Eduardo A.

    2015-01-01

    The transcription termination factor Rho associates with most nascent bacterial RNAs as they emerge from RNA polymerase. However, pharmacological inhibition of Rho derepresses only a small fraction of these transcripts. What, then, determines the specificity of Rho-dependent transcription termination? We now report the identification of a Rho-antagonizing RNA element (RARE) that hinders Rho-dependent transcription termination. We establish that RARE traps Rho in an inactive complex but does not prevent Rho binding to its recruitment sites. Although translating ribosomes normally block Rho access to an mRNA, inefficient translation of an open reading frame in the leader region of the Salmonella mgtCBR operon actually enables transcription of its associated coding region by favoring an RNA conformation that sequesters RARE. The discovery of an RNA element that inactivates Rho signifies that the specificity of nucleic-acid binding proteins is defined not only by the sequences that recruit these proteins but also by sequences that antagonize their activity. PMID:26630006

  8. [Rho GTPases as therapeutic targets in cancer and other human diseases].

    PubMed

    Lorenzano Menna, Pablo; Cardama, Georgina A; Comin, María J; Alonso, Daniel F; Gómez, Daniel E

    2010-01-01

    Rho GTPases are a key protein family controlling the transduction of external signals to cytoplasmatic and nuclear effectors. In the last few years, the development of genetic and pharmacological tools has allowed a more precise definition of the specific roles of Rho GTPases. The aim of this review is to describe the cellular functions regulated by these proteins with focus on the molecular mechanism involved. We also address the role of Rho GTPases in the development of different human diseases such as cancer. Finally, we describe different experimental therapeutic strategies with Rho GTPases as molecular targets.

  9. Candida albicans RHO1 is required for cell viability in vitro and in vivo.

    PubMed

    Smith, Susan E; Csank, Csilla; Reyes, Guadalupe; Ghannoum, Mahmoud A; Berlin, Vivian

    2002-05-01

    In Saccharomyces cerevisiae, Rho1p plays an important role in cell wall integrity by regulating beta-1,3-glucan synthase, Pkc1p and the actin cytoskeleton. To determine the physiological role of Rho1p in the dimorphic fungus Candida albicans, the major human fungal pathogen, we constructed mutants that conditionally express Rho1p from the glucose-repressible phosphoenolpyruvate carboxykinase promoter (pPCK1). We examined the growth of these cells in a range of conditions. Depletion of Rho1p from yeast cells resulted in cell death, lysis, and aggregation. The Rho1p conditional mutant was inviable on 10% serum indicating that Rho1p was also required for hyphal viability. Furthermore, in a mouse model of systemic candidiasis, strains dependent on pPCK1-driven RHO1 expression failed to colonise the kidneys and establish disease, suggesting that the level of glucose in serum was sufficient to repress the pPCK1 and that Rho1p-depleted strains were inviable within the host. Therefore, Rho1p is essential for the viability of C. albicans in vitro and in vivo.

  10. The Rho GTP exchange factor Lfc promotes spindle assembly in early mitosis

    PubMed Central

    Bakal, Christopher J.; Finan, Dina; LaRose, José; Wells, Clark D.; Gish, Gerald; Kulkarni, Sarang; DeSepulveda, Paulo; Wilde, Andrew; Rottapel, Robert

    2005-01-01

    Rho GTPases regulate reorganization of actin and microtubule cytoskeletal structures during both interphase and mitosis. The timing and subcellular compartment in which Rho GTPases are activated is controlled by the large family of Rho GTP exchange factors (RhoGEFs). Here, we show that the microtubule-associated RhoGEF Lfc is required for the formation of the mitotic spindle during prophase/prometaphase. The inability of cells to assemble a functioning spindle after Lfc inhibition resulted in a delay in mitosis and an accumulation of prometaphase cells. Inhibition of Lfc's primary target Rho GTPase during prophase/prometaphase, or expression of a catalytically inactive mutant of Lfc, also prevented normal spindle assembly and resulted in delays in mitotic progression. Coinjection of constitutively active Rho GTPase rescued the spindle defects caused by Lfc inhibition, suggesting the requirement of RhoGTP in regulating spindle assembly. Lastly, we implicate mDia1 as an important effector of Lfc signaling. These findings demonstrate a role for Lfc, Rho, and mDia1 during mitosis. PMID:15976019

  11. Rho-kinase as a novel therapeutic target in treatment of cardiovascular diseases.

    PubMed

    Shimokawa, Hiroaki

    2002-03-01

    Rho-kinase has been identified as one of the effectors of the small GTP-binding protein Rho. Accumulating evidence has demonstrated that the Rho/Rho-kinase-mediated pathway plays an important role in various cellular functions, not only in vascular smooth muscle contraction but also in actin cytoskeleton organization, cell adhesion and motility, cytokinesis, and gene expressions, all of which may be involved in the pathogenesis of arteriosclerosis/atherosclerosis. Indeed, animal experiments have demonstrated that Rho-kinase inhibitors effectively suppress coronary artery spasm and that long-term inhibition of Rho-kinase inhibits the development of coronary arteriosclerotic lesions and even causes regression of coronary vascular lesions in vivo. Recent clinical studies also have demonstrated the inhibitory effect of a Rho-kinase inhibitor on coronary artery spasm in patients with vasospastic angina and on exercise-induced myocardial ischemia in patients with stable effort angina with adequate safety. It is possible that Rho-kinase is also involved in the pathogenesis of other forms of cardiovascular diseases. Thus, Rho-kinase could be regarded as a novel therapeutic target in treatment of cardiovascular diseases.

  12. The use of GFP to localize Rho GTPases in living cells.

    PubMed

    Michaelson, David; Philips, Mark

    2006-01-01

    The green fluorescent protein (GFP) of the jellyfish Aequorea victoria has revolutionized the study of protein localization and dynamics. GFP fusions permit analysis of proteins in living cells and offer distinct advantages over conventional immunofluorescence. Among these are lower background, higher resolution, robust dual color colocalization, and avoidance of fixation artifacts. In the case of Ras and Rho family proteins, GFP fusions have allowed breakthroughs in the understanding of how CAAX proteins are targeted to specific cell membranes and how signaling at different membranes can result in different cellular responses. GFP-tagged Rho proteins have also been informative in analyzing the interactions with the cytosolic chaperone, RhoGDI. The major disadvantages of studying GFP fusion proteins is that they are generally overexpressed relative to endogenous proteins, and the GFP tag can, in principle, affect protein function. Fortunately, in the case of Ras and Rho family proteins, a GFP tag at the N terminus seems to have little effect on protein targeting and function. Nevertheless, it is prudent to confirm GFP fusion protein data with the study of the endogenous protein. This chapter describes the tagging of Rho proteins with GFP and the analysis of GFP-Rho protein localization by epifluorescence and confocal microscopy. It further describes methods of analyzing endogenous Rho proteins as confirmation of data acquired using GFP-Rho fusion proteins. These techniques will be useful for anyone studying Rho protein function and are widely applicable to many cell types and signal transduction systems.

  13. RhoA GTPase controls cytokinesis and programmed necrosis of hematopoietic progenitors

    PubMed Central

    Zhou, Xuan; Florian, Maria Carolina; Arumugam, Paritha; Chen, Xiaoyi; Cancelas, Jose A.; Lang, Richard; Malik, Punam; Geiger, Hartmut

    2013-01-01

    Hematopoietic progenitor cells (HPCs) are central to hematopoiesis as they provide large numbers of lineage-defined blood cells necessary to sustain blood homeostasis. They are one of the most actively cycling somatic cells, and their precise control is critical for hematopoietic homeostasis. The small GTPase RhoA is an intracellular molecular switch that integrates cytokine, chemokine, and adhesion signals to coordinate multiple context-dependent cellular processes. By using a RhoA conditional knockout mouse model, we show that RhoA deficiency causes a multilineage hematopoietic failure that is associated with defective multipotent HPCs. Interestingly, RhoA−/− hematopoietic stem cells retained long-term engraftment potential but failed to produce multipotent HPCs and lineage-defined blood cells. This multilineage hematopoietic failure was rescued by reconstituting wild-type RhoA into the RhoA−/− Lin−Sca-1+c-Kit+ compartment. Mechanistically, RhoA regulates actomyosin signaling, cytokinesis, and programmed necrosis of the HPCs, and loss of RhoA results in a cytokinesis failure of HPCs manifested by an accumulation of multinucleated cells caused by failed abscission of the cleavage furrow after telophase. Concomitantly, the HPCs show a drastically increased death associated with increased TNF–RIP-mediated necrosis. These results show that RhoA is a critical and specific regulator of multipotent HPCs during cytokinesis and thus essential for multilineage hematopoiesis. PMID:24101377

  14. Local RhoA activation induces cytokinetic furrows independent of spindle position and cell cycle stage

    PubMed Central

    2016-01-01

    The GTPase RhoA promotes contractile ring assembly and furrow ingression during cytokinesis. Although many factors that regulate RhoA during cytokinesis have been characterized, the spatiotemporal regulatory logic remains undefined. We have developed an optogenetic probe to gain tight spatial and temporal control of RhoA activity in mammalian cells and demonstrate that cytokinetic furrowing is primarily regulated at the level of RhoA activation. Light-mediated recruitment of a RhoGEF domain to the plasma membrane leads to rapid induction of RhoA activity, leading to assembly of cytokinetic furrows that partially ingress. Furthermore, furrow formation in response to RhoA activation is not temporally or spatially restricted. RhoA activation is sufficient to generate furrows at both the cell equator and cell poles, in both metaphase and anaphase. Remarkably, furrow formation can be initiated in rounded interphase cells, but not adherent cells. These results indicate that RhoA activation is sufficient to induce assembly of functional contractile rings and that cell rounding facilitates furrow formation. PMID:27298323

  15. The GTPase Rho has a critical regulatory role in thymus development.

    PubMed Central

    Henning, S W; Galandrini, R; Hall, A; Cantrell, D A

    1997-01-01

    The present study employs a genetic approach to explore the role of Rho GTPases in murine thymic development. Inactivation of Rho function in the thymus was achieved by thymic targeting of a transgene encoding C3 transferase from Clostridium botulinum which selectively ADP-ribosylates Rho within its effector domain and thereby abolishes its biological function. Thymi lacking functional Rho isolated from C3 transgenic mice were strikingly smaller and showed a marked (90%) decrease in cellularity compared with their normal litter mates. We also observed a similar decrease in levels of peripheral T cells in C3 transgenic mice. Analysis of the maturation status of thymocytes indicated that differentiation of progenitor cells to mature T cells can occur in the absence of Rho function, and both positive and negative selection of T cells appear to be intact. However, transgenic mice that lack Rho function in the thymus show maturational, proliferative and cell survival defects during T-cell development that severely impair the generation of normal numbers of thymocytes and mature peripheral T cells. The present study thus identifies a role for Rho-dependent signalling pathways in thymocyte development. The data show that the function of Rho GTPases is critical for the proliferative expansion of thymocytes. This defines a selective role for the GTPase Rho in early thymic development as a critical integrator of proliferation and cell survival signals. PMID:9171353

  16. Coxiella burnetii Phagocytosis Is Regulated by GTPases of the Rho Family and the RhoA Effectors mDia1 and ROCK

    PubMed Central

    Distel, Jesús S.; Aguilera, Milton O.; Colombo, María I.; Berón, Walter

    2015-01-01

    The GTPases belonging to the Rho family control the actin cytoskeleton rearrangements needed for particle internalization during phagocytosis. ROCK and mDia1 are downstream effectors of RhoA, a GTPase involved in that process. Coxiella burnetii, the etiologic agent of Q fever, is internalized by the host´s cells in an actin-dependent manner. Nevertheless, the molecular mechanism involved in this process has been poorly characterized. This work analyzes the role of different GTPases of the Rho family and some downstream effectors in the internalization of C. burnetii by phagocytic and non-phagocytic cells. The internalization of C. burnetii into HeLa and RAW cells was significantly inhibited when the cells were treated with Clostridium difficile Toxin B which irreversibly inactivates members of the Rho family. In addition, the internalization was reduced in HeLa cells that overexpressed the dominant negative mutants of RhoA, Rac1 or Cdc42 or that were knocked down for the Rho GTPases. The pharmacological inhibition or the knocking down of ROCK diminished bacterium internalization. Moreover, C. burnetii was less efficiently internalized in HeLa cells overexpressing mDia1-N1, a dominant negative mutant of mDia1, while the overexpression of the constitutively active mutant mDia1-ΔN3 increased bacteria uptake. Interestingly, when HeLa and RAW cells were infected, RhoA, Rac1 and mDia1 were recruited to membrane cell fractions. Our results suggest that the GTPases of the Rho family play an important role in C. burnetii phagocytosis in both HeLa and RAW cells. Additionally, we present evidence that ROCK and mDia1, which are downstream effectors of RhoA, are involved in that process. PMID:26674774

  17. Effect of electroacupuncture on the mRNA and protein expression of Rho-A and Rho-associated kinase II in spinal cord injury rats

    PubMed Central

    Min, You-jiang; Ding, Li-li-qiang; Cheng, Li-hong; Xiao, Wei-ping; He, Xing-wei; Zhang, Hui; Min, Zhi-yun; Pei, Jia

    2017-01-01

    Electroacupuncture is beneficial for the recovery of spinal cord injury, but the underlying mechanism is unclear. The Rho/Rho-associated kinase (ROCK) signaling pathway regulates the actin cytoskeleton by controlling the adhesive and migratory behaviors of cells that could inhibit neurite regrowth after neural injury and consequently hinder the recovery from spinal cord injury. Therefore, we hypothesized electroacupuncture could affect the Rho/ROCK signaling pathway to promote the recovery of spinal cord injury. In our experiments, the spinal cord injury in adult Sprague-Dawley rats was caused by an impact device. Those rats were subjected to electroacupuncture at Yaoyangguan (GV3), Dazhui (GV14), Zusanli (ST36) and Ciliao (BL32) and/or monosialoganglioside treatment. Behavioral scores revealed that the hindlimb motor functions improved with those treatments. Real-time quantitative polymerase chain reaction, fluorescence in situ hybridization and western blot assay showed that electroacupuncture suppressed the mRNA and protein expression of Rho-A and Rho-associated kinase II (ROCKII) of injured spinal cord. Although monosialoganglioside promoted the recovery of hindlimb motor function, monosialoganglioside did not affect the expression of Rho-A and ROCKII. However, electroacupuncture combined with monosialoganglioside did not further improve the motor function or suppress the expression of Rho-A and ROCKII. Our data suggested that the electroacupuncture could specifically inhibit the activation of the Rho/ROCK signaling pathway thus partially contributing to the repair of injured spinal cord. Monosialoganglioside could promote the motor function but did not suppress expression of RhoA and ROCKII. There was no synergistic effect of electroacupuncture combined with monosialoganglioside.

  18. trans-4-Amino-2-methylbut-2-enoic acid (2-MeTACA) and (+/-)-trans-2-aminomethylcyclopropanecarboxylic acid ((+/-)-TAMP) can differentiate rat rho3 from human rho1 and rho2 recombinant GABA(C) receptors.

    PubMed

    Vien, Jimmy; Duke, Rujee K; Mewett, Kenneth N; Johnston, Graham A R; Shingai, Ryuzo; Chebib, Mary

    2002-02-01

    1. This study investigated the effects of a number of GABA analogues on rat rho3 GABA(C) receptors expressed in Xenopus oocytes using 2-electrode voltage clamp methods. 2. The potency order of agonists was muscimol (EC(50)=1.9 +/- 0.1 microM) (+)-trans-3-aminocyclopentanecarboxylic acids ((+)-TACP; EC(50)=2.7 +/- 0.9 microM) trans-4-aminocrotonic acid (TACA; EC(50)=3.8 +/-0.3 microM) GABA (EC(50)=4.0 +/- 0.3 microM) > thiomuscimol (EC(50)=24.8 +/- 2.6 microM) > (+/-)-cis-2-aminomethylcyclopropane-carboxylic acid ((+/-)-CAMP; EC(50)=52.6 +/-8.7 microM) > cis-4-aminocrotonic acid (CACA; EC(50)=139.4 +/- 5.2 microM). 3. The potency order of antagonists was (+/-)-trans-2-aminomethylcyclopropanecarboxylic acid ((+/-)-TAMP; K(B)=4.8+/-1.8 microM) (1,2,5,6-tetrahydropyridin-4-yl)methylphosphinic acid (TPMPA; K(B)=4.8 +/-0.8 microM) > (piperidin-4-yl)methylphosphinic acid (P4MPA; K(B)=10.2+/-2.3 microM) 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP; K(B)=10.2+/-0.3 microM) imidazole-4-acetic acid (I4AA; K(B)=12.6+/-2.7 microM) > 3-aminopropylphosphonic acid (3-APA; K(B)=35.8+/-13.5 microM). 4. trans-4-Amino-2-methylbut-2-enoic acid (2-MeTACA; 300 microM) had no effect as an agonist or an antagonist indicating that the C2 methyl substituent is sterically interacting with the ligand-binding site of rat rho3 GABA(C) receptors. 5. 2-MeTACA affects rho1 and rho2 but not rho3 GABA(C) receptors. In contrast, (plus minus)-TAMP is a partial agonist at rho1 and rho2 GABA(C) receptors, while at rat rho3 GABA(C) receptors it is an antagonist. Thus, 2-MeTACA and (+/-)-TAMP could be important pharmacological tools because they may functionally differentiate between rho1, rho2 and rho3 GABA(C) receptors in vitro.

  19. Mechanism of RhoB/FTI Action in Breast Cancer

    DTIC Science & Technology

    2004-05-01

    Cancer Research describing work on cyclin B1 as a proapoptotic effector target for RhoB signaling (Kamasani, U., Huang, M., DuHadaway, J ., Prochownik...disease: a potential new marker for small breast carcinomas with metastatic ability. Am. J . Pathol. 2002; 160: 579-584. 3. Liu, A-X, Rane, N, Liu, J -P...USA 2001; 98: 6192-6197. 16a. Kamasani, U., Huang, M., DuHadaway, J ., Prochownik, E.V., Donover, P.S. and Prendergast, G.C. Cyclin B1 is a critical

  20. Riboswitches in regulation of Rho-dependent transcription termination.

    PubMed

    Proshkin, Sergey; Mironov, Alexander; Nudler, Evgeny

    2014-10-01

    Riboswitches are RNA sensors of small metabolites and ions that regulate gene expression in response to environmental changes. In bacteria, the riboswitch sensor domain usually controls the formation of a strong RNA hairpin that either functions as a potent transcription terminator or sequesters a ribosome-binding site. A recent study demonstrated a novel mechanism by which a riboswitch controls Rho-dependent transcription termination. This riboswitch mechanism is likely a widespread mode of gene regulation that determines whether a protein effector is able to attenuate transcription. This article is part of a Special Issue entitled: Riboswitches.

  1. Rho, ROCK and actomyosin contractility in metastasis as drug targets

    PubMed Central

    Bruce, Fanshawe; Sanz-Moreno, Victoria

    2016-01-01

    Metastasis is the spread of cancer cells around the body and the cause of the majority of cancer deaths. Metastasis is a very complex process in which cancer cells need to dramatically modify their cytoskeleton and cope with different environments to successfully colonize a secondary organ. In this review, we discuss recent findings pointing at Rho-ROCK or actomyosin force (or both) as major drivers of many of the steps required for metastatic success. We propose that these are important drug targets that need to be considered in the clinic to palliate metastatic disease. PMID:27158478

  2. The RhoA-ROCK-PTEN pathway as a molecular switch for anchorage dependent cell behavior.

    PubMed

    Yang, Seungwon; Kim, Hyun-Man

    2012-04-01

    The proliferation of anchorage-dependent cells of mesenchymal origin requires the attachment of the cells to substrates. Thus, cells that are poorly attached to substrates exhibit retarded cell cycle progression or apoptotic death. A major disadvantage of most polymers used in tissue engineering is their hydrophobicity; hydrophobic surfaces do not allow cells to attach firmly and, therefore, do not allow normal proliferation rates. In this study, we investigated the molecular mechanism underlying the reduced proliferation rate of cells that are poorly attached to substrates. There was an inverse relationship between the activity of the small GTPase RhoA (RhoA) and the cell proliferation rate. RhoA activity correlated inversely with the strength of cell adhesion to the substrates. The high RhoA activity in the cells poorly attached to substrates caused an increase in the activity of Rho-associated kinase (ROCK), a well-known effector of RhoA that upregulated the activity of phosphatase and tensin homolog (PTEN). The resulting activated PTEN downregulated Akt activity, which is essential for cell proliferation. Thus, the cells that were poorly attached to substrates showed low levels of cell proliferation because the RhoA-ROCK-PTEN pathway was hyperactive. In addition, RhoA activity seemed to be related to focal adhesion kinase (FAK) activity. Weak FAK activity in these poorly attached cells failed to downregulate the high RhoA activity that restrained cell proliferation. Interestingly, reducing the expression of any component of the RhoA-ROCK-PTEN pathway rescued the proliferation rate without physico-chemical surface modifications. Based on these results, we suggest that the RhoA-ROCK-PTEN pathway acts as a molecular switch to control cell proliferation and determine anchorage dependence. In cells that are poorly attached to substrates, its inhibition is sufficient to restore cell proliferation without the need for physico-chemical modification of the material

  3. Role of Rho kinase signalling in healthy and varicose human saphenous veins

    PubMed Central

    Cario-Toumaniantz, Chrystelle; Evellin, Sandrine; Maury, Séverine; Baron, Olivier; Pacaud, Pierre; Loirand, Gervaise

    2002-01-01

    The present study was performed to determine the role of Rho-Rho kinase signalling pathway in smooth muscle cells from both healthy and varicose human saphenous vein. The Rho kinase inhibitor Y-27632 inhibited the noradrenaline (NA)-induced contraction in human saphenous veins with IC50 corresponding to 0.5 μM and 10.9 μM in control and varicose veins, respectively. The maximal amplitude of the NA-induced contraction was smaller in varicose vein compared to control (1263±172 mg versus 1974±245 mg, P<0.05). In β-escin permeabilized strips, GTPγS induced a rise in tension that was inhibited by Y-27632. The amplitude of the GTPγS-induced contraction was smaller in varicose compared to control veins (23.1±2.4% versus 41.3±2.2%, P<0.002). In smooth muscle cells, Y-27632 induced disassembly of both actin cytoskeleton and extracellular fibronectin matrix. In comparison to control cells, varicose vein smooth muscle cells show decreased actin cytoskeleton organization and reduction of fibronectin matrix deposition. The Rho proteins Rnd1 and RhoA, and Rho kinase 1 are expressed in human saphenous veins. A 2.6 fold reduction of Rho kinase expression was found in varicose veins. These results indicate that RhoA-Rho kinase mediated Ca2+ sensitization of the contraction and regulated actin cytoskeleton and extracellular fibronectin matrix assembly in human saphenous smooth muscle. The decrease of Rho kinase expression and Rho kinase-dependent functions detected in smooth muscle from varicose veins supports a role of this signalling pathway in the functional alterations of the vein wall occurring in the course of the disease. PMID:12208777

  4. RhoA is dispensable for skin development, but crucial for contraction and directed migration of keratinocytes.

    PubMed

    Jackson, Ben; Peyrollier, Karine; Pedersen, Esben; Basse, Astrid; Karlsson, Richard; Wang, Zhipeng; Lefever, Tine; Ochsenbein, Alexandra M; Schmidt, Gudula; Aktories, Klaus; Stanley, Alanna; Quondamatteo, Fabio; Ladwein, Markus; Rottner, Klemens; van Hengel, Jolanda; Brakebusch, Cord

    2011-03-01

    RhoA is a small guanosine-5'-triphosphatase (GTPase) suggested to be essential for cytokinesis, stress fiber formation, and epithelial cell-cell contacts. In skin, loss of RhoA was suggested to underlie pemphigus skin blistering. To analyze RhoA function in vivo, we generated mice with a keratinocyte-restricted deletion of the RhoA gene. Despite a severe reduction of cofilin and myosin light chain (MLC) phosphorylation, these mice showed normal skin development. Primary RhoA-null keratinocytes, however, displayed an increased percentage of multinucleated cells, defective maturation of cell-cell contacts. Furthermore we observed increased cell spreading due to impaired RhoA-ROCK (Rho-associated protein kinase)-MLC phosphatase-MLC-mediated cell contraction, independent of Rac1. Rho-inhibiting toxins further increased multinucleation of RhoA-null cells but had no significant effect on spreading, suggesting that RhoB and RhoC have partially overlapping functions with RhoA. Loss of RhoA decreased directed cell migration in vitro caused by reduced migration speed and directional persistence. These defects were not related to the decreased cell contraction and were independent of ROCK, as ROCK inhibition by Y27632 increased directed migration of both control and RhoA-null keratinocytes. Our data indicate a crucial role for RhoA and contraction in regulating cell spreading and a contraction-independent function of RhoA in keratinocyte migration. In addition, our data show that RhoA is dispensable for skin development.

  5. RNA secondary structures regulate three steps of Rho-dependent transcription termination within a bacterial mRNA leader.

    PubMed

    Kriner, Michelle A; Groisman, Eduardo A

    2017-01-25

    Transcription termination events in bacteria often require the RNA helicase Rho. Typically, Rho promotes termination at the end of coding sequences, but it can also terminate transcription within leader regions to implement regulatory decisions. Rho-dependent termination requires initial recognition of a Rho utilization (rut) site on a nascent RNA by Rho's primary binding surface. However, it is presently unclear what factors determine the location of transcription termination, how RNA secondary structures influence this process and whether mechanistic differences distinguish constitutive from regulated Rho-dependent terminators. We previously demonstrated that the 5' leader mRNA of the Salmonella corA gene can adopt two mutually exclusive conformations that dictate accessibility of a rut site to Rho. We now report that the corA leader also controls two subsequent steps of Rho-dependent termination. First, the RNA conformation that presents an accessible rut site promotes pausing of RNA polymerase (RNAP) at a single Rho-dependent termination site over 100 nt downstream. Second, an additional RNA stem-loop promotes Rho activity and controls the location at which Rho-dependent termination occurs, despite having no effect on initial Rho binding to the corA leader. Thus, the multi-step nature of Rho-dependent termination may facilitate regulation of a given coding region by multiple cytoplasmic signals.

  6. RNA secondary structures regulate three steps of Rho-dependent transcription termination within a bacterial mRNA leader

    PubMed Central

    Kriner, Michelle A.; Groisman, Eduardo A.

    2017-01-01

    Transcription termination events in bacteria often require the RNA helicase Rho. Typically, Rho promotes termination at the end of coding sequences, but it can also terminate transcription within leader regions to implement regulatory decisions. Rho-dependent termination requires initial recognition of a Rho utilization (rut) site on a nascent RNA by Rho's primary binding surface. However, it is presently unclear what factors determine the location of transcription termination, how RNA secondary structures influence this process and whether mechanistic differences distinguish constitutive from regulated Rho-dependent terminators. We previously demonstrated that the 5′ leader mRNA of the Salmonella corA gene can adopt two mutually exclusive conformations that dictate accessibility of a rut site to Rho. We now report that the corA leader also controls two subsequent steps of Rho-dependent termination. First, the RNA conformation that presents an accessible rut site promotes pausing of RNA polymerase (RNAP) at a single Rho-dependent termination site over 100 nt downstream. Second, an additional RNA stem-loop promotes Rho activity and controls the location at which Rho-dependent termination occurs, despite having no effect on initial Rho binding to the corA leader. Thus, the multi-step nature of Rho-dependent termination may facilitate regulation of a given coding region by multiple cytoplasmic signals. PMID:28123036

  7. Rho-kinase mediated cytoskeletal stiffness in skinned smooth muscle

    PubMed Central

    Lan, Bo; Wang, Lu; Zhang, Jenny; Pascoe, Chris D.; Norris, Brandon A.; Liu, Jeffrey C.-Y.; Solomon, Dennis; Paré, Peter D.; Deng, Linhong

    2013-01-01

    The structurally dynamic cytoskeleton is important in many cell functions. Large gaps still exist in our knowledge regarding what regulates cytoskeletal dynamics and what underlies the structural plasticity. Because Rho-kinase is an upstream regulator of signaling events leading to phosphorylation of many cytoskeletal proteins in many cell types, we have chosen this kinase as the focus of the present study. In detergent skinned tracheal smooth muscle preparations, we quantified the proteins eluted from the muscle cells over time and monitored the muscle's ability to respond to acetylcholine (ACh) stimulation to produce force and stiffness. In a partially skinned preparation not able to generate active force but could still stiffen upon ACh stimulation, we found that the ACh-induced stiffness was independent of calcium and myosin light chain phosphorylation. This indicates that the myosin light chain-dependent actively cycling crossbridges are not likely the source of the stiffness. The results also indicate that Rho-kinase is central to the ACh-induced stiffness, because inhibition of the kinase by H1152 (1 μM) abolished the stiffening. Furthermore, the rate of relaxation of calcium-induced stiffness in the skinned preparation was faster than that of ACh-induced stiffness, with or without calcium, suggesting that different signaling pathways lead to different means of maintenance of stiffness in the skinned preparation. PMID:24072407

  8. Fasudil alleviates traumatic optic neuropathy by inhibiting Rho signaling pathway

    PubMed Central

    Yu, Jianglong; Lan, Shiying; Wang, Ruijia; Maier, Aba; Luan, Xinping

    2015-01-01

    Objectives: The present study is to investigate the pathological changes in rabbits with traumatic optic neuropathy (TON), as well as the effect of fasudil on the lesions. Methods: A total of 144 New Zealand rabbits were successfully established as TON models. Twelve hours after surgery, the rabbits in control, dexamethasone, and fasudil groups were administrated with saline, dexamethasone, and fasudil via ear veins, respectively. Then, retinas of the rabbits were obtained at 72 h and on days 7, 14 and 21 after surgery. The pathological changes in retina and optic nerves were observed by hematoxylin and eosin staining and transmission electron microscopy. The expression levels of Rho-associated genes were measured using quantitative real-time polymerase chain reaction. Results: In control group, the axons were swelling, and mitochondria showed vacuolation after optic nerve crush. Mitochondria were swelled slightly in dexamethasone group. By contrast, nerves in fasudil group were repaired. Retinal ganglion cells in control group were reduced significantly due to optic nerve crush. The loss of retinal ganglion cells was alleviated in fasudil group. Quantitative real-time polymerase chain reaction showed that the expression of Rho-associated genes were down-regulated. Conclusions: The present study demonstrates that fasudil inhibits the apoptosis of retinal ganglion cells and ameliorates damages of optic nerves in traumatic optic neuropathy. PMID:26550269

  9. A molecular ruler regulates cytoskeletal remodelling by the Rho kinases

    PubMed Central

    Truebestein, Linda; Elsner, Daniel J.; Fuchs, Elisabeth; Leonard, Thomas A.

    2015-01-01

    The Rho-associated coiled-coil kinases (ROCK) are essential regulators of the actin cytoskeleton; however, the structure of a full-length ROCK is unknown and the mechanisms by which its kinase activity is controlled are not well understood. Here we determine the low-resolution structure of human ROCK2 using electron microscopy, revealing it to be a constitutive dimer, 120 nm in length, with a long coiled-coil tether linking the kinase and membrane-binding domains. We find, in contrast to previous reports, that ROCK2 activity does not appear to be directly regulated by binding to membranes, RhoA, or by phosphorylation. Instead, we show that changing the length of the tether modulates ROCK2 function in cells, suggesting that it acts as a molecular ruler. We present a model in which ROCK activity is restricted to a discrete region of the actin cytoskeleton, governed by the length of its coiled-coil. This represents a new type of spatial control, and hence a new paradigm for kinase regulation. PMID:26620183

  10. Ab initio alpha-alpha scattering.

    PubMed

    Elhatisari, Serdar; Lee, Dean; Rupak, Gautam; Epelbaum, Evgeny; Krebs, Hermann; Lähde, Timo A; Luu, Thomas; Meißner, Ulf-G

    2015-12-03

    Processes such as the scattering of alpha particles ((4)He), the triple-alpha reaction, and alpha capture play a major role in stellar nucleosynthesis. In particular, alpha capture on carbon determines the ratio of carbon to oxygen during helium burning, and affects subsequent carbon, neon, oxygen, and silicon burning stages. It also substantially affects models of thermonuclear type Ia supernovae, owing to carbon detonation in accreting carbon-oxygen white-dwarf stars. In these reactions, the accurate calculation of the elastic scattering of alpha particles and alpha-like nuclei--nuclei with even and equal numbers of protons and neutrons--is important for understanding background and resonant scattering contributions. First-principles calculations of processes involving alpha particles and alpha-like nuclei have so far been impractical, owing to the exponential growth of the number of computational operations with the number of particles. Here we describe an ab initio calculation of alpha-alpha scattering that uses lattice Monte Carlo simulations. We use lattice effective field theory to describe the low-energy interactions of protons and neutrons, and apply a technique called the 'adiabatic projection method' to reduce the eight-body system to a two-cluster system. We take advantage of the computational efficiency and the more favourable scaling with system size of auxiliary-field Monte Carlo simulations to compute an ab initio effective Hamiltonian for the two clusters. We find promising agreement between lattice results and experimental phase shifts for s-wave and d-wave scattering. The approximately quadratic scaling of computational operations with particle number suggests that it should be possible to compute alpha scattering and capture on carbon and oxygen in the near future. The methods described here can be applied to ultracold atomic few-body systems as well as to hadronic systems using lattice quantum chromodynamics to describe the interactions of

  11. Ab initio alpha-alpha scattering

    NASA Astrophysics Data System (ADS)

    Elhatisari, Serdar; Lee, Dean; Rupak, Gautam; Epelbaum, Evgeny; Krebs, Hermann; Lähde, Timo A.; Luu, Thomas; Meißner, Ulf-G.

    2015-12-01

    Processes such as the scattering of alpha particles (4He), the triple-alpha reaction, and alpha capture play a major role in stellar nucleosynthesis. In particular, alpha capture on carbon determines the ratio of carbon to oxygen during helium burning, and affects subsequent carbon, neon, oxygen, and silicon burning stages. It also substantially affects models of thermonuclear type Ia supernovae, owing to carbon detonation in accreting carbon-oxygen white-dwarf stars. In these reactions, the accurate calculation of the elastic scattering of alpha particles and alpha-like nuclei—nuclei with even and equal numbers of protons and neutrons—is important for understanding background and resonant scattering contributions. First-principles calculations of processes involving alpha particles and alpha-like nuclei have so far been impractical, owing to the exponential growth of the number of computational operations with the number of particles. Here we describe an ab initio calculation of alpha-alpha scattering that uses lattice Monte Carlo simulations. We use lattice effective field theory to describe the low-energy interactions of protons and neutrons, and apply a technique called the ‘adiabatic projection method’ to reduce the eight-body system to a two-cluster system. We take advantage of the computational efficiency and the more favourable scaling with system size of auxiliary-field Monte Carlo simulations to compute an ab initio effective Hamiltonian for the two clusters. We find promising agreement between lattice results and experimental phase shifts for s-wave and d-wave scattering. The approximately quadratic scaling of computational operations with particle number suggests that it should be possible to compute alpha scattering and capture on carbon and oxygen in the near future. The methods described here can be applied to ultracold atomic few-body systems as well as to hadronic systems using lattice quantum chromodynamics to describe the interactions of

  12. RhoB/ROCK mediates oxygen-glucose deprivation-stimulated syncytiotrophoblast microparticle shedding in preeclampsia.

    PubMed

    Han, Jian; Yang, Bo-Ping; Li, Yi-Lin; Li, Hong-Mei; Zheng, Xiu-Hui; Yu, Li-Li; Zhang, Qiong; Zheng, Ying-Ru; Yi, Ping; Li, Li; Guo, Jian-Xin; Zhou, Yuan-Guo

    2016-11-01

    Increased circulating syncytiotrophoblast microparticles (STBMs) are often associated with preeclampsia (PE) but the molecular mechanisms regulating STBM shedding remain elusive. Experimental evidence has shown that actin plays a key role in STBM shedding and that Rho/ROCK is important in regulating actin rearrangement. To investigate the role of RhoB/ROCK-regulated actin arrangement in STBM shedding in PE, chorionic villous explants were prepared from placenta of patients with normotensive or PE pregnancies and BeWo cells were fused to imitate syncytiotrophoblasts. The oxygen-glucose deprivation (OGD) conditions were applied to imitate the pathophysiology of PE in vitro. The results showed that RhoB and ROCK were activated in the preeclamptic placenta, accompanied by increased actin polymerization and decreased outgrowing microvilli. In villous tissue cultures or BeWo cells, OGD activated RhoB, ROCK1 and ROCK2 and promoted STBM shedding and actin stress fibers formation. In BeWo cells, RhoB overexpression activated ROCK1 and ROCK2, leading to F-actin redistribution and STBM shedding and the OGD-induced actin polymerization and STBM shedding could be reversed by RhoB or ROCK knockdown. These results reveal that RhoB and ROCK play a key role in PE by targeting STBM shedding through actin rearrangement and that RhoB/ROCK intervention may be a potential therapeutic strategy for PE.

  13. Revisited and Revised: Is RhoA Always a Villain in Cardiac Pathophysiology?

    PubMed Central

    Miyamoto, Shigeki; Del Re, Dominic P.; Xiang, Sunny Y.; Zhao, Xia; Florholmen, Geir

    2010-01-01

    The neonatal rat ventricular myocyte model of hypertrophy has provided tremendous insight with regard to signaling pathways regulating cardiac growth and gene expression. Many mediators thus discovered have been successfully extrapolated to the in vivo setting, as assessed using genetically engineered mice and physiological interventions. Studies in neonatal rat ventricular myocytes demonstrated a role for the small G-protein RhoA and its downstream effector kinase, Rho-associated coiled-coil containing protein kinase (ROCK), in agonist-mediated hypertrophy. Transgenic expression of RhoA in the heart does not phenocopy this response, however, nor does genetic deletion of ROCK prevent hypertrophy. Pharmacologic inhibition of ROCK has effects most consistent with roles for RhoA signaling in the development of heart failure or responses to ischemic damage. Whether signals elicited downstream of RhoA promote cell death or survival and are deleterious or salutary is, however, context and cell-type dependent. The concepts discussed above are reviewed, and the hypothesis that RhoA might protect cardiomyocytes from ischemia and other insults is presented. Novel RhoA targets including phospholipid regulated and regulating enzymes (Akt, PI kinases, phospholipase C, protein kinases C and D) and serum response element-mediated transcriptional responses are considered as possible pathways through which RhoA could affect cardiomyocyte survival. PMID:20559774

  14. Empirical Size, Coverage, and Power of Confidence Intervals for Spearman's Rho.

    ERIC Educational Resources Information Center

    Caruso, John C.; Cliff, Norman

    1997-01-01

    Several methods of constructing confidence intervals for Spearman's rho (rank correlation coefficient) (C. Spearman, 1904) were tested in a Monte Carlo study using 2,000 samples of 3 different sizes. Results support the continued use of Spearman's rho in behavioral research. (SLD)

  15. Rho-Kinase Activation in Leukocytes Plays a Pivotal Role in Myocardial Ischemia/Reperfusion Injury

    PubMed Central

    Kitano, Katsunori; Usui, Soichiro; Ootsuji, Hiroshi; Takashima, Shin-ichiro; Kobayashi, Daisuke; Murai, Hisayoshi; Furusho, Hiroshi; Nomura, Ayano; Kaneko, Shuichi; Takamura, Masayuki

    2014-01-01

    The Rho/Rho-kinase pathway plays an important role in many cardiovascular diseases such as hypertension, atherosclerosis, heart failure, and myocardial infarction. Although previous studies have shown that Rho-kinase inhibitors reduce ischemia/reperfusion (I/R) injury and cytokine production, the role of Rho-kinase in leukocytes during I/R injury is not well understood. Mice were subjected to 30-min ischemia and reperfusion. Rho-kinase activity was significantly greater in leukocytes subjected to myocardial I/R compared to the sham-operated mice. Administration of fasudil, a Rho-kinase inhibitor, significantly reduced the I/R-induced expression of the proinflammatory cytokines interleukin (IL)-6, C-C motif chemoattractant ligand 2 (CCL2), and tumor necrosis factor (TNF)-α, in leukocytes, compared with saline as the vehicle. Furthermore, fasudil decreased I/R-induced myocardial infarction/area at risk (IA) and I/R-induced leukocyte infiltration in the myocardium. Interestingly, IA in fasudil-administered mice with leukocyte depletion was similar to that in fasudil-administered mice. I/R also resulted in remarkable increases in the mRNA expression levels of the proinflammatory cytokines TNF-α, IL-6, and CCL2 in the heart. Inhibition of Rho-kinase activation in leukocytes has an important role in fasudil-induced cardioprotective effects. Hence, inhibition of Rho-kinase may be an additional therapeutic intervention for the treatment of acute coronary syndrome. PMID:24638037

  16. RhoGEFs in cell motility: Novel links between Rgnef and focal adhesion kinase

    PubMed Central

    Miller, Nichol L. G.; Kleinschmidt, Elizabeth G.; Schlaepfer, David D.

    2014-01-01

    Rho guanine exchange factors (GEFs) are a large, diverse family of proteins defined by their ability to catalyze the exchange of GDP for GTP on small GTPase proteins such as Rho family members. GEFs act as integrators from varied intra- and extracellular sources to promote spatiotemporal activity of Rho GTPases that control signaling pathways regulating cell proliferation and movement. Here we review recent studies elucidating roles of RhoGEF proteins in cell motility. Emphasis is placed on Dbl-family GEFs and connections to development, integrin signaling to Rho GTPases regulating cell adhesion and movement, and how these signals may enhance tumor progression. Moreover, RhoGEFs have additional domains that confer distinctive functions or specificity. We will focus on a unique interaction between Rgnef (also termed Arhgef28 or p190RhoGEF) and focal adhesion kinase (FAK), a non-receptor tyrosine kinase that controls migration properties of normal and tumor cells. This Rgnef-FAK interaction activates canonical GEF-dependent RhoA GTPase activity to govern contractility and also functions as a scaffold in a GEF-independent manner to enhance FAK activation. Recent studies have also brought to light the importance of specific regions within the Rgnef pleckstrin homology (PH) domain for targeting the membrane. As revealed by ongoing Rgnef-FAK investigations, exploring GEF roles in cancer will yield fundamental new information on the molecular mechanisms promoting tumor spread and metastasis. PMID:24467206

  17. Cationic modulation of rho 1-type gamma-aminobutyrate receptors expressed in Xenopus oocytes.

    PubMed Central

    Calvo, D J; Vazquez, A E; Miledi, R

    1994-01-01

    A study was made of the effects of di- and trivalent cations on homomeric rho 1-type gamma-aminobutyrate (GABA rho 1) receptors expressed in Xenopus oocytes after injection of mRNA coding for the GABA rho 1 subunit. GABA elicited large currents with a Kd approximately 1 microM. The properties of these GABA rho 1 receptors were similar to those of native bicuculline-resistant GABA receptors expressed by retinal mRNA. GABA rho 1 currents showed very little desensitization, were blocked by picrotoxin but not by bicuculline, and were not modulated by barbiturates, benzodiazepines, or beta-carbolines. Zn2+ reversibly decreased GABA rho 1 responses (IC50 = 22 microM). Other divalent cations were also tested and their rank order of potency was: Zn2+ approximately Ni2+ approximately Cu2+ >> Cd2+, whereas Ba2+, Co2+, Sr2+, Mn2+, Mg2+, and Ca2+ showed little or no effect. In contrast, La3+ reversibly potentiated the GABA currents mediated by homomeric GABA rho 1 receptors, with an EC50 = 135 microM and a maximal potentiation of about 100% (GABA, 1 microM; La3+, 1 mM). Other lanthanides showed similar effects (Lu3+ > Eu3+ > Tb3+ > Gd3+ > Er3% > Nd3+ > La3+ > Ce3+). Thus, GABA rho 1 receptors contain sites for cationic recognition, and in particular, Zn2+ may play a role during synaptic transmission in the retina. Images Fig. 3 PMID:7809110

  18. alpha-Hexachlorocyclohexane (alpha-HCH)

    Integrated Risk Information System (IRIS)

    alpha - Hexachlorocyclohexane ( alpha - HCH ) ; CASRN 319 - 84 - 6 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Ass

  19. RhoA/ROCK downregulates FPR2-mediated NADPH oxidase activation in mouse bone marrow granulocytes.

    PubMed

    Filina, Julia V; Gabdoulkhakova, Aida G; Safronova, Valentina G

    2014-10-01

    Polymorphonuclear neutrophils (PMNs) express the high and low affinity receptors to formylated peptides (mFPR1 and mFPR2 in mice, accordingly). RhoA/ROCK (Rho activated kinase) pathway is crucial for cell motility and oxidase activity regulated via FPRs. There are contradictory data on RhoA-mediated regulation of NADPH oxidase activity in phagocytes. We have shown divergent Rho GTPases signaling via mFPR1 and mFPR2 to NADPH oxidase in PMNs from inflammatory site. The present study was aimed to find out the role of RhoA/ROCK in the respiratory burst activated via mFPR1 and mFPR2 in the bone marrow PMNs. Different kinetics of RhoA activation were detected with 0.1μM fMLF and 1μM WKYMVM operating via mFPR1 and mFPR2, accordingly. RhoA was translocated in fMLF-activated cells towards the cell center and juxtamembrane space versus uniform allocation in the resting cells. Specific inhibition of RhoA by CT04, Rho inhibitor I, weakly depressed the respiratory burst induced via mFPR1, but significantly increased the one induced via mFPR2. Inhibition of ROCK, the main effector of RhoA, by Y27632 led to the same effect on the respiratory burst. Regulation of mFPR2-induced respiratory response by ROCK was impossible under the cytoskeleton disruption by cytochalasin D, whereas it persisted in the case of mFPR1 activation. Thus we suggest RhoA to be one of the regulatory and signal transduction components in the respiratory burst through FPRs in the mouse bone marrow PMNs. Both mFPR1 and mFPR2 binding with a ligand trigger the activation of RhoA. FPR1 signaling through RhoA/ROCK increases NADPH-oxidase activity. But in FPR2 action RhoA/ROCK together with cytoskeleton-linked systems down-regulates NADPH-oxidase. This mechanism could restrain the reactive oxygen species dependent damage of own tissues during the chemotaxis of PMNs and in the resting cells.

  20. A Gα12-specific Binding Domain in AKAP-Lbc and p114RhoGEF

    PubMed Central

    Brawley, Douglas N.; Berkley, Carrie Y.; Smolski, William C.; Garcia, Ricardo D.; Towne, Autumn L.; Sims, Jonathan R.

    2016-01-01

    AKAP-Lbc is a Rho-activating guanine nucleotide exchange factor (RhoGEF) important in heart development and pro-fibrotic signaling in cardiomyocytes. Heterotrimeric G proteins of the G12/13 subfamily, comprising Gα12 and Gα13, are well characterized as stimulating a specialized group of RhoGEFs through interaction with their RGS-homology (RH) domain. Despite lacking an RH domain, AKAP-Lbc is bound by Gα12 through an unknown mechanism to activate Rho signaling. We identified a Gα12-binding region near the C-terminus of AKAP-Lbc, closely homologous to a region of p114RhoGEF that we also discovered to interact with Gα12. This binding mechanism is distinct from the well-studied interface between RH-RhoGEFs and G12/13 α subunits, as demonstrated by Gα12 mutants selectively impaired in binding either this AKAP-Lbc/p114RhoGEF region or RH-RhoGEFs. AKAP-Lbc and p114RhoGEF showed high specificity for binding Gα12 in comparison to Gα13, and experiments using chimeric G12/13 α subunits mapped determinants of this selectivity to the N-terminal region of Gα12. In cultured cells expressing constitutively GDP-bound Gα12 or Gα13, the Gα12 construct was more potent in exerting a dominant-negative effect on serum-mediated signaling to p114RhoGEF, demonstrating coupling of these signaling proteins in a cellular pathway. In addition, charge-reversal of conserved residues in AKAP-Lbc and p114RhoGEF disrupted Gα12 binding for both proteins, suggesting they harbor a common structural mechanism for interaction with this α subunit. Our results provide the first evidence of p114RhoGEF as a Gα12 signaling effector, and define a novel region conserved between AKAP-Lbc and p114RhoGEF that allows Gα12 signaling input to these non-RH RhoGEFs.

  1. Skeletal muscle differentiation and fusion are regulated by the BAR-containing Rho-GTPase-activating protein (Rho-GAP), GRAF1.

    PubMed

    Doherty, Jason T; Lenhart, Kaitlin C; Cameron, Morgan V; Mack, Christopher P; Conlon, Frank L; Taylor, Joan M

    2011-07-22

    Although RhoA activity is necessary for promoting myogenic mesenchymal stem cell fates, recent studies in cultured cells suggest that down-regulation of RhoA activity in specified myoblasts is required for subsequent differentiation and myotube formation. However, whether this phenomenon occurs in vivo and which Rho modifiers control these later events remain unclear. We found that expression of the Rho-GTPase-activating protein, GRAF1, was transiently up-regulated during myogenesis, and studies in C2C12 cells revealed that GRAF1 is necessary and sufficient for mediating RhoA down-regulation and inducing muscle differentiation. Moreover, forced expression of GRAF1 in pre-differentiated myoblasts drives robust muscle fusion by a process that requires GTPase-activating protein-dependent actin remodeling and BAR-dependent membrane binding or sculpting. Moreover, morpholino-based knockdown studies in Xenopus laevis determined that GRAF1 expression is critical for muscle development. GRAF1-depleted embryos exhibited elevated RhoA activity and defective myofibrillogenesis that resulted in progressive muscle degeneration, defective motility, and embryonic lethality. Our results are the first to identify a GTPase-activating protein that regulates muscle maturation and to highlight the functional importance of BAR domains in myotube formation.

  2. Molecular characterization of a novel RhoGAP, RRC-1 of the nematode Caenorhabditis elegans

    SciTech Connect

    Delawary, Mina; Nakazawa, Takanobu; Tezuka, Tohru; Sawa, Mariko; Iino, Yuichi; Takenawa, Tadaomi; Yamamoto, Tadashi . E-mail: tyamamot@ims.u-tokyo.ac.jp

    2007-06-01

    The GTPase-activating proteins for Rho family GTPases (RhoGAP) transduce diverse intracellular signals by negatively regulating Rho family GTPase-mediated pathways. In this study, we have cloned and characterized a novel RhoGAP for Rac1 and Cdc42, termed RRC-1, from Caenorhabditis elegans. RRC-1 was highly homologous to mammalian p250GAP and promoted GTP hydrolysis of Rac1 and Cdc42 in cells. The rrc-1 mRNA was expressed in all life stages. Using an RRC-1::GFP fusion protein, we found that RRC-1 was localized to the coelomocytes, excretory cell, GLR cells, and uterine-seam cell in adult worms. These data contribute toward understanding the roles of Rho family GTPases in C. elegans.

  3. Depleting Mycobacterium tuberculosis of the transcription termination factor Rho causes pervasive transcription and rapid death.

    PubMed

    Botella, Laure; Vaubourgeix, Julien; Livny, Jonathan; Schnappinger, Dirk

    2017-03-28

    Rifampicin, which inhibits bacterial RNA polymerase, provides one of the most effective treatments for tuberculosis. Inhibition of the transcription termination factor Rho is used to treat some bacterial infections, but its importance varies across bacteria. Here we show that Rho of Mycobacterium tuberculosis functions to both define the 3' ends of mRNAs and silence substantial fragments of the genome. Brief inactivation of Rho affects over 500 transcripts enriched for genes of foreign DNA elements and bacterial virulence factors. Prolonged inactivation of Rho causes extensive pervasive transcription, a genome-wide increase in antisense transcripts, and a rapid loss of viability of replicating and non-replicating M. tuberculosis in vitro and during acute and chronic infection in mice. Collectively, these data suggest that inhibition of Rho may provide an alternative strategy to treat tuberculosis with an efficacy similar to inhibition of RNA polymerase.

  4. Depleting Mycobacterium tuberculosis of the transcription termination factor Rho causes pervasive transcription and rapid death

    PubMed Central

    Botella, Laure; Vaubourgeix, Julien; Livny, Jonathan; Schnappinger, Dirk

    2017-01-01

    Rifampicin, which inhibits bacterial RNA polymerase, provides one of the most effective treatments for tuberculosis. Inhibition of the transcription termination factor Rho is used to treat some bacterial infections, but its importance varies across bacteria. Here we show that Rho of Mycobacterium tuberculosis functions to both define the 3′ ends of mRNAs and silence substantial fragments of the genome. Brief inactivation of Rho affects over 500 transcripts enriched for genes of foreign DNA elements and bacterial virulence factors. Prolonged inactivation of Rho causes extensive pervasive transcription, a genome-wide increase in antisense transcripts, and a rapid loss of viability of replicating and non-replicating M. tuberculosis in vitro and during acute and chronic infection in mice. Collectively, these data suggest that inhibition of Rho may provide an alternative strategy to treat tuberculosis with an efficacy similar to inhibition of RNA polymerase. PMID:28348398

  5. Molecular pathways: targeting the kinase effectors of RHO-family GTPases.

    PubMed

    Prudnikova, Tatiana Y; Rawat, Sonali J; Chernoff, Jonathan

    2015-01-01

    RHO GTPases, members of the RAS superfamily of small GTPases, are adhesion and growth factor-activated molecular switches that play important roles in tumor development and progression. When activated, RHO-family GTPases such as RAC1, CDC42, and RHOA, transmit signals by recruiting a variety of effector proteins, including the protein kinases PAK, ACK, MLK, MRCK, and ROCK. Genetically induced loss of RHO function impedes transformation by a number of oncogenic stimuli, leading to an interest in developing small-molecule inhibitors that either target RHO GTPases directly, or that target their downstream protein kinase effectors. Although inhibitors of RHO GTPases and their downstream signaling kinases have not yet been widely adopted for clinical use, their potential value as cancer therapeutics continues to facilitate pharmaceutical research and development and is a promising therapeutic strategy.

  6. Alpha Hydroxy Acids

    MedlinePlus

    ... Cosmetics Home Cosmetics Products & Ingredients Ingredients Alpha Hydroxy Acids Share Tweet Linkedin Pin it More sharing options ... for Industry: Labeling for Cosmetics Containing Alpha Hydroxy Acids The following information is intended to answer questions ...

  7. Rho GDP dissociation inhibitor beta promotes cell proliferation and invasion by modulating the AKT pathway in hepatocellular carcinoma.

    PubMed

    Fang, Yang; Yi, Jiang; Lizhi, Lv; Qiucheng, Cai

    2014-11-01

    Rho GDP dissociation inhibitor (GDI) beta, (RhoGDI2), has been identified as a proto-oncogene that is upregulated in human cancers, but the role of RhoGDI2 in hepatocellular carcinoma (HCC) remains unclear. In the present study, we investigated the RhoGDI2 expression level in HCC tissues and the function of RhoGDI2 in HCC cell growth and metastasis. We examined the RhoGDI2 mRNA expression level in 64 sets of HCC tissue and their adjacent nontumor tissue counterparts using quantitative real-time polymerase chain reaction. In vitro proliferation and invasion assays were conducted to determine the effect of RhoGDI2 on the ability of HCC cells to proliferate and invade, respectively. Western blot analysis was conducted to examine expression levels of RhoGDI2p-AKT, MMP-2, and MMP-9 in HCC cells. RhoGDI2 mRNA was significantly overexpressed in the HCC specimens compared with the nonneoplastic liver specimens, and the RhoGDI2 mRNA and protein levels were higher in the HCC cell lines, especially the highly metastatic cell lines 97L and 97H. To further investigate the role that RhoGDI2 plays in HCC, we overexpressed RhoGDI2 using a lentivirus-mediated overexpression technique in two HCC cell lines (Huh7 and 7721) that endogenously express a low level of RhoGDI2. Stable cells overexpressing RhoGDI2 demonstrated a significant increase in cell proliferation and invasion. Furthermore, our additional findings indicated that RhoGDI2-mediated cellular invasion requires the PI3K/Akt signaling-dependent expression of matrix metalloproteinases (MMPs). Our findings suggest that RhoGDI2 plays an important role in HCC growth and invasion and should be considered a novel HCC therapeutic target candidate.

  8. The Human Minor Histocompatibility Antigen1 Is a RhoGAP

    PubMed Central

    de Kreuk, Bart-Jan; Schaefer, Antje; Anthony, Eloise C.; Tol, Simon; Fernandez-Borja, Mar; Geerts, Dirk; Pool, Jos; Hambach, Lothar; Goulmy, Els; Hordijk, Peter L.

    2013-01-01

    The human minor Histocompatibility Antigen HMHA-1 is a major target of immune responses after allogeneic stem cell transplantation applied for the treatment of leukemia and solid tumors. The restriction of its expression to hematopoietic cells and many solid tumors raised questions regarding its cellular functions. Sequence analysis of the HMHA-1 encoding HMHA1 protein revealed the presence of a possible C-terminal RhoGTPase Activating Protein (GAP) domain and an N-terminal BAR domain. Rho-family GTPases, including Rac1, Cdc42, and RhoA are key regulators of the actin cytoskeleton and control cell spreading and migration. RhoGTPase activity is under tight control as aberrant signaling can lead to pathology, including inflammation and cancer. Whereas Guanine nucleotide Exchange Factors (GEFs) mediate the exchange of GDP for GTP resulting in RhoGTPase activation, GAPs catalyze the low intrinsic GTPase activity of active RhoGTPases, resulting in inactivation. Here we identify the HMHA1 protein as a novel RhoGAP. We show that HMHA1 constructs, lacking the N-terminal region, negatively regulate the actin cytoskeleton as well as cell spreading. Furthermore, we show that HMHA1 regulates RhoGTPase activity in vitro and in vivo. Finally, we demonstrate that the HMHA1 N-terminal BAR domain is auto-inhibitory as HMHA1 mutants lacking this region, but not full-length HMHA1, showed GAP activity towards RhoGTPases. In conclusion, this study shows that HMHA1 acts as a RhoGAP to regulate GTPase activity, cytoskeletal remodeling and cell spreading, which are crucial functions in normal hematopoietic and cancer cells. PMID:24086303

  9. The Andes Virus Nucleocapsid Protein Directs Basal Endothelial Cell Permeability by Activating RhoA

    PubMed Central

    Gorbunova, Elena E.; Simons, Matthew J.; Gavrilovskaya, Irina N.

    2016-01-01

    ABSTRACT Andes virus (ANDV) predominantly infects microvascular endothelial cells (MECs) and nonlytically causes an acute pulmonary edema termed hantavirus pulmonary syndrome (HPS). In HPS patients, virtually every pulmonary MEC is infected, MECs are enlarged, and infection results in vascular leakage and highly lethal pulmonary edema. We observed that MECs infected with the ANDV hantavirus or expressing the ANDV nucleocapsid (N) protein showed increased size and permeability by activating the Rheb and RhoA GTPases. Expression of ANDV N in MECs increased cell size by preventing tuberous sclerosis complex (TSC) repression of Rheb-mTOR-pS6K. N selectively bound the TSC2 N terminus (1 to 1403) within a complex containing TSC2/TSC1/TBC1D7, and endogenous TSC2 reciprocally coprecipitated N protein from ANDV-infected MECs. TSCs normally restrict RhoA-induced MEC permeability, and we found that ANDV infection or N protein expression constitutively activated RhoA. This suggests that the ANDV N protein alone is sufficient to activate signaling pathways that control MEC size and permeability. Further, RhoA small interfering RNA, dominant-negative RhoA(N19), and the RhoA/Rho kinase inhibitors fasudil and Y27632 dramatically reduced the permeability of ANDV-infected MECs by 80 to 90%. Fasudil also reduced the bradykinin-directed permeability of ANDV and Hantaan virus-infected MECs to control levels. These findings demonstrate that ANDV activation of RhoA causes MEC permeability and reveal a potential edemagenic mechanism for ANDV to constitutively inhibit the basal barrier integrity of infected MECs. The central importance of RhoA activation in MEC permeability further suggests therapeutically targeting RhoA, TSCs, and Rac1 as potential means of resolving capillary leakage during hantavirus infections. PMID:27795403

  10. The Alpha Centauri System.

    ERIC Educational Resources Information Center

    Soderblom, David R.

    1987-01-01

    Describes the Alpha Centauri star system, which is the closest star system to the sun. Discusses the difficulties associated with measurements involving Alpha Centauri, along with some of the recent advances in stellar seismology. Raises questions about the possibilities of planets around Alpha Centauri. (TW)

  11. RhoA GTPase activation by TLR2 and TLR3 ligands: connecting via Src to NF-kappa B.

    PubMed

    Manukyan, Maria; Nalbant, Perihan; Luxen, Sylvia; Hahn, Klaus M; Knaus, Ulla G

    2009-03-15

    Rho GTPases are essential regulators of signaling networks emanating from many receptors involved in innate or adaptive immunity. The Rho family member RhoA controls cytoskeletal processes as well as the activity of transcription factors such as NF-kappaB, C/EBP, and serum response factor. The multifaceted host cell activation triggered by TLRs in response to soluble and particulate microbial structures includes rapid stimulation of RhoA activity. RhoA acts downstream of TLR2 in HEK-TLR2 and monocytic THP-1 cells, but the signaling pathway connecting TLR2 and RhoA is still unknown. It is also not clear if RhoA activation is dependent on a certain TLR adapter. Using lung epithelial cells, we demonstrate TLR2- and TLR3-triggered recruitment and activation of RhoA at receptor-proximal cellular compartments. RhoA activity was dependent on TLR-mediated stimulation of Src family kinases. Both Src family kinases and RhoA were required for NF-kappaB activation, whereas RhoA was dispensable for type I IFN generation. These results suggest that RhoA plays a role downstream of MyD88-dependent and -independent TLR signaling and acts as a molecular switch downstream of TLR-Src-initiated pathways.

  12. RhoE Promotes Metastasis in Gastric Cancer through a Mechanism Dependent on Enhanced Expression of CXCR4

    PubMed Central

    Wang, Hefei; Shang, Yulong; Bai, Ming; Liang, Jie; Wang, Xin; Fan, Daiming

    2013-01-01

    RhoE, a novel member of the Rho protein family, is a key regulator of the cytoskeleton and cell migration. Our group has previously shown that RhoE as a direct target for HIF-1α and mediates hypoxia-induced epithelial to mesenchymal transition in gastric cancer cells. Therefore, we assumed that RhoE might play an important role in gastric cancer metastasis. In the present study, we have explored the role of RhoE expression in gastric cancer, cell invasion and metastasis, and the influence of RhoE on regulating the potential expression of down-stream genes. RhoE expression was elevated in gastric cancer tissues as compared with normal gastric tissues. We also found a close correlation between the histological grade and the diagnosis of the patient. Up-regulation of RhoE significantly enhanced the migratory and invasive abilities of gastric cancer cells both in vitro and in vivo. Moreover, down-regulation of RhoE diminished the metastatic potential of cancer cells. PCR array and subsequent transwell assay showed that the regulation of gastric cancer metastasis by RhoE was partially mediated by CXCR4. This observation suggested that CXCR4 might be a downstream effector for RhoE. In summary, our study identified RhoE as a novel prognostic biomarker and metastatic-promoting gene of gastric cancer. PMID:24312338

  13. Observations of heavy-element recombination lines in the Rho Ophiuchi dark cloud at 13 centimeters wavelength

    NASA Technical Reports Server (NTRS)

    Knapp, G. R.; Kuiper, T. B. H.; Brown, R. L.

    1976-01-01

    High-sensitivity observations at the carbon 140-alpha and 141-alpha recombination-line frequencies in the direction of the Rho Oph dust cloud show the presence of recombination lines of carbon and sulfur, but not of other heavy elements. These results require that the elements Si, Fe, and Mg are depleted by factors greater than 15, 10, and 2, respectively, most likely into interstellar grains, whereas sulfur is undepleted. They also require that only approximately one-sixth of the carbon is present in the gas phase in the cloud. From the observed size of the C II region together with the observed carbon depletion, it is inferred that the density in the carbon-line-emitting region is 25,000 per cu cm and that the size of the S II region is significantly larger than that of the C II region. The agreement between the widths of the recombination lines (which arise in a small region of the cloud) and those of the more widely distributed molecular species suggests that the lines in this cloud are broadened by microturbulence on a scale of much less than 0.1 pc.

  14. Depletion of calcium in the Rho Ophiuchi cloud

    SciTech Connect

    Snow, T.P.; Timothy, J.G.; Seab, C.G.

    1983-02-15

    Data on the interstellar Ca II H and K lines toward HD 147889, a star deeply embedded in the rho Ophiuchi dark cloud, were obtained at the Mauna Kea Observatory of the University of Hawaii, using a multi-anode microchannel array (MAMA) detector. An upper limit on the Ca II column density of 1.7 x 10/sup 11/ cm/sup -2/ was derived, which, together with ionization considerations (published elsewhere) and an estimated lower limit on the total hydrogen column density, implies that the calcium-to-hydrogen ratio in this line of sight is 7.95 x 10/sup -6/ or less, relative to the solar value. The logarithmic depletion is < or =-5.10, the largest yet measured, and implies that gas-grain interaction in this region has proceeded to a much greater degree than is typical of diffuse clouds.

  15. Chemotherapy resistance in diffuse type gastric adenocarcinoma is mediated by RhoA activation in cancer stem-like cells

    PubMed Central

    Yoon, Changhwan; Cho, Soo-Jeong; Aksoy, Bülent Arman; Park, Do Joong; Schultz, Nikolaus; Ryeom, Sandra W.; Yoon, Sam S.

    2016-01-01

    Purpose The Lauren diffuse type of gastric adenocarcinoma (DGA), as opposed to the intestinal type (IGA), often harbor mutations in RHOA but little is known about the role of RhoA in DGA. Experimental Design We examined RhoA activity and RhoA pathway inhibition in DGA cell lines and in two mouse xenograft models. RhoA activity was also assessed in patient tumor samples. Results RhoA activity was higher in DGA compared to IGA cell lines, and was further increased when grown as spheroids to enrich for cancer stem-like cells (CSC) or when sorted using the gastric CSC marker CD44. RhoA shRNA or the RhoA inhibitor Rhosin decreased expression of the stem cell transcription factor, Sox2, and decreased spheroid formation by 78–81%. DGA spheroid cells had 3–5 fold greater migration and invasion than monolayer cells, and this activity was Rho-dependent. Diffuse GA spheroid cells were resistant in a cytotoxicity assay to 5-fluorouracil and cisplatin chemotherapy, and this resistance could be reversed with RhoA pathway inhibition. In two xenograft models, cisplatin inhibited tumor growth by 40–50%, RhoA inhibition by 32–60%, and the combination by 77–83%. In 288 patient tumors, increased RhoA activity correlated with worse OS in DGA patients (p=0.017) but not in IGA patients (p=0.612). Conclusions RhoA signaling promotes CSC phenotypes in DGA cells. Increased RhoA activity is correlated with worse OS in DGA patients and RhoA inhibition can reverse chemotherapy resistance in DGA CSC and in tumor xenografts. Thus the RhoA pathway is a promising new target in DGA patients. PMID:26482039

  16. RhoA-mediated inhibition of vascular endothelial cell mobility: positive feedback through reduced cytosolic p21 and p27.

    PubMed

    Hsu, Yung-Ho; Chang, Chih-Cheng; Yang, Nian-Jie; Lee, Yi-Hsuan; Juan, Shu-Hui

    2014-10-01

    We previously identified that activation of the aryl hydrocarbon receptor (AhR) by 3-methylcholanthrene (3MC) exerts antiproliferative and antimigratory effects on human umbilical vein endothelial cells (HUVECs) through the upregulation of p21/p27 transcription and RhoA activation. In this study, we investigated the mechanisms of 3MC-mediated downregulation of cytosolic p21/ p27, and the effects of 3MC on RhoA activation and cell migration, in mouse cerebral vascular endothelial cells (MCVECs). Our results indicated that 3MC reduced the phosphorylation of p21/p27 through AhR/RhoA/PTEN-mediated PI3K/Akt inactivation, which reduced cytosolic p21/p27 retention, causing RhoA activation through positive feedback. Downregulation of p21/p27 by siRNA, and cytosolic p21/p27 by the nuclear export blocker leptomycin B, further reduced cell migration in the 3MC-treated cells. Reduced cytosolic p21/p27 expression led to reduced interaction between RhoA and the RhoA inhibitor p190RhoGAP, causing RhoA activation. Treatment with YS-49 activated PI3K/Akt, a downstream target of RhoA, to reduce RhoA/PTEN activation in the 3MC-treated cells, whereas treatment with wortmannin, a PI3K inhibitor, activated RhoA/PTEN. Gain- and loss-of-function analyses revealed that constitutively active (CA) Akt1, but not CA Akt2, inactivated RhoA and stimulated migratory activity. Considering the essential role of RhoA activation in cell migration, we evaluated the potential use of simvastatin, a RhoA inhibitor, as a therapeutic intervention in vivo using matrigel plug formation assays. Our results provide a molecular basis for the therapeutic application of simvastatin to reduce RhoA/PTEN activation, restore cytosolic levels of phosphorylated p21/p27, and induce angiogenic processes.

  17. Diverse activation states of RhoA in human lung cancer cells: contribution of G protein coupled receptors.

    PubMed

    Touge, Hirokazu; Chikumi, Hiroki; Igishi, Tadashi; Kurai, Jun; Makino, Haruhiko; Tamura, Yoshisato; Takata, Miyako; Yoneda, Kazuhiko; Nakamoto, Masaki; Suyama, Hisashi; Gutkind, J Silvio; Shimizu, Eiji

    2007-03-01

    Rho GTPases play an essential role in the control of various cellular functions. Accumulating evidence suggests that RhoA overexpression contributes to human cancer development. However, the activation states of RhoA are poorly defined in cancer cells. In this study, we examined both the expression levels and the activation states of RhoA in various lung cancer cells by quantitative real-time reverse transcriptase-polymerase chain reaction and in vivo Rho guanine nucleotide exchange assay, respectively. Moreover, we dissected the signaling pathway from the cell surface receptors to RhoA using a broad-spectrum G protein coupled receptor (GPCR) antagonist, [D-Arg1,D-Trp5,7,9,Leu11]Substance P (SP), and a recently reported Galphaq/11-selective inhibitor, YM-254890. We found that RhoA was expressed highly in large cell carcinoma cells but only weakly in adenocarcinoma cells. The activation states of RhoA are considerably different from its expression profiles. We found that four of six small cell lung carcinoma (SCLC) cell lines exhibited a moderate to high activation rate of RhoA. The addition of [D-Arg1,D-Trp5,7,9,Leu11]SP reduced RhoA activity by almost 60% in H69 SCLC cells. The addition of YM-254890 had no effect on RhoA activity in H69 cells. Our results suggest that RhoA is activated in various lung cancer cells independent of its expression levels, and the high activation state of RhoA in SCLC cells mainly depends on a neuroendocrine peptide autocrine system which signals through Galpha12 coupled GPCR to RhoA. This study provides new insights into RhoA signaling in lung cancer cells and may help in developing novel therapeutic strategies against lung cancer.

  18. THE INITIAL MASS FUNCTION AND DISK FREQUENCY OF THE {rho} OPHIUCHI CLOUD: AN EXTINCTION-LIMITED SAMPLE

    SciTech Connect

    Erickson, Kristen L.; Wilking, Bruce A.; Robinson, John G.; Stephenson, Lauren N.; Meyer, Michael R. E-mail: bwilking@umsl.edu E-mail: lnafff@mail.umsl.edu

    2011-10-15

    We have completed an optical spectroscopic survey of an unbiased, extinction-limited sample of candidate young stars covering 1.3 deg{sup 2} of the {rho} Ophiuchi star-forming region. While infrared, X-ray, and optical surveys of the cloud have identified many young stellar objects (YSOs), these surveys are biased toward particular stages of stellar evolution and are not optimal for studies of the disk frequency and initial mass function. We have obtained over 300 optical spectra to help identify 135 association members based on the presence of H{alpha} in emission, lithium absorption, X-ray emission, a mid-infrared excess, a common proper motion, reflection nebulosity, and/or extinction considerations. Spectral types along with R- and I-band photometry were used to derive effective temperatures and bolometric luminosities for association members to compare with theoretical tracks and isochrones for pre-main-sequence stars. An average age of 3.1 Myr is derived for this population which is intermediate between that of objects embedded in the cloud core of {rho} Ophiuchi and low-mass stars in the Upper Scorpius subgroup. Consistent with this age we find a circumstellar disk frequency of 27% {+-} 5%. We also constructed an initial mass function for an extinction-limited sample of 123 YSOs (A{sub v} {<=} 8 mag), which is consistent with the field star initial mass function for YSOs with masses >0.2 M{sub sun}. There may be a deficit of brown dwarfs but this result relies on completeness corrections and requires confirmation.

  19. Inhibition of the Rho GTPase, Rac1, decreases estrogen receptor levels and is a novel therapeutic strategy in breast cancer.

    PubMed

    Rosenblatt, Adena E; Garcia, Maria Ines; Lyons, Leah; Xie, Yingqiu; Maiorino, Carol; Désiré, Laurent; Slingerland, Joyce; Burnstein, Kerry L

    2011-04-01

    Rac1, a Rho GTPase, modulates diverse cellular processes and is hyperactive in some cancers. Estrogen receptor-alpha (ERα) in concert with intracellular signaling pathways regulates genes associated with cell proliferation, tumor development, and breast cancer cell survival. Therefore, we examined the possibility of Rac1 and ERα crosstalk in breast cancer cells. We found that Rac1 enhanced ERα transcriptional activity in breast cancer cells. Vav3, a Rho guanine nucleotide exchange factor that activates Rac1, was an upstream mediator, and P21/Cdc42/Rac1 activating kinase-1 (Pak-1) was a downstream effector of Rac1 enhancement of ERα activity. These results suggest that Rac1 may prove to be a therapeutic target. To test this hypothesis, we used a small molecule Rac inhibitor, EHT 1864, and found that EHT 1864 inhibited ERα transcriptional activity. Furthermore, EHT 1864 inhibited estrogen-induced cell proliferation in breast cancer cells and decreased tamoxifen-resistant breast cancer cell growth. EHT 1864 decreased activity of the promoter of the ERα gene resulting in down-regulation of ERα mRNA and protein levels. Therefore, ERα down-regulation by EHT 1864 is the likely mechanism of EHT 1864-mediated inhibition of ERα activity and estrogen-stimulated breast cancer cell proliferation. Since ERα plays a critical role in the pathogenesis of breast cancer and the Rac inhibitor EHT 1864 down-regulates ERα expression and breast cancer cell proliferation, further investigation of the therapeutic potential of Rac1 targeting in the treatment of breast cancer is warranted.

  20. Involvement of Rho-type GTPase in control of cell size in Saccharomyces cerevisiae.

    PubMed

    Kikuchi, Yo; Mizuuchi, Eri; Nogami, Satoru; Morishita, Shinichi; Ohya, Yoshikazu

    2007-06-01

    Maintaining specific cell size, which is important for many organisms, is achieved by coordinating cell growth and cell division. In the budding yeast Saccharomyces cerevisiae, the existence of two cell-size checkpoints is proposed: at the first checkpoint, cell size is monitored before budding at the G1/S transition, and at the second checkpoint, actin depolymerization occurring in the small bud is monitored before the G2/M transition. Morphological analyses have revealed that the small GTPase Rho1p participates in cell-size control at both the G1/S and the G2/M boundaries. One group of rho1 mutants (rho1A) underwent premature entry into mitosis, leading to the birth of abnormally small cells. In another group of rho1 mutants (rho1B), the mother cells failed to reach an appropriate size before budding, and expression of the G1 cyclin Cln2p began at an earlier phase of the cell cycle. Analyses of mutants defective in Rho1p effector proteins indicate that Skn7p, Fks1p and Mpk1p are involved in cell-size control. Thus, Rho1p and its downstream regulatory pathways are involved in controlling cell size in S. cerevisiae.

  1. Nitric oxide promotes epidermal stem cell migration via cGMP-Rho GTPase signalling

    PubMed Central

    Zhan, Rixing; He, Weifeng; Wang, Fan; Yao, Zhihui; Tan, Jianglin; Xu, Rui; Zhou, Junyi; Wang, Yuzhen; Li, Haisheng; Wu, Jun; LUO, Gaoxing

    2016-01-01

    The migration and reepithelization of epidermal stem cells (ESCs) are the most critical processes in wound healing. The gaseous messenger nitric oxide (NO) has multiple biological effects, but its actions on ESCs are poorly understood. In this study, an NO donor, S-nitroso-N-acetylpenicillamine (SNAP), was found to facilitate the in vitro migration of human ESCs (huESCs) in both live-imaging and scratch models. In addition, pull-down assays demonstrated that SNAP could activate the small GTPases RhoA and Rac1 of the Rho family, but not Cdc42. Moreover, the effects of SNAP on the migration and F-actin polymerization of ESCs could be blocked by inhibitors of cGMP, PKG, RhoA or Rac1, and by a specific siRNA of RhoA or Rac1, but not by a Cdc42 inhibitor or siRNA. Furthermore, the roles of NO in ESC migration via cGMP-Rho GTPase signalling in vivo were confirmed by tracing 5-bromo-2-deoxyuridine (BrdU)-labelled cells in a superficial, partial-thickness scald mouse model. Thus, the present study demonstrated that the NO donor SNAP could promote huESC migration in vitro. Furthermore, NO was found to induce ESC migration via cGMP-Rho GTPase RhoA and Rac1 signalling, but not Cdc42 signalling, both in vivo and in vitro. PMID:27469024

  2. Ameloblasts require active RhoA to generate normal dental enamel.

    PubMed

    Xue, Hui; Li, Yong; Everett, Eric T; Ryan, Kathleen; Peng, Li; Porecha, Rakhee; Yan, Yan; Lucchese, Anna M; Kuehl, Melissa A; Pugach, Megan K; Bouchard, Jessica; Gibson, Carolyn W

    2013-08-01

    RhoA plays a fundamental role in regulation of the actin cytoskeleton, intercellular attachment, and cell proliferation. During amelogenesis, ameloblasts (which produce the enamel proteins) undergo dramatic cytoskeletal changes and the RhoA protein level is up-regulated. Transgenic mice were generated that express a dominant-negative RhoA transgene in ameloblasts using amelogenin gene-regulatory sequences. Transgenic and wild-type (WT) molar tooth germs were incubated with sodium fluoride (NaF) or sodium chloride (NaCl) in organ culture. Filamentous actin (F-actin) stained with phalloidin was elevated significantly in WT ameloblasts treated with NaF compared with WT ameloblasts treated with NaCl or with transgenic ameloblasts treated with NaF, thereby confirming a block in the RhoA/Rho-associated protein kinase (ROCK) pathway in the transgenic mice. Little difference in quantitative fluorescence (an estimation of fluorosis) was observed between WT and transgenic incisors from mice provided with drinking water containing NaF. We subsequently found reduced transgene expression in incisors compared with molars. Transgenic molar teeth had reduced amelogenin, E-cadherin, and Ki67 compared with WT molar teeth. Hypoplastic enamel in transgenic mice correlates with reduced expression of the enamel protein, amelogenin, and E-cadherin and cell proliferation are regulated by RhoA in other tissues. Together these findings reveal deficits in molar ameloblast function when RhoA activity is inhibited.

  3. An adventitious interaction of filamin A with RhoGDI2(Tyr153Glu)

    PubMed Central

    Song, Mia; He, Qianjing; Berk, Benjamin-Andreas; Hartwig, John H.; Stossel, Thomas P.; Nakamura, Fumihiko

    2015-01-01

    Filamin A (FLNA) is an actin filament crosslinking protein with multiple intracellular binding partners. Mechanical force exposes cryptic FLNA binding sites for some of these ligands. To identify new force-dependent binding interactions, we used a fusion construct composed of two FLNA domains, one of which was previously identified as containing a force-dependent binding site as a bait in a yeast two-hybrid system and identified the Rho dissociation inhibitor 2 (RhoGDI2) as a potential interacting partner. A RhoGDI2 truncate with 81 N-terminal amino acid residues and a phosphomimetic mutant, RhoGDI(Tyr153Glu) interacted with the FLNA construct. However, neither wild-type or full-length RhoGDI2 phosphorylated at Y153 interacted with FLNA. Our interpretation of these contradictions is that truncation and/or mutation of RhoGDI2 perturbs its conformation to expose a site that adventitiously binds FLNA and is not a bona-fide interaction. Therefore, previous studies reporting that a RhoGDI(Y153E) mutant suppresses the metastasis of human bladder cancer cells must be reinvestigated in light of artificial interaction of this point mutant with FLNA. PMID:26707877

  4. Guanine nucleotide exchange factors for RhoGTPases: good therapeutic targets for cancer therapy?

    PubMed

    Lazer, Galit; Katzav, Shulamit

    2011-06-01

    Rho guanosine triphosphatases (GTPases) are a family of small proteins which function as molecular switches in a variety of signaling pathways following stimulation of cell surface receptors. RhoGTPases regulate numerous cellular processes including cytoskeleton organization, gene transcription, cell proliferation, migration, growth and cell survival. Because of their central role in regulating processes that are dysregulated in cancer, it seems reasonable that defects in the RhoGTPase pathway may be involved in the development of cancer. RhoGTPase activity is regulated by a number of protein families: guanine nucleotide exchange factors (GEFs), GTPase activating proteins (GAPs) and guanine nucleotide-dissociation inhibitors (GDIs). This review discusses the participation of RhoGTPases and their regulators, especially GEFs in human cancers. In particular, we focus on the involvement of the RhoGTPase GEF, Vav1, a hematopoietic specific signal transducer which is involved in human neuroblastoma, pancreatic ductal carcinoma and lung cancer. Finally, we summarize recent advances in the design and application of a number of molecules that specifically target individual RhoGTPases or their regulators or effectors, and discuss their potential for cancer therapy.

  5. A Rho-signaling pathway mediates cortical granule translocation in the sea urchin oocyte.

    PubMed

    Covián-Nares, Fernando; Martínez-Cadena, Guadalupe; López-Godínez, Juana; Voronina, Ekaterina; Wessel, Gary M; García-Soto, Jesús

    2004-03-01

    Cortical granules are secretory vesicles of the egg that play a fundamental role in preventing polyspermy at fertilization. In the sea urchin egg, they localize directly beneath the plasma membrane forming a compact monolayer and, upon fertilization, undergo a Ca(2+)-dependent exocytosis. Cortical granules form during early oogenesis and, during maturation, translocate from the cytosol to the oocyte cortex in a microfilament-mediated process. We tested the hypothesis that these cortical granule dynamics were regulated by Rho, a GTPase of the Ras superfamily. We observed that Rho is synthesized early in oogenesis, mainly in a soluble form. At the end of maturation, however, Rho associates with cortical granules. Inhibition of Rho with the C3 transferase from C. botulinum blocks cortical granule translocation and microfilaments undergo a significant disorganization. A similar effect is observed by GGTI-286, a geranylgeranyl transferase inhibitor, suggesting that the association of Rho with the cortical granules is indispensable for its function. In contrast, the anchorage of the cortical granules in the cortex, as well as their fusion at fertilization, are Rho-independent processes. We conclude that Rho association with the cortical granules is a critical regulatory step in their translocation to the egg cortex.

  6. Effect of oxidative stress on Rho kinase II and smooth muscle contraction in rat stomach.

    PubMed

    Al-Shboul, Othman; Mustafa, Ayman

    2015-06-01

    Recent studies have shown that both Rho kinase signaling and oxidative stress are involved in the pathogenesis of a number of human diseases, such as diabetes mellitus, hypertension, and atherosclerosis. However, very little is known about the effect of oxidative stress on the gastrointestinal (GI) smooth muscle Rho kinase pathway. The aim of the current study was to investigate the effect of oxidative stress on Rho kinase II and muscle contraction in rat stomach. The peroxynitrite donor 3-morpholinosydnonimine (SIN-1), hydrogen peroxide (H2O2), and peroxynitrite were used to induce oxidative stress. Rho kinase II expression and ACh-induced activity were measured in control and oxidant-treated cells via specifically designed enzyme-linked immunosorbent assay (ELISA) and activity assay kits, respectively. Single smooth muscle cell contraction was measured via scanning micrometry in the presence or absence of the Rho kinase blocker, Y-27632 dihydrochloride. All oxidant agents significantly increased ACh-induced Rho kinase II activity without affecting its expression level. Most important, oxidative stress induced by all three agents augmented ACh-stimulated muscle cell contraction, which was significantly inhibited by Y-27632. In conclusion, oxidative stress activates Rho kinase II and enhances contraction in rat gastric muscle, suggesting an important role in GI motility disorders associated with oxidative stress.

  7. Centralspindlin regulates ECT2 and RhoA accumulation at the equatorial cortex during cytokinesis.

    PubMed

    Nishimura, Yukako; Yonemura, Shigenobu

    2006-01-01

    During determination of the cell division plane, an actomyosin contractile ring is induced at the equatorial cell cortex by signals from the mitotic apparatus and contracts to cause cleavage furrow progression. Although the small GTPase RhoA is known to regulate the progression, probably by controlling actin filament assembly and enhancing actomyosin interaction, any involvement of RhoA in division plane determination is unknown. In this study, using a trichloroacetic acid (TCA) fixation protocol we recently developed, we show that RhoA accumulates at the equatorial cortex before furrow initiation and continues to concentrate at the cleavage furrow during cytokinesis. We also demonstrate that both Rho activity and microtubule organization are required for RhoA localization and proper furrowing. Selective disruption of microtubule organization revealed that both astral and central spindle microtubules can recruit RhoA at the equatorial cortex. We find that centralspindlin and ECT2 are required for RhoA localization and furrowing. Centralspindlin is localized both to central spindle microtubules and at the tips of astral microtubules near the equatorial cortex and recruits ECT2. Positional information for division plane determination from microtubules is transmitted to the cell cortex to organize actin cytoskeleton through a mechanism involving these proteins.

  8. Proplatelet generation in the mouse requires PKCε-dependent RhoA inhibition

    PubMed Central

    Gobbi, Giuliana; Mirandola, Prisco; Carubbi, Cecilia; Masselli, Elena; Sykes, Stephen M.; Ferraro, Francesca; Nouvenne, Antonio; Thon, Jonathan N.; Italiano, Joseph E.

    2013-01-01

    During thrombopoiesis, megakaroycytes undergo extensive cytoskeletal remodeling to form proplatelet extensions that eventually produce mature platelets. Proplatelet formation is a tightly orchestrated process that depends on dynamic regulation of both tubulin reorganization and Rho-associated, coiled-coil containing protein kinase/RhoA activity. A disruption in tubulin dynamics or RhoA activity impairs proplatelet formation and alters platelet morphology. We previously observed that protein kinase Cepsilon (PKCε), a member of the protein kinase C family of serine/threonine-kinases, expression varies during human megakaryocyte differentiation and modulates megakaryocyte maturation and platelet release. Here we used an in vitro model of murine platelet production to investigate a potential role for PKCε in proplatelet formation. By immunofluorescence we observed that PKCε colocalizes with α/β-tubulin in specific areas of the marginal tubular-coil in proplatelets. Moreover, we found that PKCε expression escalates during megakarocyte differentiation and remains elevated in proplatelets, whereas the active form of RhoA is substantially downregulated in proplatelets. PKCε inhibition resulted in lower proplatelet numbers and larger diameter platelets in culture as well as persistent RhoA activation. Finally, we demonstrate that pharmacological inhibition of RhoA is capable of reversing the proplatelet defects mediated by PKCε inhibition. Collectively, these data indicate that by regulating RhoA activity, PKCε is a critical mediator of mouse proplatelet formation in vitro. PMID:23838351

  9. Targeting of Rho kinase ameliorates impairment of diabetic endothelial function in intrarenal artery.

    PubMed

    Yin, Hongping; Ru, Hailong; Yu, Liping; Kang, Yanhua; Lin, Guohua; Liu, Chuanfei; Sun, Lixian; Shi, Liyun; Sun, Qinghua; Liu, Cuiqing

    2013-10-14

    Endothelial dysfunction in kidney vasculature is the initial and key element for nephropathy in diabetes mellitus. Accumulating evidence suggests the protective role of Rho kinase inhibitors in endothelial dysfunction via modulating eNOS activity and NO production. However, the role of Rho kinase in diabetes-related endothelial dysfunction in kidney vasculature and the relevant mechanisms remain unknown. We assessed whether pharmacological inhibition of Rho kinase attenuates endothelial dysfunction in intrarenal arteries from type 1 diabetic rats. Fasudil, a Rho kinase inhibitor effectively decreased the phosphorylated level of MYPT1 without affecting the expression of ROCKs in the kidney. Fasudil treatment showed no improvement in diabetes-related abnormality in metabolic indices, but it significantly ameliorated endothelial dysfunction in intrarenal arteries and lessened the mesangial matrix expansion in the kidney cortex. Mechanistically, superoxide production in the intrarenal artery and NOX4 member of NADPH oxidase in the renal cortex that contribute to diabetic nephropathy were also prevented by the Rho kinase inhibitor. In conclusion, the present results indicate that Rho kinase is involved in endothelial dysfunction in type 1 diabetes via enhancement of oxidative stress and provides new evidence for Rho kinase inhibitors as potential therapeutic agents for the treatment of diabetic nephropathy.

  10. A novel role for RhoA GTPase in the regulation of airway smooth muscle contraction.

    PubMed

    Zhang, Wenwu; Huang, Youliang; Wu, Yidi; Gunst, Susan J

    2015-02-01

    Recent studies have demonstrated a novel molecular mechanism for the regulation of airway smooth muscle (ASM) contraction by RhoA GTPase. In ASM tissues, both myosin light chain (MLC) phosphorylation and actin polymerization are required for active tension generation. RhoA inactivation dramatically suppresses agonist-induced tension development and completely inhibits agonist-induced actin polymerization, but only slightly reduces MLC phosphorylation. The inhibition of MLC phosphatase does not reverse the effects of RhoA inactivation on contraction or actin polymerization. Thus, RhoA regulates ASM contraction through its effects on actin polymerization rather than MLC phosphorylation. Contractile stimulation of ASM induces the recruitment and assembly of paxillin, vinculin, and focal adhesion kinase (FAK) into membrane adhesion complexes (adhesomes) that regulate actin polymerization by catalyzing the activation of cdc42 GTPase by the G-protein-coupled receptor kinase-interacting target (GIT) - p21-activated kinase (PAK) - PAK-interacting exchange factor (PIX) complex. Cdc42 is a necessary and specific activator of the actin filament nucleation activator, N-WASp. The recruitment and activation of paxillin, vinculin, and FAK is prevented by RhoA inactivation, thus preventing cdc42 and N-WASp activation. We conclude that RhoA regulates ASM contraction by catalyzing the assembly and activation of membrane adhesome signaling modules that regulate actin polymerization, and that the RhoA-mediated assembly of adhesome complexes is a fundamental step in the signal transduction process in response to a contractile agonist.

  11. {pi} and {rho} mesons, and their diquark partners, from a contact interaction

    SciTech Connect

    Roberts, H. L. L.; Bashir, A.; Gutierrez-Guerrero, L. X.; Roberts, C. D.; Wilson, D. J.

    2011-06-15

    We present a unified Dyson-Schwinger equation treatment of static and electromagnetic properties of pseudoscalar and vector mesons, and scalar and axial-vector diquark correlations, based upon a vector-vector contact interaction. A basic motivation for this paper is the need to document a comparison between the electromagnetic form factors of mesons and those diquarks that play a material role in nucleon structure. A notable result, therefore, is the large degree of similarity between related meson and diquark form factors. The simplicity of the interaction enables computation of the form factors at arbitrarily large spacelike Q{sup 2}, which enables us to expose a zero in the {rho}-meson electric form factor at z{sub Q}{sup {rho}}{approx_equal}{radical}6m{sub {rho}}. Notably, r{sub {rho}}zQ{sup {rho}}{approx_equal}r{sub D}z{sub Q}{sup D}, where r{sub {rho}} and r{sub D} are, respectively, the electric radii of the {rho}-meson and deuteron.

  12. Detection and Quantification of ADP-Ribosylated RhoA/B by Monoclonal Antibody

    PubMed Central

    Rohrbeck, Astrid; Fühner, Viola; Schröder, Anke; Hagemann, Sandra; Vu, Xuan-Khang; Berndt, Sarah; Hust, Michael; Pich, Andreas; Just, Ingo

    2016-01-01

    Clostridium botulinum exoenzyme C3 is the prototype of C3-like ADP-ribosyltransferases that modify the GTPases RhoA, B, and C. C3 catalyzes the transfer of an ADP-ribose moiety from the co-substrate nicotinamide adenine dinucleotide (NAD) to asparagine-41 of Rho-GTPases. Although C3 does not possess cell-binding/-translocation domains, C3 is able to efficiently enter intact cells, including neuronal and macrophage-like cells. Conventionally, the detection of C3 uptake into cells is carried out via the gel-shift assay of modified RhoA. Since this gel-shift assay does not always provide clear, evaluable results an additional method to confirm the ADP-ribosylation of RhoA is necessary. Therefore, a new monoclonal antibody has been generated that specifically detects ADP-ribosylated RhoA/B, but not RhoC, in Western blot and immunohistochemical assay. The scFv antibody fragment was selected by phage display using the human naive antibody gene libraries HAL9/10. Subsequently, the antibody was produced as scFv-Fc and was found to be as sensitive as a commercially available RhoA antibody providing reproducible and specific results. We demonstrate that this specific antibody can be successfully applied for the analysis of ADP-ribosylated RhoA/B in C3-treated Chinese hamster ovary (CHO) and HT22 cells. Moreover, ADP-ribosylation of RhoA was detected within 10 min in C3-treated CHO wild-type cells, indicative of C3 cell entry. PMID:27043630

  13. Detection and Quantification of ADP-Ribosylated RhoA/B by Monoclonal Antibody.

    PubMed

    Rohrbeck, Astrid; Fühner, Viola; Schröder, Anke; Hagemann, Sandra; Vu, Xuan-Khang; Berndt, Sarah; Hust, Michael; Pich, Andreas; Just, Ingo

    2016-04-01

    Clostridium botulinum exoenzyme C3 is the prototype of C3-like ADP-ribosyltransferases that modify the GTPases RhoA, B, and C. C3 catalyzes the transfer of an ADP-ribose moiety from the co-substrate nicotinamide adenine dinucleotide (NAD) to asparagine-41 of Rho-GTPases. Although C3 does not possess cell-binding/-translocation domains, C3 is able to efficiently enter intact cells, including neuronal and macrophage-like cells. Conventionally, the detection of C3 uptake into cells is carried out via the gel-shift assay of modified RhoA. Since this gel-shift assay does not always provide clear, evaluable results an additional method to confirm the ADP-ribosylation of RhoA is necessary. Therefore, a new monoclonal antibody has been generated that specifically detects ADP-ribosylated RhoA/B, but not RhoC, in Western blot and immunohistochemical assay. The scFv antibody fragment was selected by phage display using the human naive antibody gene libraries HAL9/10. Subsequently, the antibody was produced as scFv-Fc and was found to be as sensitive as a commercially available RhoA antibody providing reproducible and specific results. We demonstrate that this specific antibody can be successfully applied for the analysis of ADP-ribosylated RhoA/B in C3-treated Chinese hamster ovary (CHO) and HT22 cells. Moreover, ADP-ribosylation of RhoA was detected within 10 min in C3-treated CHO wild-type cells, indicative of C3 cell entry.

  14. MiR-31 Regulates Rho-Associated Kinase-Myosin Light Chain (ROCK-MLC) Pathway and Inhibits Gastric Cancer Invasion: Roles of RhoA

    PubMed Central

    Chen, Zhuo; Liu, Shengnan; Xia, Yuan; Wu, Kejian

    2016-01-01

    Background This study evaluated how the expression of miR-31 can be used to detect gastric cancer (GC) to help illuminate the role of miR-31 and RhoA in GC cells. Material/Methods We carried out our experiments using tissue specimens from 70 GC patients. The relative expression of miR-31 and RhoA mRNA in tissues and cells was detected by RT-PCR. The expression level of RhoA protein was detected by immunohistochemistry. GC cell line BGC-823 was transfected with six groups of vectors: blank group, NC (negative control) group, miR-31 mimics group, miR-31 mimics + RhoA group, miR-31 mimics + ROCK group, and miR-31 mimics + MLCK agonist group. AGS cells were also transfected with six groups of vectors: blank group, NC group, miR-31 inhibitor group, miR-31 inhibitor + RhoA siRNA group, miR-31 inhibitor + ROCK siRNA group, and miR-31 inhibitor + MLCK inhibitor group. Transwell assay was performed to detect the invasion and migration of cells. The protein expression in different transfected groups was detected using Western blotting. Results GC tissues exhibited significantly lower levels of miR-31 expression compared to pericarcinous tissues (p<0.01). Moreover, a significantly higher expression of RhoA in GC tissues was observed (p<0.05). MiR-31 inhibited RhoA expression by binding to 3′UTR of mRNA, whereas miR-31 mimics significantly decreased the number of invaded and migrated cells (p<0.05). The activation of RhoA, ROCK, and phosphorylation of MLC remarkably exacerbate the invasion and migration ability of GC cells (p<0.05). Conclusions We found miR-31 could downregulate the ROCK/MLC pathway by inhibiting the expression of RhoA in order to suppress the invasion and migration of GC cells. PMID:27904131

  15. MiR-31 Regulates Rho-Associated Kinase-Myosin Light Chain (ROCK-MLC) Pathway and Inhibits Gastric Cancer Invasion: Roles of RhoA.

    PubMed

    Chen, Zhuo; Liu, Shengnan; Xia, Yuan; Wu, Kejian

    2016-12-01

    BACKGROUND This study evaluated how the expression of miR-31 can be used to detect gastric cancer (GC) to help illuminate the role of miR-31 and RhoA in GC cells. MATERIAL AND METHODS We carried out our experiments using tissue specimens from 70 GC patients. The relative expression of miR-31 and RhoA mRNA in tissues and cells was detected by RT-PCR. The expression level of RhoA protein was detected by immunohistochemistry. GC cell line BGC-823 was transfected with six groups of vectors: blank group, NC (negative control) group, miR-31 mimics group, miR-31 mimics + RhoA group, miR-31 mimics + ROCK group, and miR-31 mimics + MLCK agonist group. AGS cells were also transfected with six groups of vectors: blank group, NC group, miR-31 inhibitor group, miR-31 inhibitor + RhoA siRNA group, miR-31 inhibitor + ROCK siRNA group, and miR-31 inhibitor + MLCK inhibitor group. Transwell assay was performed to detect the invasion and migration of cells. The protein expression in different transfected groups was detected using Western blotting. RESULTS GC tissues exhibited significantly lower levels of miR-31 expression compared to pericarcinous tissues (p<0.01). Moreover, a significantly higher expression of RhoA in GC tissues was observed (p<0.05). MiR-31 inhibited RhoA expression by binding to 3'UTR of mRNA, whereas miR-31 mimics significantly decreased the number of invaded and migrated cells (p<0.05). The activation of RhoA, ROCK, and phosphorylation of MLC remarkably exacerbate the invasion and migration ability of GC cells (p<0.05). CONCLUSIONS We found miR-31 could downregulate the ROCK/MLC pathway by inhibiting the expression of RhoA in order to suppress the invasion and migration of GC cells.

  16. Involvement of rho-gtpases in fibroblast adhesion and fibronectine fibrillogenesis under stretch

    NASA Astrophysics Data System (ADS)

    Guignandon, A.; Lambert, C.; Rattner, A.; Servotte, S.; Lapiere, C.; Nusgens, B.; Vico, L.

    The Rho family small GTPases play a crucial role in mediating cellular adaptation to mechanical stimulation (MS), and possibly to microgravity (μg), through effects on the cytoskeleton and cell adhesion which is, in turn, mainly regulated by fibronectin fibrillogenesis (FnF). It remains unclear how mechanical stimulation is transduced to the Rho signaling pathways and how it impacts on fibronectin (fbn) fibrillogenesis (FnF). μg (2 days, mission STS-095) led to de-adhesion of fibroblasts and modification of the underlying extracellular matrix. To determine whether GTPases modulated FnF, we generated stable cell lines expressing high level of activated RhoA and Rac1 (QL) as compared to wild type (WI26-WT). After MS application [8% deformation, 1Hz, 15 min., 3 times/day for 1-2 days], we quantified focal adhesion (vinculin, paxillin, FAKY397), f-actin stress fibers (Sf) and FnF with home-developed softwares. We reported that after MS, Sf are more rapidly (30min) formed under the nucleus in Wi26-WT (+100%) and Rac1 (+200%) than in RhoA (+20%). Vinculin & paxillin were only restricted to the cell edge in static conditions and homogeneously distributed after MS in WT and Rac1. The relative area of contacts (vinculin & paxillin) was more dramatically enhanced by MS in Rac1 (+80%) than in WT (+40%) and RhoA (+25%) indicating that new focal contacts are formed under MS and supported the presence of Sf. MS Activation of FAK (FAKY397) was clear in WT and Rac1 and reduced in RhoA. FnF was restricted to cell-cell contacts zone without any change in the relative area of fbn after a 2-days MS. However we found more numerous spots of fbn at the cell center in Rac1 as compared with RhoA & WT suggesting that these fibrillar contacts will grow upon maturation and modulate FnF. The results indicate that MS induces formation of Sf and focal adhesions and enhances FF. RhoA has been shown to induce the formation of Sf and focal adhesions, and Rac1 activation decreases Rho activity in

  17. A study of the nuclear-medium influence on {rho}{sup 0} neutrinoproduction

    SciTech Connect

    Agababyan, N. M.; Ammosov, V. V.; Atayan, M.; Grigoryan, N.; Gulkanyan, H.; Ivanilov, A.A.; Karamyan, Zh.; Korotkov, V. A.

    2007-11-15

    The nuclear effects in {rho}{sup 0}-meson neutrino production are investigated using the data obtained with the SKAT bubble chamber. The nuclear-medium influence on the {rho}{sup 0} total yield and inclusive distributions on z = E{sub {rho}/}v and Feynman x{sub F} variable is found to be approximately the same as for pions. It is shown that these distributions with increasing A tend to shift toward smaller values of z and x{sub F}, thus indicating an increasing role of secondary intranuclear interactions. The predictions of a simple model incorporating the latter are found to be in qualitative agreement with experimental data.

  18. Mathematical cognitive style and arithmetic sign comprehension: a study of EEG alpha and theta activity.

    PubMed

    Earle, J B; Garcia-Dergay, P; Manniello, A; Dowd, C

    1996-01-01

    The localization of arithmetic sign comprehension was investigated using EEG spectral parameters as indicators of cortical engagement. Right-handed male subjects were selected on the basis of scores on the Mathematics Cognitive Style Survey and assigned to 2 groups, a 'left hemisphere oriented (LHO)' (N = 9) and 'right hemisphere oriented (RHO)' (N = 9) group. Subjects were presented with 4 conditions, a motoric baseline condition, two arithmetic fact retrieval tasks employing either a sign operator or verbal operator and a sign comprehension task which required subjects to fill in a missing sign (e.g. 6 ? 4 = 24). Both across subject correlational analysis of EEG alpha 1 asymmetry and performance as well as within subject analysis of condition means indicated a somewhat unique contribution of the right hemisphere to sign comprehension. LHO subjects exhibited greater relative left mid-temporal lobe activation than RHO subjects but less relative left frontal activation (theta band) than RHO subjects during the verbal operator task. It was tentatively concluded that this frontal lobe asymmetry difference was due to a mismatch in strategy preference and coding requirements among RHO subjects.

  19. Drosophila RhoGEF4 encodes a novel RhoA-specific guanine exchange factor that is highly expressed in the embryonic central nervous system.

    PubMed

    Nahm, Minyeop; Lee, Mihye; Baek, Seung-Hak; Yoon, Jin-Ho; Kim, Hong-Hee; Lee, Zang Hee; Lee, Seungbok

    2006-12-15

    Rho family small GTPases act as molecular switches that regulate neuronal morphogenesis, including axon growth and guidance, dendritic spine formation, and synapse formation. These proteins are positively regulated by guanine nucleotide exchange factors (GEFs) of the Dbl family. This study describes the identification and characterization of Drosophila RhoGEF4 (DRhoGEF4), a novel Dbl family protein that is specifically expressed in the central nervous system during Drosophila embryogenesis. The predicted amino acid sequence of DRhoGEF4 contains a Dbl homology (DH) domain and an adjacent C-terminal pleckstrin homology (PH) domain, which are most closely related to those of mammalian frabins. In this study, the DH-PH motif is shown to enhance the dissociation of GDP from either RhoA or Rac1 but not from Cdc42 in vitro. In addition, p21-binding domain pull-down assays demonstrate that DRhoGEF4 activates RhoA, but neither Rac1 nor Cdc42 in HEK293 cells. Finally, overexpression of DRhoGEF4 is able to induce assembly of stress fibers in cultured NIH3T3 cells. Taken together, these findings suggest that DRhoGEF4 may participate in cytoskeleton-related cellular events by specifically activating RhoA in neuronal morphogenesis.

  20. Inhibition of the RhoA GTPase Activity Increases Sensitivity of Melanoma Cells to UV Radiation Effects

    PubMed Central

    Espinha, Gisele; Osaki, Juliana Harumi; Costa, Erico Tosoni; Forti, Fabio Luis

    2016-01-01

    Ultraviolet radiation is the main cause of DNA damage to melanocytes and development of melanoma, one of the most lethal human cancers, which leads to metastasis due to uncontrolled cell proliferation and migration. These phenotypes are mediated by RhoA, a GTPase overexpressed or overactivated in highly aggressive metastatic tumors that plays regulatory roles in cell cycle progression and cytoskeleton remodeling. This work explores whether the effects of UV on DNA damage, motility, proliferation, and survival of human metastatic melanoma cells are mediated by the RhoA pathway. Mutant cells expressing dominant-negative (MeWo-RhoA-N19) or constitutively active RhoA (MeWo-RhoA-V14) were generated and subjected to UV radiation. A slight reduction in migration and invasion was observed in MeWo and MeWo-RhoA-V14 cells but not in MeWo-RhoA-N19 cells, which presented inefficient motility and invasiveness associated with stress fibers fragmentation. Proliferation and survival of RhoA-deficient cells were drastically reduced by UV compared to cells displaying normal or high RhoA activity, suggesting increased sensitivity to UV. Loss of RhoA activity also caused less efficient DNA repair, with elevated levels of DNA lesions such as strand breaks and cyclobutane pyrimidine dimers (CPDs). Thus, RhoA mediates genomic stability and represents a potential target for sensitizing metastatic tumors to genotoxic agents. PMID:26823948

  1. Inhibition of the RhoA GTPase Activity Increases Sensitivity of Melanoma Cells to UV Radiation Effects.

    PubMed

    Espinha, Gisele; Osaki, Juliana Harumi; Costa, Erico Tosoni; Forti, Fabio Luis

    2016-01-01

    Ultraviolet radiation is the main cause of DNA damage to melanocytes and development of melanoma, one of the most lethal human cancers, which leads to metastasis due to uncontrolled cell proliferation and migration. These phenotypes are mediated by RhoA, a GTPase overexpressed or overactivated in highly aggressive metastatic tumors that plays regulatory roles in cell cycle progression and cytoskeleton remodeling. This work explores whether the effects of UV on DNA damage, motility, proliferation, and survival of human metastatic melanoma cells are mediated by the RhoA pathway. Mutant cells expressing dominant-negative (MeWo-RhoA-N19) or constitutively active RhoA (MeWo-RhoA-V14) were generated and subjected to UV radiation. A slight reduction in migration and invasion was observed in MeWo and MeWo-RhoA-V14 cells but not in MeWo-RhoA-N19 cells, which presented inefficient motility and invasiveness associated with stress fibers fragmentation. Proliferation and survival of RhoA-deficient cells were drastically reduced by UV compared to cells displaying normal or high RhoA activity, suggesting increased sensitivity to UV. Loss of RhoA activity also caused less efficient DNA repair, with elevated levels of DNA lesions such as strand breaks and cyclobutane pyrimidine dimers (CPDs). Thus, RhoA mediates genomic stability and represents a potential target for sensitizing metastatic tumors to genotoxic agents.

  2. RhoA-Mediated Functions in C3H10T1/2 Osteoprogenitors Are Substrate Topography Dependent.

    PubMed

    Ogino, Yoichiro; Liang, Ruiwei; Mendonça, Daniela B S; Mendonça, Gustavo; Nagasawa, Masako; Koyano, Kiyoshi; Cooper, Lyndon F

    2016-03-01

    Surface topography broadly influences cellular responses. Adherent cell activities are regulated, in part, by RhoA, a member of the Rho-family of GTPases. In this study, we evaluated the influence of surface topography on RhoA activity and associated cellular functions. The murine mesenchymal stem cell line C3H10T1/2 cells (osteoprogenitor cells) were cultured on titanium substrates with smooth topography (S), microtopography (M), and nanotopography (N) to evaluate the effect of surface topography on RhoA-mediated functions (cell spreading, adhesion, migration, and osteogenic differentiation). The influence of RhoA activity in the context of surface topography was also elucidated using RhoA pharmacologic inhibitor. Following adhesion, M and N adherent cells developed multiple projections, while S adherent cells had flattened and widespread morphology. RhoA inhibitor induced remarkable longer and thinner cytoplasmic projections on all surfaces. Cell adhesion and osteogenic differentiation was topography dependent with S < M and N surfaces. RhoA inhibition increased adhesion on S and M surfaces, but not N surfaces. Cell migration in a wound healing assay was greater on S versus M versus N surfaces and RhoA inhibitor increased S adherent cell migration, but not N adherent cell migration. RhoA inhibitor enhanced osteogenic differentiation in S adherent cells, but not M or N adherent cells. RhoA activity was surface topography roughness dependent (S < M, N). RhoA activity and -mediated functions are influenced by surface topography. Smooth surface adherent cells appear highly sensitive to RhoA function, while nano-scale topography adherent cell may utilize alternative cellular signaling pathway(s) to influence adherent cellular functions regardless of RhoA activity.

  3. Cloning of the RHO1 gene from Candida albicans and its regulation of beta-1,3-glucan synthesis.

    PubMed Central

    Kondoh, O; Tachibana, Y; Ohya, Y; Arisawa, M; Watanabe, T

    1997-01-01

    The Saccharomyces cerevisiae RHO1 gene encodes a low-molecular-weight GTPase. One of its recently identified functions is the regulation of beta-1,3-glucan synthase, which synthesizes the main component of the fungal cell wall (J. Drgonova et al., Science 272:277-279, 1996; T. Mazur and W. Baginsky, J. Biol. Chem. 271:14604-14609, 1996; and H. Qadota et al., Science 272:279-281, 1996). From the opportunistic pathogenic fungus Candida albicans, we cloned the RHO1 gene by the PCR and cross-hybridization methods. Sequence analysis revealed that the Candida RHO1 gene has a 597-nucleotide region which encodes a putative 22.0-kDa peptide. The deduced amino acid sequence predicts that Candida albicans Rho1p is 82.9% identical to Saccharomyces Rho1p and contains all the domains conserved among Rho-type GTPases from other organisms. The Candida albicans RHO1 gene could rescue a S. cerevisiae strain containing a rho1 deletion. Furthermore, recombinant Candida albicans Rho1p could reactivate the beta-1,3-glucan synthesis activities of both C. albicans and S. cerevisiae membranes in which endogenous Rho1p had been depleted by Tergitol NP-40-NaCl treatment. Candida albicans Rho1p was copurified with the beta-1,3-glucan synthase putative catalytic subunit, Candida albicans Gsc1p, by product entrapment. Candida albicans Rho1p was shown to interact directly with Candida albicans Gsc1p in a ligand overlay assay and a cross-linking study. These results indicate that Candida albicans Rho1p acts in the same manner as Saccharomyces cerevisiae Rho1p to regulate beta-1,3-glucan synthesis. PMID:9401032

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

    PubMed

    Somers, W Gregory; Saint, Robert

    2003-01-01

    The mechanism that positions the cytokinetic contractile ring is unknown, but derives from the spindle midzone. We show that an interaction between the Rho GTP exchange factor, Pebble, and the Rho family GTPase-activating protein, RacGAP50C, connects the contractile ring to cortical microtubules at the site of furrowing in D. melanogaster cells. Pebble regulates actomyosin organization, while RacGAP50C and its binding partner, the Pavarotti kinesin-like protein, regulate microtubule bundling. All three factors are required for cytokinesis. As furrowing begins, these proteins colocalize to a cortical equatorial ring. We propose that RacGAP50C-Pavarotti complexes travel on cortical microtubules to the cell equator, where they associate with the Pebble RhoGEF to position contractile ring formation and coordinate F-actin and microtubule remodeling during cytokinesis.

  5. Nu Rho Psi, The National Honor Society in Neuroscience: A decade of progress

    PubMed Central

    Hesp, Zoe C.; Cousens, Graham A.; Becker, Lora; Zee, Michele C.; Mickley, G. Andrew

    2016-01-01

    Nu Rho Psi, the National Honor Society in Neuroscience, celebrates its 10th anniversary by reflecting back upon a decade’s worth of growth, successes, and accomplishments of its membership. Fundamentally, Nu Rho Psi seeks to engage the nation’s best and brightest science students early in their educational pursuits and steer them towards future careers in neuroscience, thereby driving higher quality neuroscience education and research at all levels. This article details the history of Nu Rho Psi since its founding by the Faculty for Undergraduate Neuroscience (FUN) and reviews the current programs, benefits, and future initiatives of the Society. We make the case that Nu Rho Psi has enhanced the opportunities for undergraduate students of neuroscience and created a new culture among this vital cohort of budding scientists, reminiscent of the substantial network of faculty educators and departments of neuroscience established by FUN. PMID:27385933

  6. Measurement of the CP asymmetry and branching fraction of B0-->rho0K0.

    PubMed

    Aubert, B; Barate, R; Bona, M; Boutigny, D; Couderc, F; Karyotakis, Y; Lees, J P; Poireau, V; Tisserand, V; Zghiche, A; Grauges, E; Palano, A; Chen, J C; Qi, N D; Rong, G; Wang, P; Zhu, Y S; Eigen, G; Ofte, I; Stugu, B; Abrams, G S; Battaglia, M; Brown, D N; Button-Shafer, J; Cahn, R N; Charles, E; Gill, M S; Groysman, Y; Jacobsen, R G; Kadyk, J A; Kerth, L T; Kolomensky, Yu G; Kukartsev, G; Lynch, G; Mir, L M; Orimoto, T J; Pripstein, M; Roe, N A; Ronan, M T; Wenzel, W A; del Amo Sanchez, P; Barrett, M; Ford, K E; Harrison, T J; Hart, A J; Hawkes, C M; Morgan, S E; Watson, A T; Held, T; Koch, H; Lewandowski, B; Pelizaeus, M; Peters, K; Schroeder, T; Steinke, M; Boyd, J T; Burke, J P; Cottingham, W N; Walker, D; Cuhadar-Donszelmann, T; Fulsom, B G; Hearty, C; Knecht, N S; Mattison, T S; McKenna, J A; Khan, A; Kyberd, P; Saleem, M; Sherwood, D J; Teodorescu, L; Blinov, V E; Bukin, A D; Druzhinin, V P; Golubev, V B; Onuchin, A P; Serednyakov, S I; Skovpen, Yu I; Solodov, E P; Todyshev, K Yu; Best, D S; Bondioli, M; Bruinsma, M; Chao, M; Curry, S; Eschrich, I; Kirkby, D; Lankford, A J; Lund, P; Mandelkern, M; Mommsen, R K; Roethel, W; Stoker, D P; Abachi, S; Buchanan, C; Foulkes, S D; Gary, J W; Long, O; Shen, B C; Wang, K; Zhang, L; Hadavand, H K; Hill, E J; Paar, H P; Rahatlou, S; Sharma, V; Berryhill, J W; Campagnari, C; Cunha, A; Dahmes, B; Hong, T M; Kovalskyi, D; Richman, J D; Beck, T W; Eisner, A M; Flacco, C J; Heusch, C A; Kroseberg, J; Lockman, W S; Nesom, G; Schalk, T; Schumm, B A; Seiden, A; Spradlin, P; Williams, D C; Wilson, M G; Albert, J; Chen, E; Dvoretskii, A; Fang, F; Hitlin, D G; Narsky, I; Piatenko, T; Porter, F C; Ryd, A; Samuel, A; Mancinelli, G; Meadows, B T; Mishra, K; Sokoloff, M D; Blanc, F; Bloom, P C; Chen, S; Ford, W T; Hirschauer, J F; Kreisel, A; Nagel, M; Nauenberg, U; Olivas, A; Ruddick, W O; Smith, J G; Ulmer, K A; Wagner, S R; Zhang, J; Chen, A; Eckhart, E A; Soffer, A; Toki, W H; Wilson, R J; Winklmeier, F; Zeng, Q; Altenburg, D D; Feltresi, E; Hauke, A; Jasper, H; Petzold, A; Spaan, B; Brandt, T; Klose, V; Lacker, H M; Mader, W F; Nogowski, R; Schubert, J; Schubert, K R; Schwierz, R; Sundermann, J E; Volk, A; Bernard, D; Bonneaud, G R; Grenier, P; Latour, E; Thiebaux, Ch; Verderi, M; Clark, P J; Gradl, W; Muheim, F; Playfer, S; Robertson, A I; Xie, Y; Andreotti, M; Bettoni, D; Bozzi, C; Calabrese, R; Cibinetto, G; Luppi, E; Negrini, M; Petrella, A; Piemontese, L; Prencipe, E; Anulli, F; Baldini-Ferroli, R; Calcaterra, A; de Sangro, R; Finocchiaro, G; Pacetti, S; Patteri, P; Peruzzi, I M; Piccolo, M; Rama, M; Zallo, A; Buzzo, A; Capra, R; Contri, R; Lo Vetere, M; Macri, M M; Monge, M R; Passaggio, S; Patrignani, C; Robutti, E; Santroni, A; Tosi, S; Brandenburg, G; Chaisanguanthum, K S; Morii, M; Wu, J; Dubitzky, R S; Marks, J; Schenk, S; Uwer, U; Bard, D J; Bhimji, W; Bowerman, D A; Dauncey, P D; Egede, U; Flack, R L; Nash, J A; Nikolich, M B; Panduro Vazquez, W; Behera, P K; Chai, X; Charles, M J; Mallik, U; Meyer, N T; Ziegler, V; Cochran, J; Crawley, H B; Dong, L; Eyges, V; Meyer, W T; Prell, S; Rosenberg, E I; Rubin, A E; Gritsan, A V; Denig, A G; Fritsch, M; Schott, G; Arnaud, N; Davier, M; Grosdidier, G; Höcker, A; Le Diberder, F; Lepeltier, V; Lutz, A M; Oyanguren, A; Pruvot, S; Rodier, S; Roudeau, P; Schune, M H; Stocchi, A; Wang, W F; Wormser, G; Cheng, C H; Lange, D J; Wright, D M; Chavez, C A; Forster, I J; Fry, J R; Gabathuler, E; Gamet, R; George, K A; Hutchcroft, D E; Payne, D J; Schofield, K C; Touramanis, C; Bevan, A J; Di Lodovico, F; Menges, W; Sacco, R; Cowan, G; Flaecher, H U; Hopkins, D A; Jackson, P S; McMahon, T R; Ricciardi, S; Salvatore, F; Wren, A C; Brown, D N; Davis, C L; Allison, J; Barlow, N R; Barlow, R J; Chia, Y M; Edgar, C L; Lafferty, G D; Naisbit, M T; Williams, J C; Yi, J I; Chen, C; Hulsbergen, W D; Jawahery, A; Lae, C K; Roberts, D A; Simi, G; Blaylock, G; Dallapiccola, C; Hertzbach, S S; Li, X; Moore, T B; Saremi, S; Staengle, H; Cowan, R; Sciolla, G; Sekula, S J; Spitznagel, M; Taylor, F; Yamamoto, R K; Kim, H; McLachlin, S E; Patel, P M; Robertson, S H; Lazzaro, A; Lombardo, V; Palombo, F; Bauer, J M; Cremaldi, L; Eschenburg, V; Godang, R; Kroeger, R; Sanders, D A; Summers, D J; Zhao, H W; Brunet, S; Côté, D; Simard, M; Taras, P; Viaud, F B; Nicholson, H; Cavallo, N; De Nardo, G; Fabozzi, F; Gatto, C; Lista, L; Monorchio, D; Paolucci, P; Piccolo, D; Sciacca, C; Baak, M; Raven, G; Snoek, H L; Jessop, C P; Losecco, J M; Allmendinger, T; Benelli, G; Gan, K K; Honscheid, K; Hufnagel, D; Jackson, P D; Kagan, H; Kass, R; Rahimi, A M; Ter-Antonyan, R; Wong, Q K; Blount, N L; Brau, J; Frey, R; Igonkina, O; Lu, M; Rahmat, R; Sinev, N B; Strom, D; Strube, J; Torrence, E; Gaz, A; Margoni, M; Morandin, M; Pompili, A; Posocco, M; Rotondo, M; Simonetto, F; Stroili, R; Voci, C; Benayoun, M; Chauveau, J; Briand, H; David, P; Del Buono, L; de la Vaissière, Ch; Hamon, O; Hartfiel, B L; John, M J J; Leruste, Ph; Malclès, J; Ocariz, J; Roos, L; Therin, G; Gladney, L; Panetta, J; Biasini, M; Covarelli, R; Angelini, C; Batignani, G; Bettarini, S; Bucci, F; Calderini, G; Carpinelli, M; Cenci, R; Forti, F; Giorgi, M A; Lusiani, A; Marchiori, G; Mazur, M A; Morganti, M; Neri, N; Paoloni, E; Rizzo, G; Walsh, J J; Haire, M; Judd, D; Wagoner, D E; Biesiada, J; Danielson, N; Elmer, P; Lau, Y P; Lu, C; Olsen, J; Smith, A J S; Telnov, A V; Bellini, F; Cavoto, G; D'Orazio, A; del Re, D; Di Marco, E; Faccini, R; Ferrarotto, F; Ferroni, F; Gaspero, M; Li Gioi, L; Mazzoni, M A; Morganti, S; Piredda, G; Polci, F; Safai Tehrani, F; Voena, C; Ebert, M; Schröder, H; Waldi, R; Adye, T; De Groot, N; Franek, B; Olaiya, E O; Wilson, F F; Aleksan, R; Emery, S; Gaidot, A; Ganzhur, S F; Hamel de Monchenault, G; Kozanecki, W; Legendre, M; Vasseur, G; Yèche, Ch; Zito, M; Chen, X R; Liu, H; Park, W; Purohit, M V; Wilson, J R; Allen, M T; Aston, D; Bartoldus, R; Bechtle, P; Berger, N; Claus, R; Coleman, J P; Convery, M R; Cristinziani, M; Dingfelder, J C; Dorfan, J; Dubois-Felsmann, G P; Dujmic, D; Dunwoodie, W; Field, R C; Glanzman, T; Gowdy, S J; Graham, M T; Halyo, V; Hast, C; Hryn'ova, T; Innes, W R; Kelsey, M H; Kim, P; Leith, D W G S; Li, S; Luitz, S; Luth, V; Lynch, H L; MacFarlane, D B; Marsiske, H; Messner, R; Muller, D R; O'Grady, C P; Ozcan, V E; Perazzo, A; Perl, M; Pulliam, T; Ratcliff, B N; Roodman, A; Salnikov, A A; Schindler, R H; Schwiening, J; Snyder, A; Stelzer, J; Su, D; Sullivan, M K; Suzuki, K; Swain, S K; Thompson, J M; Va'vra, J; van Bakel, N; Weaver, M; Weinstein, A J R; Wisniewski, W J; Wittgen, M; Wright, D H; Yarritu, A K; Yi, K; Young, C C; Burchat, P R; Edwards, A J; Majewski, S A; Petersen, B A; Roat, C; Wilden, L; Ahmed, S; Alam, M S; Bula, R; Ernst, J A; Jain, V; Pan, B; Saeed, M A; Wappler, F R; Zain, S B; Bugg, W; Krishnamurthy, M; Spanier, S M; Eckmann, R; Ritchie, J L; Satpathy, A; Schilling, C J; Schwitters, R F; Izen, J M; Lou, X C; Ye, S; Bianchi, F; Gallo, F; Gamba, D; Bomben, M; Bosisio, L; Cartaro, C; Cossutti, F; Della Ricca, G; Dittongo, S; Lanceri, L; Vitale, L; Azzolini, V; Martinez-Vidal, F; Banerjee, Sw; Bhuyan, B; Brown, C M; Fortin, D; Hamano, K; Kowalewski, R; Nugent, I M; Roney, J M; Sobie, R J; Back, J J; Harrison, P F; Latham, T E; Mohanty, G B; Pappagallo, M; Band, H R; Chen, X; Cheng, B; Dasu, S; Datta, M; Flood, K T; Hollar, J J; Kutter, P E; Mellado, B; Mihalyi, A; Pan, Y; Pierini, M; Prepost, R; Wu, S L; Yu, Z; Neal, H

    2007-02-02

    We present a measurement of the branching fraction and time-dependent CP asymmetry of B(0)-->rho0K0. The results are obtained from a data sample of 227x10(6) Upsilon(4S)-->BB decays collected with the BABAR detector at the PEP-II asymmetric-energy B factory at Stanford Linear Accelerator Center. From a time-dependent maximum likelihood fit yielding 111+/-19 signal events, we find B(B(0)-->rho0K0)=(4.9+/-0.8+/-0.9)x10(-6), where the first error is statistical and the second systematic. We report the measurement of the CP parameters S(rho)(0)K(0)S=0.20+/-0.52+/-0.24 and C(rho)(0)K(0)S=0.64+/-0.41+/-0.20.

  7. Lovastatin-induced RhoA modulation and its effect on senescence in prostate cancer cells

    SciTech Connect

    Lee, Jeeyun; Lee, Inkyoung; Park, Chaehwa; Kang, Won Ki . E-mail: wkkang@smc.samsung.co.kr

    2006-01-20

    Lovastatin inhibits a 3-hydroxy 3-methylglutaryl coenzyme A reductase and prevents the synthesis of cholesterol precursors, such as farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP), responsible for important cell signaling in cell proliferation and migration. Recently, the anti-cancer effect of lovastatin has been suggested in various tumor types. In this study, we showed that a low dose lovastatin induced senescence and G1 cell cycle arrest in human prostate cancer cells. Addition of GGPP or mevalonate, but not FPP, prevented the lovastatin-induced G1 phase cell cycle arrest and cell senescence. We found that constitutively active RhoA (caRhoA) reversed lovastatin-induced senescence in caRhoA-transfected PC-3 cells. Thus, we postulate that modulation of RhoA may be critical in lovastatin-induced senescence in PC-3 cells.

  8. The structure of RNA-free Rho termination factor indicates a dynamic mechanism of transcript capture.

    PubMed

    Canals, Albert; Usón, Isabel; Coll, Miquel

    2010-07-02

    The Rho factor is a ring-shaped ATP-dependent helicase that mediates transcription termination in most prokaryotic cells by disengaging the transcription elongation complex formed by the RNA polymerase, DNA, and the nascent RNA transcript. The crystal structures of key intermediates along the kinetic pathway of RNA binding to Rho unveiled an unprecedented mode of helicase loading and provided a model for the ATP turnover coupled to coordinated strand movement. Here we report the structure of the early RNA-free state of Rho, which had eluded crystallization for many years but now completes the series. The structure allows the characterization of the apo-form Rho from Thermotoga maritima to 2.3 A resolution, reveals an RNA-recruiting site that becomes hidden after occupancy of the adjacent specific primary RNA-binding site, and suggests an enriched model for mRNA capture that is consistent with previous data.

  9. Interpreting EEG alpha activity.

    PubMed

    Bazanova, O M; Vernon, D

    2014-07-01

    Exploring EEG alpha oscillations has generated considerable interest, in particular with regards to the role they play in cognitive, psychomotor, psycho-emotional and physiological aspects of human life. However, there is no clearly agreed upon definition of what constitutes 'alpha activity' or which of the many indices should be used to characterize it. To address these issues this review attempts to delineate EEG alpha-activity, its physical, molecular and morphological nature, and examine the following indices: (1) the individual alpha peak frequency; (2) activation magnitude, as measured by alpha amplitude suppression across the individual alpha bandwidth in response to eyes opening, and (3) alpha "auto-rhythmicity" indices: which include intra-spindle amplitude variability, spindle length and steepness. Throughout, the article offers a number of suggestions regarding the mechanism(s) of alpha activity related to inter and intra-individual variability. In addition, it provides some insights into the various psychophysiological indices of alpha activity and highlights their role in optimal functioning and behavior.

  10. Errors in quantitative T1rho imaging and the correction methods

    PubMed Central

    2015-01-01

    The spin-lattice relaxation time constant in rotating frame (T1rho) is useful for assessment of the properties of macromolecular environment inside tissue. Quantification of T1rho is found promising in various clinical applications. However, T1rho imaging is prone to image artifacts and quantification errors, which remains one of the greatest challenges to adopt this technique in routine clinical practice. The conventional continuous wave spin-lock is susceptible to B1 radiofrequency (RF) and B0 field inhomogeneity, which appears as banding artifacts in acquired images. A number of methods have been reported to modify T1rho prep RF pulse cluster to mitigate this effect. Adiabatic RF pulse can also be used for spin-lock with insensitivity to both B1 RF and B0 field inhomogeneity. Another source of quantification error in T1rho imaging is signal evolution during imaging data acquisition. Care is needed to affirm such error does not take place when specific pulse sequence is used for imaging data acquisition. Another source of T1rho quantification error is insufficient signal-to-noise ratio (SNR), which is common among various quantitative imaging approaches. Measurement of T1rho within an ROI can mitigate this issue, but at the cost of reduced resolution. Noise-corrected methods are reported to address this issue in pixel-wise quantification. For certain tissue type, T1rho quantification can be confounded by magic angle effect and the presence of multiple tissue components. Review of these confounding factors from inherent tissue properties is not included in this article. PMID:26435922

  11. Galectin-8 Promotes Cytoskeletal Rearrangement in Trabecular Meshwork Cells through Activation of Rho Signaling

    PubMed Central

    Cao, Zhiyi; Gyawali, Smita; Gong, Haiyan; Soza, Andrea; González, Alfonso; Panjwani, Noorjahan

    2012-01-01

    Purpose The trabecular meshwork (TM) cell-matrix interactions and factors that influence Rho signaling in TM cells are thought to play a pivotal role in the regulation of aqueous outflow. The current study was designed to evaluate the role of a carbohydrate-binding protein, galectin-8 (Gal8), in TM cell adhesion and Rho signaling. Methods Normal human TM cells were assayed for Gal8 expression by immunohistochemistry and Western blot analysis. To assess the role of Gal8 in TM cell adhesion and Rho signaling, the cell adhesion and spreading assays were performed on Gal8-coated culture plates in the presence and the absence of anti-β1 integrin antibody and Rho and Rho-kinase inhibitors. In addition, the effect of Gal8-mediated cell-matrix interactions on TM cell cytoskeleton arrangement and myosin light chain 2 (MLC2) phosphorylation was examined. Principal Findings We demonstrate here that Gal8 is expressed in the TM and a function-blocking anti-β1 integrin antibody inhibits the adhesion and spreading of TM cells to Gal8-coated wells. Cell spreading on Gal8 substratum was associated with the accumulation of phosphorylated myosin light chain and the formation of stress fibers that was inhibited by the Rho inhibitor, C3 transferase, as well as by the Rho-kinase inhibitor, Y27632. Conclusions/Significance The above findings present a novel function for Gal8 in activating Rho signaling in TM cells. This function may allow Gal8 to participate in the regulation of aqueous outflow. PMID:22973445

  12. RhoA Modulates Smad Signaling during Transforming Growth Factor-β-induced Smooth Muscle Differentiation*

    PubMed Central

    Chen, Shiyou; Crawford, Michelle; Day, Regina M.; Briones, Victorino R.; Leader, Jennifer E.; Jose, Pedro A.; Lechleider, Robert J.

    2007-01-01

    We recently reported that transforming growth factor (TGF)-β induced the neural crest stem cell line Monc-1 to differentiate into a spindle-like contractile smooth muscle cell (SMC) phenotype and that Smad signaling played an important role in this phenomenon. In addition to Smad signaling, other pathways such as mitogen-activated protein kinase (MAPK), phosphoinositol-3 kinase, and RhoA have also been shown to mediate TGF-β actions. The objectives of this study were to examine whether these signaling pathways contribute to TGF-β-induced SMC development and to test whether Smad signaling cross-talks with other pathway(s) during SMC differentiation induced by TGF-β. We demonstrate here that RhoA signaling is critical to TGF-β-induced SMC differentiation. RhoA kinase (ROCK) inhibitor Y27632 significantly blocks the expression of multiple SMC markers such as smooth muscle α-actin, SM22α, and calponin in TGF-β-treated Monc-1 cells. In addition, Y27632 reversed the cell morphology and abolished the contractility of TGF-β-treated cells. RhoA signaling was activated as early as 5 min following TGF-β addition. Dominant negative RhoA blocked nuclear translocation of Smad2 and Smad3 because of the inhibition of phosphorylation of both Smads and inhibited Smad-dependent SBE promoter activity, whereas constitutively active RhoA significantly enhanced SBE promoter activity. Consistent with these results, C3 exotoxin, an inhibitor of RhoA activation, significantly attenuated SBE promoter activity and inhibited Smad nuclear translocation. Taken together, these data point to a new role for RhoA as a modulator of Smad activation while regulating TGF-β-induced SMC differentiation. PMID:16317010

  13. Effects of phased joint intervention on Rho/ROCK expression levels in patients with portal hypertension

    PubMed Central

    Shi, Min; Wei, Jue; Meng, Wen-Ying; Wang, Na; Wang, Ting; Wang, Yu-Gang

    2016-01-01

    The current study investigated the effects of phased joint intervention on clinical efficacy and Rho/Rho-associated coil protein kinase (ROCK) expression in patients with portal hypertension complicated by esophageal variceal bleeding (EVB) and hypersplenism. Patients with portal hypertension (n=53) caused by liver cirrhosis complicated by EVB and hypersplenism treated with phased joint intervention were assessed, and portal hemodynamics, blood, liver function, complications, and rebleeding incidence were analyzed. Reverse transcription-quantitative polymerase chain reaction was used to measure Rho, ROCK1 and ROCK2 mRNA expression levels in peripheral blood mononuclear cells prior to and following phased joint intervention, and western blotting was employed to determine the protein expression levels of Rho, ROCK1, ROCK2, phosphorylated (p) myosin phosphatase target subunit 1 (MYPT1) and total-MYPT1. All patients underwent an emergency assessment of hemostasis with a 100% success rate. Varicose veins were alleviated, and portal hemodynamics and liver function improved following intervention. Furthermore, preoperative and postoperative expression levels of Rho, ROCK1 and ROCK2 mRNA were higher compared with the control group. Notably, the mRNA expression levels of Rho, ROCK1 and ROCK2 in the postoperative group were significantly lower when compared with the preoperative group. Protein expression levels of Rho, ROCK1, ROCK2 and pMYPT1 in the postoperative group were lower, as compared with the preoperative group. Concentration levels of transforming growth factor-β1, connective tissue growth factor and platelet-derived growth factor in peripheral blood were significantly reduced following phased joint intervention. Therefore, the present findings demonstrated that phased joint intervention is able to effectively treat EVB and hypersplenism, and improve liver function. The efficacy of phased joint intervention may be associated with its role in the regulation of the

  14. Leptin augments coronary vasoconstriction and smooth muscle proliferation via a Rho-kinase-dependent pathway.

    PubMed

    Noblet, Jillian N; Goodwill, Adam G; Sassoon, Daniel J; Kiel, Alexander M; Tune, Johnathan D

    2016-05-01

    Leptin has been implicated as a key upstream mediator of pathways associated with coronary vascular dysfunction and disease. The purpose of this investigation was to test the hypothesis that leptin modifies the coronary artery proteome and promotes increases in coronary smooth muscle contraction and proliferation via influences on Rho kinase signaling. Global proteomic assessment of coronary arteries from lean swine cultured with obese concentrations of leptin (30 ng/mL) for 3 days revealed significant alterations in the coronary artery proteome (68 proteins) and identified an association between leptin treatment and calcium signaling/contraction (four proteins) and cellular growth and proliferation (35 proteins). Isometric tension studies demonstrated that both acute (30 min) and chronic (3 days, serum-free media) exposure to obese concentrations of leptin potentiated depolarization-induced contraction of coronary arteries. Inhibition of Rho kinase significantly reduced leptin-mediated increases in coronary artery contractions. The effects of leptin on the functional expression of Rho kinase were time-dependent, as acute treatment increased Rho kinase activity while chronic (3 day) exposure was associated with increases in Rho kinase protein abundance. Proliferation assays following chronic leptin administration (8 day, serum-containing media) demonstrated that leptin augmented coronary vascular smooth muscle proliferation and increased Rho kinase activity. Inhibition of Rho kinase significantly reduced these effects of leptin. Taken together, these findings demonstrate that leptin promotes increases in coronary vasoconstriction and smooth muscle proliferation and indicate that these phenotypic effects are associated with alterations in the coronary artery proteome and dynamic effects on the Rho kinase pathway.

  15. T1rho Magnetic Resonance Imaging at 3T Detects Knee Cartilage Changes After Viscosupplementation.

    PubMed

    Shah, Roshan P; Stambough, Jeffrey B; Fenty, Matthew; Mauck, Robert L; Kelly, John D; Reddy, Ravinder; Tjoumakaris, Fotios P

    2015-07-01

    Viscosupplementation may affect cartilage. Changes in T1rho magnetic resonance imaging (MRI) relaxation times correlate with proteoglycan changes in cartilage. The authors hypothesized that T1rho MRI will show an improvement in proteoglycan content at 6 weeks and 3 months after viscosupplementation and that this improvement will correlate with functional outcome scores. Ten patients (mean age, 56 years; Kellgren-Lawrence grade 1 or 2) underwent T1rho MRI at baseline, 6 weeks, and 3 months after viscosupplementation. Volumetric T1rho means were calculated by depth and region. Visual analog scale (VAS), International Knee Documentation Committee (IKDC), and Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) scores were obtained. Mean T1rho values decreased in the superficial patella at 6 weeks (10.3%, P=.002) and 3 months (7.9%, P=.018) and in the middle patella at 6 weeks (7.0%, P=.014) compared with baseline values. Deep patella T1rho values increased at 3 months compared with 6 weeks (9.9%, P=.033), returning to values similar to baseline. Mean T1rho values increased in the deep tibia at 6 weeks (4.7%, P=.048) and in the middle tibia (5.2%, P=.004) and deep tibia (11.2%, P=.002) at 3 months compared with baseline. At 6 weeks, improvement was seen in VAS (5.9 to 3.9, P<.01), IKDC-9 (55.3 to 63.7, P=.03), and WOMAC (43.9 to 32.8, P=.03) scores. Functional VAS (4.0, P=.02), IKDC-9 (67.8, P=.04), and WOMAC (30.0, P=.04) scores remained better at 3 months. T1rho MRI is a feasible noninvasive method of studying molecular changes in cartilage. Some segments improved after viscosupplementation, and others worsened, possibly reflecting natural history or symptom relief and subsequent increase in activity-related wear.

  16. Analysis of a minimal Rho-GTPase circuit regulating cell shape

    NASA Astrophysics Data System (ADS)

    Holmes, William R.; Edelstein-Keshet, Leah

    2016-08-01

    Networks of Rho-family GTPases regulate eukaryotic cell polarization and motility by controlling assembly and contraction of the cytoskeleton. The mutually inhibitory Rac-Rho circuit is emerging as a central, regulatory hub that can affect the shape and motility phenotype of eukaryotic cells. Recent experimental manipulation of the amounts of Rac and Rho or their regulators (guanine nucleotide-exchange factors, GTPase-activating proteins, guanine nucleotide dissociation inhibitors) have been shown to bias the prevalence of these different states and promote transitions between them. Here we show that part of this data can be understood in terms of inherent Rac-Rho mutually inhibitory dynamics. We analyze a spatio-temporal mathematical model of Rac-Rho dynamics to produce a detailed set of predictions of how parameters such as GTPase rates of activation and total amounts affect cell decisions (such as Rho-dominated contraction, Rac-dominated spreading, and spatially segregated Rac-Rho polarization). We find that in some parameter regimes, a cell can take on any of these three fates depending on its environment or stimuli. We also predict how experimental manipulations (corresponding to parameter variations) can affect cell shapes observed. Our methods are based on local perturbation analysis (a kind of nonlinear stability analysis), and an approximation of nonlinear feedback by sharp switches. We compare the Rac-Rho model to an even simpler single-GTPase (‘wave-pinning’) model and demonstrate that the overall behavior is inherent to GTPase properties, rather than stemming solely from network topology.

  17. An ALMA Survey of Planet Forming Disks in Rho Ophiuchus

    NASA Astrophysics Data System (ADS)

    Guilfoil Cox, Erin; Looney, Leslie; Harris, Robert J.; Dong, Jiayin; Segura-Cox, Dominique; Tobin, John J.; Sadavoy, Sarah; Li, Zhi-Yun; Dunham, Michael; Perez, Laura M.; Chandler, Claire J.; Kratter, Kaitlin M.; Melis, Carl; Chiang, Hsin-Fang

    2017-01-01

    Relatively evolved (~ 1 Myr old) protostars with little residual natal envelope, but massive disks, are commonly assumed to be the sites of ongoing planet formation. Critical to our study of these objects is information about the available mass reservior and dust structure, as they directly tie in to how much mass is available for planets as well as the modes of planet formation that occur (i.e., core-accretion vs. gravitational instability). Millimeter-wave observations provide this critical information as continuum emission is relatively optically thin, allowing for mass estimates, and the availability of high-resolution interferometry, allowing structure constraints. We present high-resolution observations of the population of Class II protostars in the Rho-Ophiuchus cloud (d ~ 130 pc). Our survey observed ~50 of these older protostars at 870µm, using the Atacama Large Millimeter/submillimeter Array (ALMA). Out of these sources, there are ~10 transition disks, where we see a ring of dust emission surrounding the central protostar -- indicative of ongoing planet formation -- as well as many binary systems. Both of these stages have implications for star and planet formation. We present results from both 1-D and 2-D disk modeling, where we try to understand disk substructure that might indicate on-going planet formation, in particular, transition disk cavities, disk gaps, and asymmetries in the dust emission.

  18. Rho kinase regulates fragmentation and phagocytosis of apoptotic cells

    SciTech Connect

    Orlando, Kelly A.; Stone, Nicole L.; Pittman, Randall N. . E-mail: pittman@pharm.med.upenn.edu

    2006-01-01

    During the execution phase of apoptosis, a cell undergoes cytoplasmic and nuclear changes that prepare it for death and phagocytosis. The end-point of the execution phase is condensation into a single apoptotic body or fragmentation into multiple apoptotic bodies. Fragmentation is thought to facilitate phagocytosis; however, mechanisms regulating fragmentation are unknown. An isoform of Rho kinase, ROCK-I, drives membrane blebbing through its activation of actin-myosin contraction; this raises the possibility that ROCK-I may regulate other execution phase events, such as cellular fragmentation. Here, we show that COS-7 cells fragment into a number of small apoptotic bodies during apoptosis; treating with ROCK inhibitors (Y-27632 or H-1152) prevents fragmentation. Latrunculin B and blebbistatin, drugs that interfere with actin-myosin contraction, also inhibit fragmentation. During apoptosis, ROCK-I is cleaved and activated by caspases, while ROCK-II is not activated, but rather translocates to a cytoskeletal fraction. siRNA knock-down of ROCK-I but not ROCK-II inhibits fragmentation of dying cells, consistent with ROCK-I being required for apoptotic fragmentation. Finally, cells dying in the presence of the ROCK inhibitor Y-27632 are not efficiently phagocytized. These data show that ROCK plays an essential role in fragmentation and phagocytosis of apoptotic cells.

  19. Exclusive rho^0 electroproduction on the proton at CLAS

    SciTech Connect

    Morrow, Steven; Guidal, Michel; Garcon, Michel; Laget, Jean; Smith, Elton; Adams, Gary; Adhikari, Krishna; Aghasyan, Mher; Amaryan, Moscov; Amaryan, Moskov; Anghinolfi, Marco; Asryan, G.; Audit, Gerard; Avagyan, Harutyun; Bagdasaryan, H.; Baillie, Nathan; Ball, J.P.; Ball, Jacques; Baltzell, Nathan; Barrow, Steve; Battaglieri, Marco; Bedlinskiy, Ivan; Bektasoglu, Mehmet; Bellis, Matthew; Benmouna, Nawal; Berman, Barry; Biselli, Angela; Blaszczyk, Lukasz; Bonner, Billy; Bookwalter, Craig; Bouchigny, Sylvain; Boyarinov, Sergey; Bradford, Robert; Branford, Derek; Briscoe, William; Brooks, William; Bultmann, S.; Bueltmann, Stephen; Burkert, Volker; Butuceanu, Cornel; Calarco, John; Careccia, Sharon; Carman, Daniel; Carnahan, Bryan; Casey, Liam; Cazes, Antoine; Chen, Shifeng; Cheng, Lu; Cole, Philip; Collins, Patrick; Coltharp, Philip; Cords, Dieter; Corvisiero, Pietro; Crabb, Donald; Crannell, Hall; Crede, Volker; Cummings, John; Dale, Daniel; Dashyan, Natalya; De Masi, Rita; De Vita, Raffaella; De Sanctis, Enzo; Degtiarenko, Pavel; Denizli, Haluk; Dennis, Lawrence; Deur, Alexandre; Dhamija, Seema; Dharmawardane, Kahanawita; Dhuga, Kalvir; Dickson, Richard; Didelez, Jean-Pierre; Djalali, Chaden; Dodge, Gail; Doughty, David; Dugger, Michael; Dytman, Steven; Dzyubak, Oleksandr; Egiyan, Hovanes; Egiyan, Kim; Elfassi, Lamiaa; Elouadrhiri, Latifa; Eugenio, Paul; Fatemi, Renee; Fedotov, Gleb; Fersch, Robert; Feuerbach, Robert; Forest, Tony; Fradi, Ahmed; Gavalian, Gagik; Gevorgyan, Nerses; Gilfoyle, Gerard; Giovanetti, Kevin; Girod, Francois-Xavier; Goetz, John; Gohn, Wesley; Gordon, Christopher; Gothe, Ralf; Graham, Lewis; Griffioen, Keith; Guillo, Matthieu; Guler, Nevzat; Guo, Lei; Gyurjyan, Vardan; Hadjidakis, Cynthia; Hafidi, Kawtar; Hakobyan, Hayk; Hanretty, Charles; Hardie, John; Hassall, Neil; Heddle, David; Hersman, F.; Hicks, Kenneth; Hleiqawi, Ishaq; Holtrop, Maurik; Hourany, E.; Hyde, Charles; Ilieva, Yordanka; Ireland, David; Ishkhanov, Boris; Isupov, Evgeny; Ito, Mark; Jenkins, David; Jo, Hyon-Suk; Johnstone, John; Joo, Kyungseon; Juengst, Henry; Kalantarians, Narbe; Keller, Dustin; Kellie, James; Khandaker, Mahbubul; Khetarpal, Puneet; Kim, Wooyoung; Klein, Andreas; Klein, Franz; Klimenko, Alexei; Kossov, Mikhail; Kramer, Laird; Kubarovsky, Valery; Kuhn, Joachim; Kuhn, Sebastian; Kuleshov, Sergey; Kuznetsov, Viacheslav; Lachniet, Jeff; Langheinrich, Jorn; Lawrence, Dave; Li, Ji; Livingston, Kenneth; Lu, Haiyun; MacCormick, Marion; Marchand, Claude; Markov, Nikolai; Mattione, Paul; McAleer, Simeon; McCracken, Michael; McKinnon, Bryan; McNabb, John; Mecking, Bernhard; Mehrabyan, Surik; Melone, Joseph; Mestayer, Mac; Meyer, Curtis; Mibe, Tsutomu; Mikhaylov, Konstantin; Minehart, Ralph; Mirazita, Marco; Miskimen, Rory; Mokeev, Viktor; Morand, Ludyvine; Moreno, Brahim; Moriya, Kei; Moteabbed, Maryam; Mueller, James; Munevar Espitia, Edwin; Mutchler, Gordon; Nadel-Turonski, Pawel; Nasseripour, Rakhsha; Niccolai, Silvia; Niculescu, Gabriel; Niculescu, Maria-Ioana; Niczyporuk, Bogdan; Niroula, Megh; Niyazov, Rustam; Nozar, Mina; O'Rielly, Grant; Osipenko, Mikhail; Ostrovidov, Alexander; Park, Kijun; Park, Sungkyun; Pasyuk, Evgueni; Paterson, Craig; Pereira, S.Anefalos; Philips, Sasha; Pierce, Joshua; Pivnyuk, Nikolay; Pocanic, Dinko; Pogorelko, Oleg; Polli, Ermanno; Popa, Iulian; Pozdnyakov, Sergey; Preedom, Barry; Price, John; Procureur, Sebastien; Prok, Yelena; Protopopescu, Dan; Qin, Liming; Raue, Brian; Riccardi, Gregory; Ricco, Giovanni; Ripani, Marco; Ritchie, Barry; Rosner, Guenther; Rossi, Patrizia; Rubin, Philip; Sabatie, Franck; Saini, Mukesh; Salamanca, Julian; Salgado, Carlos; Santoro, Joseph; Sapunenko, Vladimir; Schott, Diane; Schumacher, Reinhard; Serov, Vladimir; Sharabian, Youri; Sharov, Dmitri; Shvedunov, Nikolay; Skabelin, Alexander; Smith, Lee; Sober, Daniel; Sokhan, Daria; Stavinskiy, Aleksey; Stepanyan, Samuel; Stepanyan, Stepan; Stokes, Burnham; Stoler, Paul; Strakovski, Igor; Strauch, Steffen; Taiuti, Mauro

    2009-01-01

    The $e p\\to e^\\prime p \\rho^0$ reaction has been measured, using the 5.754 GeV electron beam of Jefferson Lab and the CLAS detector. This represents the largest ever set of data for this reaction in the valence region. Integrated and differential cross sections are presented. The $W$, $Q^2$ and $t$ dependences of the cross section are compared to theoretical calculations based on $t$-channel meson-exchange Regge theory on the one hand and on quark handbag diagrams related to Generalized Parton Distributions (GPDs) on the other hand. The Regge approach can describe at the $\\approx$ 30% level most of the features of the present data while the two GPD calculations that are presented in this article which succesfully reproduce the high energy data strongly underestimate the present data. The question is then raised whether this discrepancy originates from an incomplete or inexact way of modelling the GPDs or the associated hard scattering amplitude or whether the GPD formalism is simply in

  20. TNF-induced endothelial barrier disruption: beyond actin and Rho.

    PubMed

    Marcos-Ramiro, B; García-Weber, D; Millán, J

    2014-12-01

    The decrease of endothelial barrier function is central to the long-term inflammatory response. A pathological alteration of the ability of endothelial cells to modulate the passage of cells and solutes across the vessel underlies the development of inflammatory diseases such as atherosclerosis and multiple sclerosis. The inflammatory cytokine tumour necrosis factor (TNF) mediates changes in the barrier properties of the endothelium. TNF activates different Rho GTPases, increases filamentous actin and remodels endothelial cell morphology. However, inhibition of actin-mediated remodelling is insufficient to prevent endothelial barrier disruption in response to TNF, suggesting that additional molecular mechanisms are involved. Here we discuss, first, the pivotal role of Rac-mediated generation of reactive oxygen species (ROS) to regulate the integrity of endothelial cell-cell junctions and, second, the ability of endothelial adhesion receptors such as ICAM-1, VCAM-1 and PECAM-1, involved in leukocyte transendothelial migration, to control endothelial permeability to small molecules, often through ROS generation. These adhesion receptors regulate endothelial barrier function in ways both dependent on and independent of their engagement by immune cells, and orchestrate the crosstalk between leukocyte transendothelial migration and endothelial permeability during inflammation.

  1. JAK tyrosine kinases promote hierarchical activation of Rho and Rap modules of integrin activation.

    PubMed

    Montresor, Alessio; Bolomini-Vittori, Matteo; Toffali, Lara; Rossi, Barbara; Constantin, Gabriela; Laudanna, Carlo

    2013-12-23

    Lymphocyte recruitment is regulated by signaling modules based on the activity of Rho and Rap small guanosine triphosphatases that control integrin activation by chemokines. We show that Janus kinase (JAK) protein tyrosine kinases control chemokine-induced LFA-1- and VLA-4-mediated adhesion as well as human T lymphocyte homing to secondary lymphoid organs. JAK2 and JAK3 isoforms, but not JAK1, mediate CXCL12-induced LFA-1 triggering to a high affinity state. Signal transduction analysis showed that chemokine-induced activation of the Rho module of LFA-1 affinity triggering is dependent on JAK activity, with VAV1 mediating Rho activation by JAKs in a Gαi-independent manner. Furthermore, activation of Rap1A by chemokines is also dependent on JAK2 and JAK3 activity. Importantly, activation of Rap1A by JAKs is mediated by RhoA and PLD1, thus establishing Rap1A as a downstream effector of the Rho module. Thus, JAK tyrosine kinases control integrin activation and dependent lymphocyte trafficking by bridging chemokine receptors to the concurrent and hierarchical activation of the Rho and Rap modules of integrin activation.

  2. Toward understanding RhoGTPase specificity: structure, function and local activation

    PubMed Central

    Schaefer, Antje; Reinhard, Nathalie R; Hordijk, Peter L

    2014-01-01

    Cell adhesion and migration are regulated through the concerted action of cytoskeletal dynamics and adhesion proteins, the activity of which is governed by RhoGTPases. Specific RhoGTPase signaling requires spatio-temporal activation and coordination of subsequent protein-protein and protein-lipid interactions. The nature, location and duration of these interactions are dependent on polarized extracellular triggers, such as cell-cell contact, and intracellular modifying events, such as phosphorylation. RhoA, RhoB, and RhoC are highly homologous GTPases that, however, succeed in generating specific intracellular responses. Here, we discuss the key features that contribute to this specificity. These not only include the well-studied switch regions, the conformation of which is nucleotide-dependent, but also additional regions and seemingly small differences in primary sequence that also contribute to specific interactions. These differences translate into differential surface charge distribution, local exposure of amino acid side-chains and isoform-specific post-translational modifications. The available evidence supports the notion that multiple regions in RhoA/B/C cooperate to provide specificity in binding to regulators and effectors. These specific interactions are highly regulated in time and space. We therefore subsequently discuss current approaches means to visualize and analyze localized GTPase activation using biosensors that allow imaging of isoform-specific, localized regulation. PMID:25483298

  3. Rho protein GTPases and their interactions with NFκB: crossroads of inflammation and matrix biology

    PubMed Central

    Tong, Louis; Tergaonkar, Vinay

    2014-01-01

    The RhoGTPases, with RhoA, Cdc42 and Rac being major members, are a group of key ubiquitous proteins present in all eukaryotic organisms that subserve such important functions as cell migration, adhesion and differentiation. The NFκB (nuclear factor κB) is a family of constitutive and inducible transcription factors that through their diverse target genes, play a major role in processes such as cytokine expression, stress regulation, cell division and transformation. Research over the past decade has uncovered new molecular links between the RhoGTPases and the NFκB pathway, with the RhoGTPases playing a positive or negative regulatory role on NFκB activation depending on the context. The RhoA–NFκB interaction has been shown to be important in cytokine-activated NFκB processes, such as those induced by TNFα (tumour necrosis factor α). On the other hand, Rac is important for activating the NFκB response downstream of integrin activation, such as after phagocytosis. Specific residues of Rac1 are important for triggering NFκB activation, and mutations do obliterate this response. Other upstream triggers of the RhoGTPase–NFκB interactions include the suppressive p120 catenin, with implications for skin inflammation. The networks described here are not only important areas for further research, but are also significant for discovery of targets for translational medicine. PMID:24877606

  4. RhoA Regulates Peroxisome Association to Microtubules and the Actin Cytoskeleton

    PubMed Central

    Lay, Dorothee; Wiese, Sebastian; Meyer, Helmut E.; Warscheid, Bettina; Saffrich, Rainer; Peränen, Johan; Gorgas, Karin; Just, Wilhelm W.

    2010-01-01

    The current view of peroxisome inheritance provides for the formation of new peroxisomes by both budding from the endoplasmic reticulum and autonomous division. Here we investigate peroxisome-cytoskeleton interactions and show by proteomics, biochemical and immunofluorescence analyses that actin, non-muscle myosin IIA (NMM IIA), RhoA, Rho kinase II (ROCKII) and Rab8 associate with peroxisomes. Our data provide evidence that (i) RhoA in its inactive state, maintained for example by C. botulinum toxin exoenzyme C3, dissociates from peroxisomes enabling microtubule-based peroxisomal movements and (ii) dominant-active RhoA targets to peroxisomes, uncouples the organelles from microtubules and favors Rho kinase recruitment to peroxisomes. We suggest that ROCKII activates NMM IIA mediating local peroxisomal constrictions. Although our understanding of peroxisome-cytoskeleton interactions is still incomplete, a picture is emerging demonstrating alternate RhoA-dependent association of peroxisomes to the microtubular and actin cytoskeleton. Whereas association of peroxisomes to microtubules clearly serves bidirectional, long-range saltatory movements, peroxisome-acto-myosin interactions may support biogenetic functions balancing peroxisome size, shape, number, and clustering. PMID:21079737

  5. Dihydrotestosterone modulates endothelial progenitor cell function via RhoA/ROCK pathway

    PubMed Central

    Zhang, Hao; Shi, Liang; Ren, Guo-Qing; Sun, Wen-Wen; Wang, Yi-Bin; Chen, Yi-Kun; Yin, Jiang-Ning; Wan, Bing

    2016-01-01

    Background: Previous findings indicate that testosterone level is negatively correlated with the incidence and mortality of cardiovascular diseases in men. Endothelial progenitor cells (EPCs) play a critical role in endothelial healing and vascular integrity. This study aimed to examine the effects of dihydrotestosterone (DHT), an active metabolite of testosterone, on human EPC function and investigate the underlying mechanism. Methods: EPCs were isolated from peripheral blood of healthy adult males and incubated with a series of concentrations (1, 10, and 100 nmol/L in dimethyl sulfoxide) of DHT for 24 h or with 10 nmol/L DHT for different periods (6, 12, 24, 36, and 48 h). EPC proliferation, migration, and adhesion were determined by MTT assay, modified Boyden chamber assay, and cell counting, respectively. Furthermore, vascular endothelial growth factor (VEGF) production was examined by ELISA, RhoA activity was determined through pull-down assay. The protein level of RhoA was quantified by Western blot analysis. Results: DHT significantly increased the proliferative, migratory, and adhesive abilities of EPCs in a dose- and time-dependent manner and upregulated the levels of VEGF and activated RhoA. However, RhoA inhibitor C3 exoenzyme or ROCK inhibitor Y-27632 significantly inhibited DHT-induced proliferation, migration, and adhesion, as well as VEGF production. Moreover, C3 exoenzyme inhibited the activation of RhoA stimulated by DHT. Conclusions: DHT promotes EPC proliferation, migration, and adhesion activities via RhoA/ROCK pathway. PMID:27830013

  6. Ameloblasts require active RhoA to generate normal dental enamel

    PubMed Central

    Li, Yong; Xue, Hui; Everett, Eric T.; Ryan, Kathleen; Peng, Li; Porecha, Rakhee; Yan, Yan; Lucchese, Anna M.; Kuehl, Melissa A.; Pugach, Megan K.; Bouchard, Jessica; Gibson, Carolyn W.

    2013-01-01

    RhoA plays a fundamental role in regulation of the actin cytoskeleton, intercellular attachment and cell proliferation. During amelogenesis, ameloblasts which produce the enamel proteins undergo dramatic cytoskeletal changes and RhoA protein level is upregulated. Transgenic mice were generated that express a dominant-negative RhoA transgene in ameloblasts using amelogenin gene regulatory sequences. Transgenic and WT molar tooth germs were incubated with NaF or NaCl in organ culture. F-actin stained with phalloidin was elevated significantly in WT ameloblasts treated with NaF compared to WT ameloblasts treated with NaCl or compared to transgenic ameloblasts treated with NaF, thereby confirming a block in the RhoA/ROCK pathway in the transgenic mice. Little difference in quantitative fluorescence (estimation of fluorosis) was observed between WT and transgenic incisors from mice provided NaF in their drinking water. We subsequently found reduced transgene expression in incisors compared to molars. Transgenic molar teeth had reduced amelogenin, E-cadherin and Ki67 compared to WT. Hypoplastic enamel in transgenic mice correlates with reduced expression of the enamel protein amelogenin, and E-cadherin and cell proliferation are regulated by RhoA in other tissues. Together these findings reveal deficits in molar ameloblast function when RhoA activity is inhibited. PMID:23841780

  7. Elevated Intraocular Pressure Induces Rho GTPase Mediated Contractile Signaling in the Trabecular Meshwork

    PubMed Central

    Pattabiraman, Padmanabhan P; Inoue, Toshihiro; Rao, P. Vasantha

    2015-01-01

    Rho GTPase regulated contractile signaling in the trabecular meshwork (TM) has been shown to modulate aqueous humor (AH) outflow and intraocular pressure (IOP). To explore whether elevated IOP, a major risk factor for primary open angle glaucoma (POAG) influences Rho GTPase signaling in the TM, we recorded AH outflow in enucleated contralateral porcine eyes perfused for 4–5 hours at either 15 mm or 50 mm Hg pressure. After perfusion, TM tissue extracted from perfused eyes was evaluated for the activation status of Rho GTPase, myosin light chain (MLC), myosin phosphatase target substrate 1 (MYPT1), myristoylated alanine-rich C-kinase substrate (MARCKS) and paxillin. Eyes perfused at 50 mm Hg exhibited a significant decrease in AH outflow facility compared with those perfused at 15 mm Hg. Additionally, TM tissue from eyes perfused at 50 mm Hg revealed significantly increased levels of activated RhoA and phosphorylated MLC, MYPT1, MARCKS and paxillin compared to TM tissue derived from eyes perfused at 15 mm Hg. Taken together, these observations indicate that elevated IOP-induced activation of Rho GTPase-dependent contractile signaling in the TM is associated with increased resistance to AH outflow through the trabecular pathway, and demonstrate the sensitivity of Rho GTPase signaling to mechanical force in the AH outflow pathway. PMID:25956210

  8. Concentric zones of active RhoA and Cdc42 around single cell wounds

    PubMed Central

    Benink, Hélène A.; Bement, William M.

    2005-01-01

    Rho GTPases control many cytoskeleton-dependent processes, but how they regulate spatially distinct features of cytoskeletal function within a single cell is poorly understood. Here, we studied active RhoA and Cdc42 in wounded Xenopus oocytes, which assemble and close a dynamic ring of actin filaments (F-actin) and myosin-2 around wound sites. RhoA and Cdc42 are rapidly activated around wound sites in a calcium-dependent manner and segregate into distinct, concentric zones around the wound, with active Cdc42 in the approximate middle of the F-actin array and active RhoA on the interior of the array. These zones form before F-actin accumulation, and then move in concert with the closing array. Microtubules and F-actin are required for normal zone organization and dynamics, as is crosstalk between RhoA and Cdc42. Each of the zones makes distinct contributions to the organization and function of the actomyosin wound array. We propose that similar rho activity zones control related processes such as cytokinesis. PMID:15684032

  9. Regulation of Rho-GEF Rgf3 by the arrestin Art1 in fission yeast cytokinesis

    PubMed Central

    Davidson, Reshma; Laporte, Damien; Wu, Jian-Qiu

    2015-01-01

    Rho GTPases, activated by guanine nucleotide exchange factors (GEFs), are essential regulators of polarized cell growth, cytokinesis, and many other cellular processes. However, the regulation of Rho-GEFs themselves is not well understood. Rgf3 is an essential GEF for Rho1 GTPase in fission yeast. We show that Rgf3 protein levels and localization are regulated by arrestin-related protein Art1. art1∆ cells lyse during cell separation with a thinner and defective septum. As does Rgf3, Art1 concentrates to the contractile ring starting at early anaphase and spreads to the septum during and after ring constriction. Art1 localization depends on its C-terminus, and Art1 is important for maintaining Rgf3 protein levels. Biochemical experiments reveal that the Rgf3 C-terminus binds to Art1. Using an Rgf3 conditional mutant and mislocalization experiments, we found that Art1 and Rgf3 are interdependent for localization to the division site. As expected, active Rho1 levels at the division site are reduced in art1∆ and rgf3 mutant cells. Taken together, these data reveal that the arrestin family protein Art1 regulates the protein levels and localization of the Rho-GEF Rgf3, which in turn modulates active Rho1 levels during fission yeast cytokinesis. PMID:25473118

  10. Computer vision profiling of neurite outgrowth dynamics reveals spatiotemporal modularity of Rho GTPase signaling

    PubMed Central

    Fusco, Ludovico; Lefort, Riwal; Smith, Kevin; Benmansour, Fethallah; Gonzalez, German; Barillari, Caterina; Rinn, Bernd; Fleuret, Francois; Fua, Pascal

    2016-01-01

    Rho guanosine triphosphatases (GTPases) control the cytoskeletal dynamics that power neurite outgrowth. This process consists of dynamic neurite initiation, elongation, retraction, and branching cycles that are likely to be regulated by specific spatiotemporal signaling networks, which cannot be resolved with static, steady-state assays. We present NeuriteTracker, a computer-vision approach to automatically segment and track neuronal morphodynamics in time-lapse datasets. Feature extraction then quantifies dynamic neurite outgrowth phenotypes. We identify a set of stereotypic neurite outgrowth morphodynamic behaviors in a cultured neuronal cell system. Systematic RNA interference perturbation of a Rho GTPase interactome consisting of 219 proteins reveals a limited set of morphodynamic phenotypes. As proof of concept, we show that loss of function of two distinct RhoA-specific GTPase-activating proteins (GAPs) leads to opposite neurite outgrowth phenotypes. Imaging of RhoA activation dynamics indicates that both GAPs regulate different spatiotemporal Rho GTPase pools, with distinct functions. Our results provide a starting point to dissect spatiotemporal Rho GTPase signaling networks that regulate neurite outgrowth. PMID:26728857

  11. Nicotine facilitates VSMC dysfunction through a miR-200b/RhoGDIA/cytoskeleton module

    PubMed Central

    Liang, Dongli; Wang, Zhaoxia; Yan, Zhiqiang; Hou, Shangwei; Xu, Wangjie; Wang, Lianyun; Shang, Meisheng; Qiao, Zhongdong

    2017-01-01

    Nicotine can induce the abnormal migration and proliferation of vascular smooth muscle cells (VSMCs). We have previously shown that cytoskeletal proteins and RhoGDIA, a negative regulator of the Rho GTPase pathway, are involved in the nicotine-induced dysfunction of VSMCs. Here, we found that nicotine can activate the Rho GTPase pathway and induce the synthesis of the cytoskeletal proteins in VSMCs through the activation of intracellular downstream signaling pathways, including targets such as MYPT1, PAK1 and PI3K/AKT. Upon nicotine treatment, the mRNA level of RhoGDIA is increased but protein level is decreased both in vitro and in vivo, which suggested a mechanism of post-translational regulation. By the dual luciferase reporter assay, we identified the microRNA-200b (miR-200b) as a modulator of the behavioural changes of VSMCs in response to nicotine through targeting RhoGDIA directly. Introducing miR-200b inhibitors into cultured VSMCs significantly attenuated cell proliferation and migration. Additionally, we found that hypomethylation in the CpG island shore region of miR-200b was responsible for the nicotine-induced miR-200b up-regulation in VSMCs. The study demonstrates that nicotine facilitates VSMC dysfunction through a miR-200b/RhoGDIA/cytoskeleton module through the hypomethylation of miR-200b promoter and suggests that epigenetic modifications may play an important role in the pathological progression. PMID:28252009

  12. EspM2 is a RhoA guanine nucleotide exchange factor

    PubMed Central

    Arbeloa, Ana; Garnett, James; Lillington, James; Bulgin, Richard R; Berger, Cedric N; Lea, Susan M; Matthews, Steve; Frankel, Gad

    2010-01-01

    We investigated how the type III secretion system WxxxE effectors EspM2 of enterohaemorrhagic Escherichia coli, which triggers stress fibre formation, and SifA of Salmonella enterica serovar Typhimurium, which is involved in intracellular survival, modulate Rho GTPases. We identified a direct interaction between EspM2 or SifA and nucleotide-free RhoA. Nuclear Magnetic Resonance Spectroscopy revealed that EspM2 has a similar fold to SifA and the guanine nucleotide exchange factor (GEF) effector SopE. EspM2 induced nucleotide exchange in RhoA but not in Rac1 or H-Ras, while SifA induced nucleotide exchange in none of them. Mutating W70 of the WxxxE motif or L118 and I127 residues, which surround the catalytic loop, affected the stability of EspM2. Substitution of Q124, located within the catalytic loop of EspM2, with alanine, greatly attenuated the RhoA GEF activity in vitro and the ability of EspM2 to induce stress fibres upon ectopic expression. These results suggest that binding of SifA to RhoA does not trigger nucleotide exchange while EspM2 is a unique Rho GTPase GEF. PMID:20039879

  13. The RhoA-ROCK pathway in the regulation of T and B cell responses

    PubMed Central

    Ricker, Edd; Chowdhury, Luvana; Yi, Woelsung; Pernis, Alessandra B.

    2016-01-01

    Effective immune responses require the precise regulation of dynamic interactions between hematopoietic and non-hematopoietic cells. The Rho subfamily of GTPases, which includes RhoA, is rapidly activated downstream of a diverse array of biochemical and biomechanical signals, and is emerging as an important mediator of this cross-talk. Key downstream effectors of RhoA are the Rho kinases, or ROCKs. The ROCKs are two serine-threonine kinases that can act as global coordinators of a tissue’s response to stress and injury because of their ability to regulate a wide range of biological processes. Although the RhoA-ROCK pathway has been extensively investigated in the non-hematopoietic compartment, its role in the immune system is just now becoming appreciated. In this commentary, we provide a brief overview of recent findings that highlight the contribution of this pathway to lymphocyte development and activation, and the impact that dysregulation in the activation of RhoA and/or the ROCKs may exert on a growing list of autoimmune and lymphoproliferative disorders. PMID:27785353

  14. Fine regulation of RhoA and Rock is required for skeletal muscle differentiation.

    PubMed

    Castellani, Loriana; Salvati, Erica; Alemà, Stefano; Falcone, Germana

    2006-06-02

    The RhoA GTPase controls a variety of cell functions such as cell motility, cell growth, and gene expression. Previous studies suggested that RhoA mediates signaling inputs that promote skeletal myogenic differentiation. We show here that levels and activity of RhoA protein are down-regulated in both primary avian myoblasts and mouse satellite cells undergoing differentiation, suggesting that a fine regulation of this GTPase is required. In addition, ectopic expression of activated RhoA in primary quail myocytes, but not in mouse myocytes, inhibits accumulation of muscle-specific proteins and cell fusion. By disrupting RhoA signaling with specific inhibitors, we have shown that this GTPase, although required for cell identity in proliferating myoblasts, is not essential for commitment to terminal differentiation and muscle gene expression. Ectopic expression of an activated form of its downstream effector, Rock, impairs differentiation of both avian and mouse myoblasts. Conversely, Rock inhibition with specific inhibitors and small interfering RNA-mediated gene silencing leads to accelerated progression in the lineage and enhanced cell fusion, underscoring a negative regulatory function of Rock in myogenesis. Finally, we have reported that Rock acts independently from RhoA in preventing myoblast exit from the cell cycle and commitment to differentiation and may receive signaling inputs from Raf-1 kinase.

  15. RhoA mediates defective stem cell function and heterotopic ossification in dystrophic muscle of mice.

    PubMed

    Mu, Xiaodong; Usas, Arvydas; Tang, Ying; Lu, Aiping; Wang, Bing; Weiss, Kurt; Huard, Johnny

    2013-09-01

    Heterotopic ossification (HO) and fatty infiltration (FI) often occur in diseased skeletal muscle and have been previously described in various animal models of Duchenne muscular dystrophy (DMD); however, the pathological mechanisms remain largely unknown. Dystrophin-deficient mdx mice and dystrophin/utrophin double-knockout (dKO) mice are mouse models of DMD; however, mdx mice display a strong muscle regeneration capacity, while dKO mice exhibit a much more severe phenotype, which is similar to patients with DMD. Our results revealed that more extensive HO, but not FI, occurred in the skeletal muscle of dKO mice versus mdx mice, and RhoA activation specifically occurred at the sites of HO. Moreover, the gene expression of RhoA, BMPs, and several inflammatory factors were significantly up-regulated in muscle stem cells isolated from dKO mice; while inactivation of RhoA in the cells with RhoA/ROCK inhibitor Y-27632 led to reduced osteogenic potential and improved myogenic potential. Finally, inactivation of RhoA signaling in the dKO mice with Y-27632 improved muscle regeneration and reduced the expression of BMPs, inflammation, HO, and intramyocellular lipid accumulation in both skeletal and cardiac muscle. Our results revealed that RhoA represents a major molecular switch in the regulation of HO and muscle regeneration in dystrophic skeletal muscle of mice.

  16. Genetic analysis demonstrates a direct link between rho signaling and nonmuscle myosin function during Drosophila morphogenesis.

    PubMed Central

    Halsell, S R; Chu, B I; Kiehart, D P

    2000-01-01

    A dynamic actomyosin cytoskeleton drives many morphogenetic events. Conventional nonmuscle myosin-II (myosin) is a key chemomechanical motor that drives contraction of the actin cytoskeleton. We have explored the regulation of myosin activity by performing genetic screens to identify gene products that collaborate with myosin during Drosophila morphogenesis. Specifically, we screened for second-site noncomplementors of a mutation in the zipper gene that encodes the nonmuscle myosin-II heavy chain. We determined that a single missense mutation in the zipper(Ebr) allele gives rise to its sensitivity to second-site noncomplementation. We then identify the Rho signal transduction pathway as necessary for proper myosin function. First we show that a lethal P-element insertion interacts genetically with zipper. Subsequently we show that this second-site noncomplementing mutation disrupts the RhoGEF2 locus. Next, we show that two EMS-induced mutations, previously shown to interact genetically with zipper(Ebr), disrupt the RhoA locus. Further, we have identified their molecular lesions and determined that disruption of the carboxyl-terminal CaaX box gives rise to their mutant phenotype. Finally, we show that RhoA mutations themselves can be utilized in genetic screens. Biochemical and cell culture analyses suggest that Rho signal transduction regulates the activity of myosin. Our studies provide direct genetic proof of the biological relevance of regulation of myosin by Rho signal transduction in an intact metazoan. PMID:10880486

  17. Extracting Diffusive States of Rho GTPase in Live Cells: Towards In Vivo Biochemistry.

    PubMed

    Koo, Peter K; Weitzman, Matthew; Sabanaygam, Chandran R; van Golen, Kenneth L; Mochrie, Simon G J

    2015-10-01

    Resolving distinct biochemical interaction states when analyzing the trajectories of diffusing proteins in live cells on an individual basis remains challenging because of the limited statistics provided by the relatively short trajectories available experimentally. Here, we introduce a novel, machine-learning based classification methodology, which we call perturbation expectation-maximization (pEM), that simultaneously analyzes a population of protein trajectories to uncover the system of diffusive behaviors which collectively result from distinct biochemical interactions. We validate the performance of pEM in silico and demonstrate that pEM is capable of uncovering the proper number of underlying diffusive states with an accurate characterization of their diffusion properties. We then apply pEM to experimental protein trajectories of Rho GTPases, an integral regulator of cytoskeletal dynamics and cellular homeostasis, in vivo via single particle tracking photo-activated localization microscopy. Remarkably, pEM uncovers 6 distinct diffusive states conserved across various Rho GTPase family members. The variability across family members in the propensities for each diffusive state reveals non-redundant roles in the activation states of RhoA and RhoC. In a resting cell, our results support a model where RhoA is constantly cycling between activation states, with an imbalance of rates favoring an inactive state. RhoC, on the other hand, remains predominantly inactive.

  18. Rho signaling regulates pannexin 1-mediated ATP release from airway epithelia.

    PubMed

    Seminario-Vidal, Lucia; Okada, Seiko F; Sesma, Juliana I; Kreda, Silvia M; van Heusden, Catharina A; Zhu, Yunxiang; Jones, Lisa C; O'Neal, Wanda K; Penuela, Silvia; Laird, Dale W; Boucher, Richard C; Lazarowski, Eduardo R

    2011-07-29

    ATP released from airway epithelial cells promotes purinergic receptor-regulated mucociliary clearance activities necessary for innate lung defense. Cell swelling-induced membrane stretch/strain is a common stimulus that promotes airway epithelial ATP release, but the mechanisms transducing cell swelling into ATP release are incompletely understood. Using knockdown and knockout approaches, we tested the hypothesis that pannexin 1 mediates ATP release from hypotonically swollen airway epithelia and investigated mechanisms regulating this activity. Well differentiated primary cultures of human bronchial epithelial cells subjected to hypotonic challenge exhibited enhanced ATP release, which was paralleled by the uptake of the pannexin probe propidium iodide. Both responses were reduced by pannexin 1 inhibitors and by knocking down pannexin 1. Importantly, hypotonicity-evoked ATP release from freshly excised tracheas and dye uptake in primary tracheal epithelial cells were impaired in pannexin 1 knockout mice. Hypotonicity-promoted ATP release and dye uptake in primary well differentiated human bronchial epithelial cells was accompanied by RhoA activation and myosin light chain phosphorylation and was reduced by the RhoA dominant negative mutant RhoA(T19N) and Rho and myosin light chain kinase inhibitors. ATP release and Rho activation were reduced by highly selective inhibitors of transient receptor potential vanilloid 4 (TRPV4). Lastly, knocking down TRPV4 impaired hypotonicity-evoked airway epithelial ATP release. Our data suggest that TRPV4 and Rho transduce cell membrane stretch/strain into pannexin 1-mediated ATP release in airway epithelia.

  19. Rho Signaling Regulates Pannexin 1-mediated ATP Release from Airway Epithelia*

    PubMed Central

    Seminario-Vidal, Lucia; Okada, Seiko F.; Sesma, Juliana I.; Kreda, Silvia M.; van Heusden, Catharina A.; Zhu, Yunxiang; Jones, Lisa C.; O'Neal, Wanda K.; Penuela, Silvia; Laird, Dale W.; Boucher, Richard C.; Lazarowski, Eduardo R.

    2011-01-01

    ATP released from airway epithelial cells promotes purinergic receptor-regulated mucociliary clearance activities necessary for innate lung defense. Cell swelling-induced membrane stretch/strain is a common stimulus that promotes airway epithelial ATP release, but the mechanisms transducing cell swelling into ATP release are incompletely understood. Using knockdown and knockout approaches, we tested the hypothesis that pannexin 1 mediates ATP release from hypotonically swollen airway epithelia and investigated mechanisms regulating this activity. Well differentiated primary cultures of human bronchial epithelial cells subjected to hypotonic challenge exhibited enhanced ATP release, which was paralleled by the uptake of the pannexin probe propidium iodide. Both responses were reduced by pannexin 1 inhibitors and by knocking down pannexin 1. Importantly, hypotonicity-evoked ATP release from freshly excised tracheas and dye uptake in primary tracheal epithelial cells were impaired in pannexin 1 knockout mice. Hypotonicity-promoted ATP release and dye uptake in primary well differentiated human bronchial epithelial cells was accompanied by RhoA activation and myosin light chain phosphorylation and was reduced by the RhoA dominant negative mutant RhoA(T19N) and Rho and myosin light chain kinase inhibitors. ATP release and Rho activation were reduced by highly selective inhibitors of transient receptor potential vanilloid 4 (TRPV4). Lastly, knocking down TRPV4 impaired hypotonicity-evoked airway epithelial ATP release. Our data suggest that TRPV4 and Rho transduce cell membrane stretch/strain into pannexin 1-mediated ATP release in airway epithelia. PMID:21606493

  20. RhoGEF and positioning of rappaport-like furrows in the early Drosophila embryo.

    PubMed

    Crest, Justin; Concha-Moore, Kirsten; Sullivan, William

    2012-11-06

    Early Drosophila embryogenesis is characterized by shifting from astral microtubule-based to central spindle-based positioning of cleavage furrows. Before cellularization, astral microtubules determine metaphase furrow position by producing Rappaport-like furrows, which encompass rather than bisect the spindle. Their positioning is explained by our finding that the conserved central spindle components centralspindlin (mKLP1 and RacGAP50C), Polo, and Fascetto (Prc1) localize to the astral microtubule overlap region. These components and the chromosomal passenger complex localize to the central spindle, though no furrow forms there. We identify the maternally supplied RhoGEF2 as a key factor in metaphase furrow positioning. Unlike the zygotic, central spindle-localized RhoGEF (Pebble), RhoGEF2 localizes to metaphase furrows, a function distinct from RhoGEF/Pebble and likely due to the absence of a RacGAP50C binding domain. Accordingly, we find that ectopic activation of Rho GTPase generates furrows perpendicular to the central spindle during syncytial divisions. Whereas metaphase furrow formation is myosin independent, these ectopic furrows, like conventional furrows, require myosin as well as microtubules. These studies demonstrate that early Drosophila embryogenesis is primed to form furrows at either overlapping astral microtubules or the central spindle. We propose that the shift to the latter is driven by a corresponding shift from RhoGEF2 to Pebble in controlling furrow formation.

  1. Formin-like2 regulates Rho/ROCK pathway to promote actin assembly and cell invasion of colorectal cancer.

    PubMed

    Zeng, Yuanfeng; Xie, Huijun; Qiao, Yudan; Wang, Jianmei; Zhu, Xiling; He, Guoyang; Li, Yuling; Ren, Xiaoli; Wang, Feifei; Liang, Li; Ding, Yanqing

    2015-10-01

    Formin-like2 (FMNL2) is a member of the diaphanous-related formins family, which act as effectors and upstream modulators of Rho GTPases signaling and control the actin-dependent processes, such as cell motility or invasion. FMNL2 has been identified as promoting the motility and metastasis in colorectal carcinoma (CRC). However, whether FMNL2 regulates Rho signaling to promote cancer cell invasion remains unclear. In this study, we demonstrated an essential role for FMNL2 in the activations of Rho/ROCK pathway, SRF transcription or actin assembly, and subsequent CRC cell invasion. FMNL2 could activate Rho/ROCK pathway, and required ROCK to promote CRC cell invasion. Moreover, FMNL2 promoted the formation of filopodia and stress fiber, and activated the SRF transcription in a Rho-dependent manner. We also demonstrated that FMNL2 was necessary for LPA-induced invasion, RhoA/ROCK activation, actin assembly and SRF activation. FMNL2 was an essential component of LPA signal transduction toward RhoA by directly interacting with LARG. LARG silence inhibited RhoA/ROCK pathway and CRC cell invasion. Collectively, these data indicate that FMNL2, acting as upstream of RhoA by interacting with LARG, can promote actin assembly and CRC cell invasion through a Rho/ROCK-dependent mechanism.

  2. TrkB-T1 regulates the RhoA signaling and actin cytoskeleton in glioma cells

    SciTech Connect

    Ohira, Koji; Homma, Koichi J.; Hirai, Hirohisa; Nakamura, Shun; Hayashi, Motoharu . E-mail: hayashi@pri.kyoto-u.ac.jp

    2006-04-14

    Recently, the truncated TrkB receptor, T1, has been reported to be involved in the control of cell morphology via the regulation of Rho proteins, through which T1 binds Rho guanine nucleotide dissociation inhibitor (Rho GDI) 1 and dissociates it in a brain-derived neurotrophic factor (BDNF)-dependent manner. However, it is unclear whether T1 signaling regulates the downstream of Rho signaling and the actin cytoskeleton. In this study, we investigated this question using C6 rat glioma cells, which express T1 endogenously. Rho GDI1 was dissociated from T1 in a BDNF-dependent manner, which also causes decreases in the activities of Rho-signaling molecules such as RhoA, Rho-associated kinase, p21-activated kinase, and extracellular-signal regulated kinase1/2. Moreover, BDNF treatment resulted in the disappearance of stress fibers in the cells treated with lysophosphatidic acid, an activator of RhoA, and in morphological changes in cells. Furthermore, a competitive assay with cyan fluorescent protein fusion proteins of T1-specific sequences reduced the effects of BDNF. These results suggest that T1 regulates the Rho-signaling pathways and the actin cytoskeleton.

  3. Knockdown of RhoA expression alters ovarian cancer biological behavior in vitro and in nude mice.

    PubMed

    Wang, Xiaoxia; Jiang, Wenyan; Kang, Jiali; Liu, Qicai; Nie, Miaoling

    2015-08-01

    RhoA regulates cell proliferation, migration, angiogenesis and gene expression. Altered RhoA activity contributes to cancer progression. The present study investigated the effects of RhoA knockdown on the regulation of ovarian cancer biological behavior in vitro and in nude mice. The expression of RhoA was knocked down using a lentivirus carrying RhoA short hairpin RNA (shRNA) in ovarian cancer cells and was confirmed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis. The altered ovarian cancer biological behaviors were assayed by cell viability, terminal deoxynucleotidyltransferase-mediated dUTP nick end-labeling (TUNEL), migration, invasion, and nude mice tumorigenicity assays, while the altered gene expression was detected by RT-qPCR and western blot analysis. The results showed that lentivirus-carrying RhoA shRNA significantly suppressed RhoA expression in ovarian cancer cells, which suppressed tumor cell viability, migration, invasion and adhesion in vitro. RhoA silencing also inhibited the tumorigenicity of ovarian cancer cells in nude mice, which was characterized by the suppression of tumor xenograft formation and growth and induction of tumor cell apoptosis. The results of the present study demonstrated that knockdown of RhoA expression had a significant antitumor effect on ovarian cancer cells in vitro and in nude mice, suggesting that RhoA may be a target for the development of a novel therapeutic strategy in the control of ovarian cancer.

  4. ALPHA CONTAMINATION MONITORING

    DTIC Science & Technology

    This project was conducted to determine the alpha hazard existing in the vicinity of the missile launch pad following the destruction of a missile ...were used for plutonium particle collection. Because all warhead-carrying missiles were properly launched after Project 2.3 was approved, no alpha contamination data was obtained.

  5. Imaging alpha particle detector

    DOEpatents

    Anderson, David F.

    1985-01-01

    A method and apparatus for detecting and imaging alpha particles sources is described. A conducting coated high voltage electrode (1) and a tungsten wire grid (2) constitute a diode configuration discharge generator for electrons dislodged from atoms or molecules located in between these electrodes when struck by alpha particles from a source (3) to be quantitatively or qualitatively analyzed. A thin polyester film window (4) allows the alpha particles to pass into the gas enclosure and the combination of the glass electrode, grid and window is light transparent such that the details of the source which is imaged with high resolution and sensitivity by the sparks produced can be observed visually as well. The source can be viewed directly, electronically counted or integrated over time using photographic methods. A significant increase in sensitivity over other alpha particle detectors is observed, and the device has very low sensitivity to gamma or beta emissions which might otherwise appear as noise on the alpha particle signal.

  6. Imaging alpha particle detector

    DOEpatents

    Anderson, D.F.

    1980-10-29

    A method and apparatus for detecting and imaging alpha particles sources is described. A dielectric coated high voltage electrode and a tungsten wire grid constitute a diode configuration discharge generator for electrons dislodged from atoms or molecules located in between these electrodes when struck by alpha particles from a source to be quantitatively or qualitatively analyzed. A thin polyester film window allows the alpha particles to pass into the gas enclosure and the combination of the glass electrode, grid and window is light transparent such that the details of the source which is imaged with high resolution and sensitivity by the sparks produced can be observed visually as well. The source can be viewed directly, electronically counted or integrated over time using photographic methods. A significant increase in sensitivity over other alpha particle detectors is observed, and the device has very low sensitivity to gamma or beta emissions which might otherwise appear as noise on the alpha particle signal.

  7. Event counting alpha detector

    DOEpatents

    Bolton, Richard D.; MacArthur, Duncan W.

    1996-01-01

    An electrostatic detector for atmospheric radon or other weak sources of alpha radiation. In one embodiment, nested enclosures are insulated from one another, open at the top, and have a high voltage pin inside and insulated from the inside enclosure. An electric field is produced between the pin and the inside enclosure. Air ions produced by collision with alpha particles inside the decay volume defined by the inside enclosure are attracted to the pin and the inner enclosure. With low alpha concentrations, individual alpha events can be measured to indicate the presence of radon or other alpha radiation. In another embodiment, an electrical field is produced between parallel plates which are insulated from a single decay cavity enclosure.

  8. Event counting alpha detector

    DOEpatents

    Bolton, R.D.; MacArthur, D.W.

    1996-08-27

    An electrostatic detector is disclosed for atmospheric radon or other weak sources of alpha radiation. In one embodiment, nested enclosures are insulated from one another, open at the top, and have a high voltage pin inside and insulated from the inside enclosure. An electric field is produced between the pin and the inside enclosure. Air ions produced by collision with alpha particles inside the decay volume defined by the inside enclosure are attracted to the pin and the inner enclosure. With low alpha concentrations, individual alpha events can be measured to indicate the presence of radon or other alpha radiation. In another embodiment, an electrical field is produced between parallel plates which are insulated from a single decay cavity enclosure. 6 figs.

  9. Alpha-particle diagnostics

    SciTech Connect

    Young, K.M.

    1991-01-01

    This paper will focus on the state of development of diagnostics which are expected to provide the information needed for {alpha}- physics studies in the future. Conventional measurement of detailed temporal and spatial profiles of background plasma properties in DT will be essential for such aspects as determining heating effectiveness, shaping of the plasma profiles and effects of MHD, but will not be addressed here. This paper will address (1) the measurement of the neutron source, and hence {alpha}-particle birth profile, (2) measurement of the escaping {alpha}-particles and (3) measurement of the confined {alpha}-particles over their full energy range. There will also be a brief discussion of (4) the concerns about instabilities being generated by {alpha}-particles and the methods necessary for measuring these effects. 51 refs., 10 figs.

  10. Reexamination of the {alpha}-{alpha}''fishbone'' potential

    SciTech Connect

    Day, J. P.; McEwen, J. E.; Elhanafy, M.; Smith, E.; Woodhouse, R.; Papp, Z.

    2011-09-15

    The fishbone potential of composite particles simulates the Pauli effect by nonlocal terms. We determine the {alpha}-{alpha} fishbone potential by simultaneously fitting to two-{alpha} resonance energies, experimental phase shifts, and three-{alpha} binding energies. We found that, essentially, a simple Gaussian can provide a good description of two-{alpha} and three-{alpha} experimental data without invoking three-body potentials.

  11. The IL-1 receptor and Rho directly associate to drive cell activation in inflammation

    PubMed Central

    Singh, R.; Wang, B.; Shirvaikar, A.; Khan, S.; Kamat, S.; Schelling, J.R.; Konieczkowski, M.; Sedor, J.R.

    1999-01-01

    IL-1–stimulated mesenchymal cells model molecular mechanisms of inflammation. Binding of IL-1 to the type I IL-1 receptor (IL-1R) clusters a multi-subunit signaling complex at focal adhesion complexes. Since Rho family GTPases coordinately organize actin cytoskeleton and signaling to regulate cell phenotype, we hypothesized that the IL-1R signaling complex contained these G proteins. IL-1 stimulated actin stress fiber formation in serum-starved HeLa cells in a Rho-dependent manner and rapidly activated nucleotide exchange on RhoA. Glutathione S-transferase (GST) fusion proteins, containing either the full-length IL-1R cytosolic domain (GST-IL-1Rcd) or the terminal 68 amino acids of IL-1R required for IL-1–dependent signal transduction, specifically coprecipitated both RhoA and Rac-1, but not p21ras, from Triton-soluble HeLa cell extracts. In whole cells, a small-molecular-weight G protein coimmunoprecipitated by anti–IL-1R antibody was a substrate for C3 transferase, which specifically ADP-ribosylates Rho GTPases. Constitutively activated RhoA, loaded with [γ-32P]GTP, directly interacted with GST-IL-1Rcd in a filter-binding assay. The IL-1Rcd-RhoA interaction was functionally important, since a dominant inhibitory mutant of RhoA prevented IL-1Rcd–directed transcriptional activation of the IL-6 gene. Consistent with our previous data demonstrating that IL-1R–associated myelin basic protein (MBP) kinases are necessary for IL-1–directed gene expression, cellular incorporation of C3 transferase inhibited IL-1R–associated MBP kinase activity both in solution and in gel kinase assays. In summary, IL-1 activated RhoA, which was physically associated with IL-1Rcd and necessary for activation of cytosolic nuclear signaling pathways. These findings suggest that IL-1–stimulated, Rho-dependent cytoskeletal reorganization may cluster signaling molecules in specific architectures that are necessary for persistent cell activation in chronic inflammatory disease

  12. Activator-inhibitor coupling between Rho signaling and actin assembly make the cell cortex an excitable medium

    PubMed Central

    Bement, William M.; Leda, Marcin; Moe, Alison M.; Kita, Angela M.; Larson, Matthew E.; Golding, Adriana E.; Pfeuti, Courtney; Su, Kuan-Chung; Miller, Ann L.; Goryachev, Andrew B.; von Dassow, George

    2016-01-01

    Animal cell cytokinesis results from patterned activation of the small GTPase Rho, which directs assembly of actomyosin in the equatorial cortex. Cytokinesis is restricted to a portion of the cell cycle following anaphase onset in which the cortex is responsive to signals from the spindle. We show that shortly after anaphase onset oocytes and embryonic cells of frogs and echinoderms exhibit cortical waves of Rho activity and F-actin polymerization. The waves are modulated by cyclin-dependent kinase 1 (Cdk1) activity and require the Rho GEF (guanine nucleotide exchange factor), Ect2. Surprisingly, during wave propagation, while Rho activity elicits F-actin assembly, F-actin subsequently inactivates Rho. Experimental and modeling results show that waves represent excitable dynamics of a reaction diffusion system with Rho as the activator and F-actin the inhibitor. We propose that cortical excitability explains fundamental features of cytokinesis including its cell cycle regulation. PMID:26479320

  13. Mycobacterium tuberculosis Rho Is an NTPase with Distinct Kinetic Properties and a Novel RNA-Binding Subdomain

    PubMed Central

    Mitra, Anirban; Misquitta, Rachel; Nagaraja, Valakunja

    2014-01-01

    Two mechanisms – factor independent and dependent termination – ensure the completion of RNA synthesis in eubacteria. Factor-dependent mechanism relies on the Rho protein to terminate transcription by interacting with RNA polymerase. Although well studied in Escherichia coli, the properties of the Rho homologs from most bacteria are not known. The rho gene is unusually large in genus Mycobacterium and other members of actinobacteria, having ∼150 additional residues towards the amino terminal end. We describe the distinct properties of Rho from Mycobacterium tuberculosis. It is an NTPase with a preference for purine nucleoside triphosphates with kinetic properties different from E. coli homolog and an ability to use various RNA substrates. The N-terminal subdomain of MtbRho can bind to RNA by itself, and appears to contribute to the interaction of the termination factor with RNAs. Furthermore, the interaction with RNA induces changes in conformation and oligomerization of MtbRho. PMID:25229539

  14. Simple enzymatic assays for the in vitro motor activity of transcription termination factor Rho from Escherichia coli.

    PubMed

    Boudvillain, Marc; Walmacq, Céline; Schwartz, Annie; Jacquinot, Frédérique

    2010-01-01

    The transcription termination factor Rho from Escherichia coli is a ring-shaped homo-hexameric protein that preferentially interacts with naked cytosine-rich Rut (Rho utilization) regions of nascent RNA transcripts. Once bound to the RNA chain, Rho uses ATP as an energy source to produce mechanical work and disruptive forces that ultimately lead to the dissociation of the ternary transcription complex. Although transcription termination assays have been useful to study Rho activity in various experimental contexts, they do not report directly on Rho mechanisms and kinetics. Here, we describe complementary ATP-dependent RNA-DNA helicase and streptavidin displacement assays that can be used to monitor in vitro Rho's motor activity in a more direct and quantitative manner.

  15. Quantitative characterization of gene regulation by Rho dependent transcription termination.

    PubMed

    Hussein, Razika; Lee, Tiffany Y; Lim, Han N

    2015-08-01

    Rho factor dependent transcription termination (RTT) is common within the coding sequences of bacterial genes and it acts to couple transcription and translation levels. Despite the importance of RTT for gene regulation, its effects on mRNA and protein concentrations have not been quantitatively characterized. Here we demonstrate that the exogenous cfp gene encoding the cyan fluorescent protein can serve as a model for gene regulation by RTT. This was confirmed by showing that Psu and bicyclomycin decrease RTT and increase full length cfp mRNAs (but remarkably they have little effect on protein production). We then use cfp to characterize the relationship between its protein and full length mRNA concentrations when the translation initiation rate is varied by sequence modifications of the translation initiation region (TIR). These experiments reveal that the fold change in protein concentration (RP) and the fold change in full length mRNA concentration (Rm) have the relationship RP≈Rm(b), where b is a constant. The average value of b was determined from three separate data sets to be ~3.6. We demonstrate that the above power law function can predict how altering the translation initiation rate of a gene in an operon will affect the mRNA concentrations of downstream genes and specify a lower bound for the associated changes in protein concentrations. In summary, this study defines a simple phenomenological model to help program expression from single genes and operons that are regulated by RTT, and to guide molecular models of RTT.

  16. PRL-1 protein promotes ERK1/2 and RhoA protein activation through a non-canonical interaction with the Src homology 3 domain of p115 Rho GTPase-activating protein.

    PubMed

    Bai, Yunpeng; Luo, Yong; Liu, Sijiu; Zhang, Lujuan; Shen, Kui; Dong, Yuanshu; Walls, Chad D; Quilliam, Lawrence A; Wells, Clark D; Cao, Youjia; Zhang, Zhong-Yin

    2011-12-09

    Phosphatases of the regenerating liver (PRL) play oncogenic roles in cancer development and metastasis. Although previous studies indicate that PRL-1 promotes cell growth and migration by activating both the ERK1/2 and RhoA pathways, the mechanism by which it activates these signaling events remains unclear. We have identified a PRL-1-binding peptide (Peptide 1) that shares high sequence identity with a conserved motif in the Src homology 3 (SH3) domain of p115 Rho GTPase-activating protein (GAP). p115 RhoGAP directly binds PRL-1 in vitro and in cells via its SH3 domain. Structural analyses of the PRL-1·Peptide 1 complex revealed a novel protein-protein interaction whereby a sequence motif within the PxxP ligand-binding site of the p115 RhoGAP SH3 domain occupies a folded groove within PRL-1. This prevents the canonical interaction between the SH3 domain of p115 RhoGAP and MEKK1 and results in activation of ERK1/2. Furthermore, PRL-1 binding activates RhoA signaling by inhibiting the catalytic activity of p115 RhoGAP. The results demonstrate that PRL-1 binding to p115 RhoGAP provides a coordinated mechanism underlying ERK1/2 and RhoA activation.

  17. The alpha channeling effect

    SciTech Connect

    Fisch, N. J.

    2015-12-10

    Alpha particles born through fusion reactions in a tokamak reactor tend to slow down on electrons, but that could take up to hundreds of milliseconds. Before that happens, the energy in these alpha particles can destabilize on collisionless timescales toroidal Alfven modes and other waves, in a way deleterious to energy confinement. However, it has been speculated that this energy might be instead be channeled into useful energy, so as to heat fuel ions or to drive current. Such a channeling needs to be catalyzed by waves Waves can produce diffusion in energy of the alpha particles in a way that is strictly coupled to diffusion in space. If these diffusion paths in energy-position space point from high energy in the center to low energy on the periphery, then alpha particles will be cooled while forced to the periphery. The energy from the alpha particles is absorbed by the wave. The amplified wave can then heat ions or drive current. This process or paradigm for extracting alpha particle energy collisionlessly has been called alpha channeling. While the effect is speculative, the upside potential for economical fusion is immense. The paradigm also operates more generally in other contexts of magnetically confined plasma.

  18. Recognition and activation of Rho GTPases by Vav1 and Vav2 guanine nucleotide exchange factors.

    PubMed

    Heo, Jongyun; Thapar, Roopa; Campbell, Sharon L

    2005-05-03

    Vav proteins are Rho GTPase-specific guanine nucleotide exchange factors (GEFs) that are distinguished by the tandem arrangement of Dbl homology (DH), Pleckstrin homology (PH), and cysteine rich domains (CRD). Whereas the tandem DH-PH arrangement is conserved among Rho GEFs, the presence of the CRD is unique to Vav family members and is required for efficient nucleotide exchange. We provide evidence that Vav2-mediated nucleotide exchange of Rho GTPases follows the Theorell-Chance mechanism in which the Vav2.Rho GTPase complex is the major species during the exchange process and the Vav2.GDP-Mg(2+).Rho GTPase ternary complex is present only transiently. The GTPase specificity for the DH-PH-CRD Vav2 in vitro follows this order: Rac1 > Cdc42 > RhoA. Results obtained from fluorescence anisotropy and NMR chemical shift mapping experiments indicate that the isolated Vav1 CRD is capable of directly associating with Rac1, and residues K116 and S83 that are in the proximity of the P-loop and the guanine base either are part of this binding interface or undergo a conformational change in response to CRD binding. The NMR studies are supported by kinetic measurements on Rac1 mutants S83A, K116A, and K116Q and Vav2 CRD mutant K533A in that these mutants affect both the initial binding event of Vav2 with Rac1 (k(on)) and the rate-limiting dissociation of Vav2 from the Vav2.Rac1 binary complex (thereby influencing the enzyme turnover number, k(cat)). The results suggest that the CRD domain in Vav proteins plays an active role, affecting both the k(on) and the k(cat) for Vav-mediated nucleotide exchange on Rho GTPases.

  19. Autophagy suppresses cell migration by degrading GEF-H1, a RhoA GEF.

    PubMed

    Yoshida, Tatsushi; Tsujioka, Masatsune; Honda, Shinya; Tanaka, Masato; Shimizu, Shigeomi

    2016-06-07

    Cell migration is a process crucial for a variety of biological events, such as morphogenesis and wound healing. Several reports have described the possible regulation of cell migration by autophagy; however, this remains controversial. We here demonstrate that mouse embryonic fibroblasts (MEFs) lacking autophagy protein 5 (Atg5), an essential molecule of autophagy, moved faster than wild-type (WT) MEFs. Similar results were obtained for MEFs lacking Atg7 and unc-51-like kinase 1 (Ulk1), which are molecules required for autophagy. This phenotype was also observed in Atg7-deficient macrophages. WT MEFs moved by mesenchymal-type migration, whereas Atg5 knockout (KO) MEFs moved by amoeba-like migration. This difference was thought to be mediated by the level of RhoA activity, because Atg5 KO MEFs had higher RhoA activity, and treatment with a RhoA inhibitor altered Atg5 KO MEF migration from the amoeba type to the mesenchymal type. Autophagic regulation of RhoA activity was dependent on GEF-H1, a member of the RhoA family of guanine nucleotide exchange factors. In WT MEFs, GEF-H1 directly bound to p62 and was degraded by autophagy, resulting in low RhoA activity. In contrast, the loss of autophagy increased GEF-H1 levels and thereby activated RhoA, which caused cells to move by amoeba-like migration. This amoeba-like migration was cancelled by the silencing of GEF-H1. These results indicate that autophagy plays a role in the regulation of migration by degrading GEF-H1.

  20. Amphetamine activates Rho GTPase signaling to mediate dopamine transporter internalization and acute behavioral effects of amphetamine

    PubMed Central

    Wheeler, David S.; Underhill, Suzanne M.; Stolz, Donna B.; Murdoch, Geoffrey H.; Thiels, Edda; Romero, Guillermo; Amara, Susan G.

    2015-01-01

    Acute amphetamine (AMPH) exposure elevates extracellular dopamine through a variety of mechanisms that include inhibition of dopamine reuptake, depletion of vesicular stores, and facilitation of dopamine efflux across the plasma membrane. Recent work has shown that the DAT substrate AMPH, unlike cocaine and other nontransported blockers, can also stimulate endocytosis of the plasma membrane dopamine transporter (DAT). Here, we show that when AMPH enters the cytoplasm it rapidly stimulates DAT internalization through a dynamin-dependent, clathrin-independent process. This effect, which can be observed in transfected cells, cultured dopamine neurons, and midbrain slices, is mediated by activation of the small GTPase RhoA. Inhibition of RhoA activity with C3 exotoxin or a dominant-negative RhoA blocks AMPH-induced DAT internalization. These actions depend on AMPH entry into the cell and are blocked by the DAT inhibitor cocaine. AMPH also stimulates cAMP accumulation and PKA-dependent inactivation of RhoA, thus providing a mechanism whereby PKA- and RhoA-dependent signaling pathways can interact to regulate the timing and robustness of AMPH’s effects on DAT internalization. Consistent with this model, the activation of D1/D5 receptors that couple to PKA in dopamine neurons antagonizes RhoA activation, DAT internalization, and hyperlocomotion observed in mice after AMPH treatment. These observations support the existence of an unanticipated intracellular target that mediates the effects of AMPH on RhoA and cAMP signaling and suggest new pathways to target to disrupt AMPH action. PMID:26553986

  1. Regulation of pancreatic cancer cell migration and invasion by RhoC GTPase and Caveolin-1

    PubMed Central

    Lin, Min; DiVito, Melinda M; Merajver, Sofia D; Boyanapalli, Madanamohan; van Golen, Kenneth L

    2005-01-01

    Background In the current study we investigated the role of caveolin-1 (cav-1) in pancreatic adenocarcinoma (PC) cell migration and invasion; initial steps in metastasis. Cav-1 is the major structural protein in caveolae; small Ω-shaped invaginations within the plasma membrane. Caveolae are involved in signal transduction, wherein cav-1 acts as a scaffolding protein to organize multiple molecular complexes regulating a variety of cellular events. Recent evidence suggests a role for cav-1 in promoting cancer cell migration, invasion and metastasis; however, the molecular mechanisms have not been described. The small monomeric GTPases are among several molecules which associate with cav-1. Classically, the Rho GTPases control actin cytoskeletal reorganization during cell migration and invasion. RhoC GTPase is overexpressed in aggressive cancers that metastasize and is the predominant GTPase in PC. Like several GTPases, RhoC contains a putative cav-1 binding motif. Results Analysis of 10 PC cell lines revealed high levels of cav-1 expression in lines derived from primary tumors and low expression in those derived from metastases. Comparison of the BxPC-3 (derived from a primary tumor) and HPAF-II (derived from a metastasis) demonstrates a reciprocal relationship between cav-1 expression and p42/p44 Erk activation with PC cell migration, invasion, RhoC GTPase and p38 MAPK activation. Furthermore, inhibition of RhoC or p38 activity in HPAF-II cells leads to partial restoration of cav-1 expression. Conclusion Cav-1 expression inhibits RhoC GTPase activation and subsequent activation of the p38 MAPK pathway in primary PC cells thus restricting migration and invasion. In contrast, loss of cav-1 expression leads to RhoC-mediated migration and invasion in metastatic PC cells. PMID:15969750

  2. Inhibition of Rho-kinase differentially affects axon regeneration of peripheral motor and sensory nerves.

    PubMed

    Joshi, Abhijeet R; Bobylev, Ilja; Zhang, Gang; Sheikh, Kazim A; Lehmann, Helmar C

    2015-01-01

    The small GTPase RhoA and its down-stream effector Rho-kinase (ROCK) are important effector molecules of the neuronal cytoskeleton. Modulation of the RhoA/ROCK pathway has been shown to promote axonal regeneration, however in vitro and animal studies are inconsistent regarding the extent of axonal outgrowth induced by pharmacological inhibition of ROCK. We hypothesized that injury to sensory and motor nerves result in diverse activation levels of RhoA, which may impact the response of those nerve fiber modalities to ROCK inhibition. We therefore examined the effects of Y-27632, a chemical ROCK inhibitor, on the axonal outgrowth of peripheral sensory and motor neurons grown in the presence of growth-inhibiting chondroitin sulfate proteoglycans (CSPGs). In addition we examined the effects of three different doses of Y-27632 on nerve regeneration of motor and sensory nerves in animal models of peripheral nerve crush. In vitro, sensory neurons were less responsive to Y-27632 compared to motor neurons in a non-growth permissive environment. These differences were associated with altered expression and activation of RhoA in sensory and motor axons. In vivo, systemic treatment with high doses of Y-27632 significantly enhanced the regeneration of motor axons over short distances, while the regeneration of sensory fibers remained largely unchanged. Our results support the concept that in a growth non-permissive environment, the regenerative capacity of sensory and motor axons is differentially affected by the RhoA/ROCK pathway, with motor neurons being more responsive compared to sensory. Future treatments, that are aimed to modulate RhoA activity, should consider this functional diversity.

  3. Asbestos induces nitric oxide synthesis in mesothelioma cells via Rho signaling inhibition.

    PubMed

    Riganti, Chiara; Orecchia, Sara; Silvagno, Francesca; Pescarmona, Gianpiero; Betta, Pier Giacomo; Gazzano, Elena; Aldieri, Elisabetta; Ghigo, Dario; Bosia, Amalia

    2007-06-01

    We have observed that in three human malignant mesothelioma cell lines, crocidolite asbestos induced the activation of the transcription factor NF-kappaB and the synthesis of nitric oxide (NO) by inhibiting the RhoA signaling pathway. The incubation with crocidolite decreased the level of GTP-bound RhoA and the activity of Rho-dependent kinase, and induced the activation of Akt/PKB and IkBalpha kinase, leading to the nuclear translocation of NF-kappaB. The effects of crocidolite fibers on NF-kappaB activation and NO synthesis were mimicked by Y27632 (an inhibitor of the Rho-dependent kinases) and toxin B (an inhibitor of RhoA GTPase activity), while they were reverted by mevalonic acid, the product of 3-hydroxy-3-methylglutaryl coenzyme A (HMGCoA) reductase. Furthermore, crocidolite, similarly to mevastatin, inhibited the synthesis of cholesterol and ubiquinone and the prenylation of RhoA: these effects were prevented in the presence of mevalonic acid. This suggests that crocidolite fibers might inhibit the synthesis of isoprenoid molecules at the level of the HMGCoA reductase reaction or of an upstream step, thus impairing the prenylation and subsequent activation of RhoA. Akt can stimulate NO synthesis via a double mechanism: it can activate the inducible NO synthase via the NF-kappaB pathway and the endothelial NO synthase via a direct phosphorylation. Our results suggest that crocidolite increases the NO levels in mesothelioma cells by modulating both NO synthase isoforms.

  4. WinRho: Rh immune globulin prepared by ion exchange for intravenous use.

    PubMed Central

    Bowman, J M; Friesen, A D; Pollock, J M; Taylor, W E

    1980-01-01

    An Rh immune globulin [Rh IgG] for intravenous use, WinRho, has been prepared by the Winnipeg Rh Institute by a modification of the ion-exchange column method of Hoppe and colleagues. When administered to Rh-negative male and nonpregnant female volunteers WinRho was found to be nonpyrogenic, nontoxic, safe and protective against Rh alloimmunization. In a clinical trial with 240 microgram given at about 28 weeks' gestation and 120 microgram given after delivery to Rh-negative women at risk of Rh immunization WinRho was effective in preventing Rh immunization. Of the 870 women carrying Rh-positive fetuses who were treated with WinRho during pregnancy and were not tested several months after delivery 14 would have shown evidence of Rh immunization by the time of delivery if WinRho had been ineffective; none showed such evidence. Of the 1122 women carrying Rh-positive fetuses who were retested 4 to 6 months after delivery 83 would have shown evidence of Rh immunization at that time if WinRho had been ineffective; only 1 showed such evidence. The efficiency of yield of anti-D with the modified method of production, the fct that it can be given intravenously (a route that causes the patient less discomfort and immediately results in high anti-D levels) and the lower levels of contaminating IgA and IgM make WinRho the preparation of choice for preventing Rh immunization. PMID:6161687

  5. GDP dissociation inhibitor D4-GDI (Rho-GDI 2), but not the homologous rho-GDI 1, is cleaved by caspase-3 during drug-induced apoptosis.

    PubMed Central

    Essmann, F; Wieder, T; Otto, A; Müller, E C; Dörken, B; Daniel, P T

    2000-01-01

    Different cytotoxic drugs induce cell death by activating the apoptotic programme; a family of cysteinyl aspartate proteases named caspases has been shown to be involved in the initiation as well as the execution of this kind of cell death. In the present study, cleavage of D4-GDI (Rho-GDI 2), an abundant haemopoietic-cell GDP dissociation inhibitor for the Ras-related Rho family GTPases, was demonstrated after treatment of BJAB Burkitt-like lymphoma cells with taxol or epirubicin. The cleavage of D4-GDI occurred simultaneously with the activation of caspase-3 but preceded DNA fragmentation and the morphological changes associated with apoptotic cell death. By using high-resolution two-dimensional gel electrophoresis it was shown that this cleavage is specific: whereas the level of the homologous protein Rho-GDI 1 was not significantly altered during drug-induced apoptosis and in cytochrome c/dATP-activated cellular extracts, D4-GDI disappeared owing to proteolytic cleavage. Inhibitor experiments with Z-DEVD-fmk (in which Z stands for benzyloxycarbonyl and fmk for fluoromethyl ketone) and microsequencing of the D4-GDI fragment revealed that this occurs at the caspase-3 cleavage site. Our results strongly suggest the differential regulation of the homologous GDP dissociation inhibitors Rho-GDI 1 and D4-GDI during drug-induced apoptosis by proteolysis mediated by caspase-3 but not by caspase-1. Owing to their crucial role as modulators of Rho GTPases, this might in turn have a significant impact on the mechanisms that induce the cytoskeletal and morphological changes in apoptotic cells. PMID:10698706

  6. Role of Wasp and the small GTPases RhoA, RhoB, and Cdc42 during capacitation and acrosome reaction in spermatozoa of English guinea pigs.

    PubMed

    Delgado-Buenrostro, Norma L; Mújica, Adela; Chiquete-Felix, Natalia; Déciga-Alcaraz, Alejandro; Medina-Reyes, Estefany I; Uribe-Carvajal, Salvador; Chirino, Yolanda I

    2016-10-01

    Cytoskeleton remodeling is necessary for capacitation and the acrosome reaction in spermatozoa. F-actin is located in the acrosome and equatorial region during capacitation, but is relocated in the post-acrosomal region during the acrosome reaction in spermatozoa from bull, rat, mice, and guinea pig. Actin polymerization and relocalization are generally regulated by small GTPases that activate Wasp protein, which coordinates with Arp2/3, profilin I, and profilin II to complete cytoskeletal remodeling. This sequence of events is not completely described in spermatozoa, though. Therefore, the aim of this study was to determine if Wasp interacts with small GTPases (RhoA, RhoB, and Cdc42) and proteins (Arp2/3, profilin I, and profilin II) that co-localize with F-actin during capacitation and the acrosome reaction in English guinea pig spermatozoa obtained from the vas deferens. The spermatozoa were capacitated in calcium-free medium, incubated with an activator or an inhibitor of GTPases, and then induced to acrosome react using calcium. The distribution patterns of F-actin were compared to the patterns of Wasp and its putative interaction partners: Wasp and RhoB, but not RhoA or Cdc42, localization overlap with F-actin during capacitation and the acrosome reaction. Activation of small GTPases localized RhoB to the post-acrosomal region whereas their inhibition prevented acrosome exocytosis. Arp2/3 and profilin II appear to interact with Wasp in the post-acrosomal region and flagellum, while profilin I and Wasp could be found in the equatorial region. Thus, Wasp and F-actin distribution overlap during capacitation and acrosome reaction, and small GTPases play an important role in cytoskeleton remodeling during these processes in spermatozoa. Mol. Reprod. Dev. 83: 927-937, 2016 © 2016 Wiley Periodicals, Inc.

  7. Role of Rho GDP dissociation inhibitor α in control of epithelial sodium channel (ENaC)-mediated sodium reabsorption.

    PubMed

    Pavlov, Tengis S; Levchenko, Vladislav; Staruschenko, Alexander

    2014-10-10

    The epithelial sodium channel (ENaC) is expressed in the aldosterone-sensitive distal nephron where it performs sodium reabsorption from the lumen. We have recently shown that ENaC activity contributes to the development of salt-induced hypertension as a result of deficiency of EGF level. Previous studies revealed that Rho GDP-dissociation inhibitor α (RhoGDIα) is involved in the control of salt-sensitive hypertension and renal injury via Rac1, which is one of the small GTPases activating ENaC. Here we investigated the intracellular mechanism mediating the involvement of the RhoGDIα/Rac1 axis in the control of ENaC and the effect of EGF on ENaC in this pathway. We demonstrated that RhoGDIα is highly expressed in the cortical collecting ducts of mice and rats, and its expression is down-regulated in Dahl salt-sensitive rats fed a high salt diet. Knockdown of RhoGDIα in cultured cortical collecting duct principal cells increased ENaC subunits expression and ENaC-mediated sodium reabsorption. Furthermore, RhoGDIα deficiency causes enhanced response to EGF treatment. Patch clamp analysis reveals that RhoGDIα significantly decreases ENaC current density and prevents its up-regulation by RhoA and Rac1. Inhibition of Rho kinase with Y27632 had no effects on ENaC response to EGF either in control or RhoGDIα knocked down cells. However, EGF treatment increased levels of active Rac1, which was further enhanced in RhoGDIα-deficient cells. We conclude that changes in the RhoGDIα-dependent pathway have a permissive role in the Rac1-mediated enhancement of ENaC activity observed in salt-induced hypertension.

  8. Lentivirus-Mediated RNA Interference Targeting RhoA Slacks the Migration, Proliferation, and Myelin Formation of Schwann Cells.

    PubMed

    Wen, Jinkun; Qian, Changhui; Pan, Mengjie; Wang, Xianghai; Li, Yuanyuan; Lu, Yanmeng; Zhou, Zhitao; Yan, Qing; Li, Lixia; Liu, Zhongying; Wu, Wutian; Guo, Jiasong

    2017-03-01

    RhoA, a member of Rho GTPases family, is known to play an important role in remodeling actin cytoskeleton. During the development of the peripheral nervous system (PNS), Schwann cells undergo proliferation, migration, and radial sorting and finally wrap the related axons compactly to form myelin sheath. All these processes involve actin cytoskeletal remodeling. However, the role of RhoA on Schwann cell during development is still unclear. To address this question, we first used a lentiviral vector-mediated short hairpin (sh) RNA targeting RhoA to knock down the expression of RhoA in the cultured Schwann cells in vitro. Effects of RhoA on Schwann cell proliferation and migration were examined by BrdU assay and transwell assay, respectively. Results of the present study indicated that downregulated RhoA expression in cultured Schwann cells significantly slacked the cells' capabilities of migration and proliferation. Then, we investigated the role of RhoA in the developing rat sciatic nerves. Immunohistology and Western blotting showed that RhoA was mainly expressed in Schwann cells in the sciatic nerves and was peaked at 2 weeks postnatal then kept in low level up to 8 weeks. In the subjected rats whose sciatic nerves were microinjected with lentiviral vectors at postnatal 3 days, we found that the lentiviruses mainly transfected Schwann cells, and the RhoA expression in the transfected Schwann cells was significantly knocked down. Four weeks after lentivirus microinjection, immunohistology and transmission electron microscopy illustrated that RhoA knockdown resulted in hypomyelination and significant decrease of the thickness of myelin in the transfected area. Overall data of current study suggested that RhoA plays a critical role in Schwann cell biology and is essential for myelination in developing peripheral nerve.

  9. RhoB Acts as a Tumor Suppressor That Inhibits Malignancy of Clear Cell Renal Cell Carcinoma

    PubMed Central

    Ma, Xin; Zhang, Peng; Gao, Yu; Fan, Yang; Pang, Haigang; Gong, Huijie; Shen, Donglai; Gu, Liangyou; Zhang, Yu

    2016-01-01

    This study aims to investigate the biological role of RhoB in clear cell renal cell carcinoma (ccRCC). The expression of RhoB was examined in specimens of patients and cell lines by Western blot and Immunohistochemistry. The correlation between RhoB expression and clinicopathologic variables was also analyzed. The effects of RhoB on cell proliferation, cell cycle, cell apoptosis, and invasion/migration were detected by over-expression and knockdown of RhoB level in ccRCC cells via plasmids and RNAi. The results showed that RhoB was low-expressed in ccRCC surgical specimens and cell lines compared with adjacent normal renal tissues and normal human renal proximal tubular epithelial cell lines (HKC), and its protein expression level was significantly associated with the tumor pathologic parameter embracing tumor size(P = 0.0157), pT stage(P = 0.0035), TNM stage(P = 0.0024) and Fuhrman tumor grade(P = 0.0008). Further, over-expression of RhoB remarkably inhibited the cancer cell proliferation, colony formation and promoted cancer cell apoptosis, and aslo reduced the invasion and migration ability of ccRCC cells. Interestingly, up-regulation of RhoB could induce cell cycle arrest in G2/M phase and led to cell cycle regulators(CyclineB1,CDK1) and pro-apoptotic protein(casp3,casp9) aberrant expression. Moreover, knockdown of RhoB in HKC cells promoted cell proliferation and migration. Taken together, our study indicates that RhoB expression is decreased in ccRCC carcinogenesis and progression. Up-regulation of RhoB significantly inhibits ccRCC cell malignant phenotype. These findings show that RhoB may play a tumor suppressive role in ccRCC cells, raising its potential value in futural therapeutic target for the patients of ccRCC. PMID:27384222

  10. Prenylation of Rho G-proteins: a novel mechanism regulating gene expression and protein stability in human trabecular meshwork cells.

    PubMed

    Stubbs, Evan B; Von Zee, Cynthia L

    2012-08-01

    Endogenous prenylation with sesquiterpene or diterpene isoprenoids facilitates membrane localization and functional activation of small monomeric GTP-binding proteins. A direct effect of isoprenoids on regulation of gene expression and protein stability has also been proposed. In this study, we determined the role of sesquiterpene or diterpene isoprenoids on the regulation of Rho G-protein expression, activation, and stability in human trabecular meshwork (TM) cells. In both primary and transformed human TM cells, limiting endogenous isoprenoid synthesis with lovastatin, a potent HMG-CoA reductase inhibitor, elicited marked increases in RhoA and RhoB mRNA and protein content. The effect of lovastatin was dose-dependent with newly synthesized inactive protein accumulating in the cytosol. Supplementation with geranylgeranyl pyrophosphate (GGPP) prevented, while inhibition of geranylgeranyl transferase-I mimicked, the effects of lovastatin on RhoA and RhoB protein content. Similarly, lovastatin-dependent increases in RhoA and RhoB mRNA expression were mimicked by geranylgeranyl transferase-I inhibition. Interestingly, GGPP supplementation selectively promoted the degradation of newly synthesized Rho proteins which was mediated, in part, through the 20S proteasome. Functionally, GGPP supplementation prevented lovastatin-dependent decreases in actin stress fiber organization while selectively facilitating the subcellular redistribution of accumulated Rho proteins from the cytosol to the membrane and increasing RhoA activation. Post-translational prenylation with geranylgeranyl diterpenes selectively facilitates the expression, membrane translocation, functional activation, and turnover of newly synthesized Rho proteins. Geranylgeranyl prenylation represents a novel mechanism by which active Rho proteins are targeted to the 20S proteasome for degradation in human TM cells.

  11. RhoC is essential for TGF-{beta}1-induced invasive capacity of rat ascites hepatoma cells

    SciTech Connect

    Mukai, M.; Endo, H.; Iwasaki, T.; Tatsuta, M.; Togawa, A.; Nakamura, H.; Inoue, M. . E-mail: inoue-ma2@mc.pref.osaka.jp

    2006-07-21

    Transforming growth factor-{beta}1 (TGF-{beta}1) is a multifunctional growth factor that plays a role in cell proliferation, differentiation, extracellular matrix production, apoptosis, and cell motility. We show here that TGF-{beta}1 increased the invasiveness of MM1 cells, which are a highly invasive clone of rat ascites hepatoma cells. Both mRNA and protein levels of RhoC but not RhoA in TGF-{beta}1-treated MM1 cells increased. In parallel with this increase in expression, RhoC activity was induced by TGF-{beta}1 treatment. When RhoC was overexpressed in MM1 cells, the invasive capacity increased. The RhoC-overexpressing cells formed more nodules than did mock cells when injected into rat peritoneum. Furthermore, when RhoC expression was reduced by transfection with shRNA/RhoC, the invasiveness of MM1 cells decreased with concomitant suppression of RhoC expression. Thus, the induced expression of RhoC by TGF-{beta}1 in MM1 cells plays a critical role in TGF-{beta}1-induced cell migration.

  12. RhoB Promotes γH2AX Dephosphorylation and DNA Double-Strand Break Repair

    PubMed Central

    Mamouni, Kenza; Cristini, Agnese; Guirouilh-Barbat, Josée; Monferran, Sylvie; Lemarié, Anthony; Faye, Jean-Charles; Lopez, Bernard S.

    2014-01-01

    Unlike other Rho GTPases, RhoB is rapidly induced by DNA damage, and its expression level decreases during cancer progression. Because inefficient repair of DNA double-strand breaks (DSBs) can lead to cancer, we investigated whether camptothecin, an anticancer drug that produces DSBs, induces RhoB expression and examined its role in the camptothecin-induced DNA damage response. We show that in camptothecin-treated cells, DSBs induce RhoB expression by a mechanism that depends notably on Chk2 and its substrate HuR, which binds to RhoB mRNA and protects it against degradation. RhoB-deficient cells fail to dephosphorylate γH2AX following camptothecin removal and show reduced efficiency of DSB repair by homologous recombination. These cells also show decreased activity of protein phosphatase 2A (PP2A), a phosphatase for γH2AX and other DNA damage and repair proteins. Thus, we propose that DSBs activate a Chk2-HuR-RhoB pathway that promotes PP2A-mediated dephosphorylation of γH2AX and DSB repair. Finally, we show that RhoB-deficient cells accumulate endogenous γH2AX and chromosomal abnormalities, suggesting that RhoB loss increases DSB-mediated genomic instability and tumor progression. PMID:24912678

  13. Flavone inhibits migration through DLC1/RhoA pathway by decreasing ROS generation in breast cancer cells.

    PubMed

    Zhu, Wenzhen; Ma, Long; Yang, Bingwu; Zheng, Zhaodi; Chai, Rongfei; Liu, Tingting; Liu, Zhaojun; Song, Taiyu; Li, Fenglin; Li, Guorong

    2016-05-01

    Tumor suppressor protein deleted in liver cancer 1 (DLC1) is a RhoGTPase-activating protein (RhoGAP) and inhibits cancer cell migration by inactivating downstream target protein RhoA. A few studies have reported the regulations of reactive oxygen species (ROS) on RhoGAP. In this study, we investigated flavone (the core structure of flavonoids)-induced regulation on ROS generation and DLC1/RhoA pathway in MCF-7 and MDA-MB-231 breast cancer cells and explored whether flavone-induced upregulation of DLC1 is mediated by ROS. Our results showed that flavone decreased ROS production and inhibited cell migration through DLC1/RhoA pathway. To further investigate the role of ROS in flavone-induced regulation on DLC1/RhoA pathway, hydrogen peroxide was added to restore the ROS levels. Flavone-induced upregulation of DLC1 expression, downregulation of RhoA activity, and inhibition of cell migration were all restrained by hydrogen peroxide. We also found that flavone increased DLC1 stability by inhibiting DLC1 protein degradation in breast cancer cells. In summary, our study demonstrated that flavone inhibited cell migration through DLC1/RhoA pathway by decreasing ROS generation and suppressed DLC1 degradation in MCF-7 and MDA-MB-231 breast cancer cells.

  14. Alpha One Foundation

    MedlinePlus

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  15. Crystal structure and novel recognition motif of rho ADP-ribosylating C3 exoenzyme from Clostridium botulinum: structural insights for recognition specificity and catalysis.

    PubMed

    Han, S; Arvai, A S; Clancy, S B; Tainer, J A

    2001-01-05

    Clostridium botulinum C3 exoenzyme inactivates the small GTP-binding protein family Rho by ADP-ribosylating asparagine 41, which depolymerizes the actin cytoskeleton. C3 thus represents a major family of the bacterial toxins that transfer the ADP-ribose moiety of NAD to specific amino acids in acceptor proteins to modify key biological activities in eukaryotic cells, including protein synthesis, differentiation, transformation, and intracellular signaling. The 1.7 A resolution C3 exoenzyme structure establishes the conserved features of the core NAD-binding beta-sandwich fold with other ADP-ribosylating toxins despite little sequence conservation. Importantly, the central core of the C3 exoenzyme structure is distinguished by the absence of an active site loop observed in many other ADP-ribosylating toxins. Unlike the ADP-ribosylating toxins that possess the active site loop near the central core, the C3 exoenzyme replaces the active site loop with an alpha-helix, alpha3. Moreover, structural and sequence similarities with the catalytic domain of vegetative insecticidal protein 2 (VIP2), an actin ADP-ribosyltransferase, unexpectedly implicates two adjacent, protruding turns, which join beta5 and beta6 of the toxin core fold, as a novel recognition specificity motif for this newly defined toxin family. Turn 1 evidently positions the solvent-exposed, aromatic side-chain of Phe209 to interact with the hydrophobic region of Rho adjacent to its GTP-binding site. Turn 2 evidently both places the Gln212 side-chain for hydrogen bonding to recognize Rho Asn41 for nucleophilic attack on the anomeric carbon of NAD ribose and holds the key Glu214 catalytic side-chain in the adjacent catalytic pocket. This proposed bipartite ADP-ribosylating toxin turn-turn (ARTT) motif places the VIP2 and C3 toxin classes into a single ARTT family characterized by analogous target protein recognition via turn 1 aromatic and turn 2 hydrogen-bonding side-chain moieties. Turn 2 centrally anchors

  16. Inhibition of Rho protein stimulates iNOS expression in rat vascular smooth muscle cells.

    PubMed

    Muniyappa, R; Xu, R; Ram, J L; Sowers, J R

    2000-06-01

    Inducible nitric oxide synthase (iNOS) in vascular smooth muscle cells (VSMCs) is upregulated in arterial injury and plays a role in regulating VSMC proliferation and restenosis. Inflammatory cytokines [e.g., interleukin-1beta (IL-1beta)] released during vascular injury induce iNOS. Small GTP-binding proteins of the Ras superfamily play a major role in IL-1beta-dependent signaling pathways. In this study, we examined the role of Rho GTPases in regulating iNOS expression in VSMCs. Treatment of VSMCs with mevastatin, which inhibits isoprenylation of Rho and other small GTP-binding proteins, produced significantly higher amounts of IL-1beta-evoked NO and iNOS protein compared with control. Similarly, bacterial toxins [Toxin B from Clostridium difficile and C3 ADP-ribosyl transferase (C3) toxin from Clostridium botulinium] that specifically inactivate Rho proteins increased NOS products (NO and citrulline) and iNOS expression. Toxin B increased the activity of iNOS promoter-reporter construct in VSMCs. Both toxins enhanced IL-1beta-stimulated iNOS expression and NO production. These data demonstrate for the first time that inhibition of Rho induces iNOS and suggest a role for Rho protein in IL-1beta-stimulated NO production in VSMCs.

  17. Tyrosine glycosylation of Rho by Yersinia toxin impairs blastomere cell behaviour in zebrafish embryos

    PubMed Central

    Jank, Thomas; Eckerle, Stephanie; Steinemann, Marcus; Trillhaase, Christoph; Schimpl, Marianne; Wiese, Sebastian; van Aalten, Daan M. F.; Driever, Wolfgang; Aktories, Klaus

    2015-01-01

    Yersinia species cause zoonotic infections, including enterocolitis and plague. Here we studied Yersinia ruckeri antifeeding prophage 18 (Afp18), the toxin component of the phage tail-derived protein translocation system Afp, which causes enteric redmouth disease in salmonid fish species. Here we show that microinjection of the glycosyltransferase domain Afp18G into zebrafish embryos blocks cytokinesis, actin-dependent motility and cell blebbing, eventually abrogating gastrulation. In zebrafish ZF4 cells, Afp18G depolymerizes actin stress fibres by mono-O-GlcNAcylation of RhoA at tyrosine-34; thereby Afp18G inhibits RhoA activation by guanine nucleotide exchange factors, and blocks RhoA, but not Rac and Cdc42 downstream signalling. The crystal structure of tyrosine-GlcNAcylated RhoA reveals an open conformation of the effector loop distinct from recently described structures of GDP- or GTP-bound RhoA. Unravelling of the molecular mechanism of the toxin component Afp18 as glycosyltransferase opens new perspectives in studies of phage tail-derived protein translocation systems, which are preserved from archaea to human pathogenic prokaryotes. PMID:26190758

  18. Bacterial factors exploit eukaryotic Rho GTPase signaling cascades to promote invasion and proliferation within their host

    PubMed Central

    Popoff, Michel R

    2014-01-01

    Actin cytoskeleton is a main target of many bacterial pathogens. Among the multiple regulation steps of the actin cytoskeleton, bacterial factors interact preferentially with RhoGTPases. Pathogens secrete either toxins which diffuse in the surrounding environment, or directly inject virulence factors into target cells. Bacterial toxins, which interfere with RhoGTPases, and to some extent with RasGTPases, catalyze a covalent modification (ADPribosylation, glucosylation, deamidation, adenylation, proteolysis) blocking these molecules in their active or inactive state, resulting in alteration of epithelial and/or endothelial barriers, which contributes to dissemination of bacteria in the host. Injected bacterial virulence factors preferentially manipulate the RhoGTPase signaling cascade by mimicry of eukaryotic regulatory proteins leading to local actin cytoskeleton rearrangement, which mediates bacterial entry into host cells or in contrast escape to phagocytosis and immune defense. Invasive bacteria can also manipulate RhoGTPase signaling through recognition and stimulation of cell surface receptor(s). Changes in RhoGTPase activation state is sensed by the innate immunity pathways and allows the host cell to adapt an appropriate defense response. PMID:25203748

  19. Cancer Stem Cells and Radioresistance: Rho/ROCK Pathway Plea Attention

    PubMed Central

    Pranatharthi, Annapurna; Ross, Cecil

    2016-01-01

    Radiation is the most potent mode of cancer therapy; however, resistance to radiation therapy results in tumor relapse and subsequent fatality. The cancer stem cell (CSC), which has better DNA repair capability, has been shown to contribute to tumor resistance and is an important target for treatment. Signaling molecules such as Notch, Wnt, and DNA repair pathways regulate molecular mechanisms in CSCs; however, none of them have been translated into therapeutic targets. The RhoGTPases and their effector ROCK-signaling pathway, though important for tumor progression, have not been well studied in the context of radioresistance. There are reports that implicate RhoA in radioresistance. ROCK2 has also been shown to interact with BRCA2 in the regulation of cell division. Incidentally, statins (drug for cardiovascular ailment) are functional inhibitors of RhoGTPases. Studies suggest that patients on statins have a better prognosis in cancers. Data from our lab suggest that ROCK signaling regulates radioresistance in cervical cancer cells. Collectively, these findings suggest that Rho/ROCK signaling may be important for radiation resistance. In this review, we enumerate the role of Rho/ROCK signaling in stemness and radioresistance and highlight the need to explore these molecules for a better understanding of radioresistance and development of therapeutics. PMID:27597870

  20. Measurements of branching fractions and CP-violating asymmetries in B0-->rho(+/-)h(-/+) decays.

    PubMed

    Aubert, B; Barate, R; Boutigny, D; Gaillard, J-M; Hicheur, A; Karyotakis, Y; Lees, J P; Robbe, P; Tisserand, V; Zghiche, A; Palano, A; Pompili, A; Chen, J C; Qi, N D; Rong, G; Wang, P; Zhu, Y S; Eigen, G; Ofte, I; Stugu, B; Abrams, G S; Borgland, A W; Breon, A B; Brown, D N; Button-Shafer, J; Cahn, R N; Charles, E; Day, C T; Gill, M S; Gritsan, A V; Groysman, Y; Jacobsen, R G; Kadel, R W; Kadyk, J; Kerth, L T; Kolomensky, Yu G; Kral, J F; Kukartsev, G; LeClerc, C; Levi, M E; Lynch, G; Mir, L M; Oddone, P J; Orimoto, T J; Pripstein, M; Roe, N A; Romosan, A; Ronan, M T; Shelkov, V G; Telnov, A V; Wenzel, W A; Ford, K; Harrison, T J; Hawkes, C M; Knowles, D J; Morgan, S E; Penny, R C; Watson, A T; Watson, N K; Deppermann, T; Goetzen, K; Koch, H; Lewandowski, B; Pelizaeus, M; Peters, K; Schmuecker, H; Steinke, M; Barlow, N R; Boyd, J T; Chevalier, N; Cottingham, W N; Kelly, M P; Latham, T E; Mackay, C; Wilson, F F; Abe, K; Cuhadar-Donszelmann, T; Hearty, C; Mattison, T S; McKenna, J A; Thiessen, D; Kyberd, P; McKemey, A K; Blinov, V E; Bukin, A D; Golubev, V B; Ivanchenko, V N; Kravchenko, E A; Onuchin, A P; Serednyakov, S I; Skovpen, Yu I; Solodov, E P; Yushkov, A N; Best, D; Chao, M; Kirkby, D; Lankford, A J; Mandelkern, M; McMahon, S; Mommsen, R K; Roethel, W; Stoker, D P; Buchanan, C; del Re, D; Hadavand, H K; Hill, E J; MacFarlane, D B; Paar, H P; Rahatlou, Sh; Schwanke, U; Sharma, V; Berryhill, J W; Campagnari, C; Dahmes, B; Kuznetsova, N; Levy, S L; Long, O; Lu, A; Mazur, M A; Richman, J D; Verkerke, W; Beck, T W; Beringer, J; Eisner, A M; Heusch, C A; Lockman, W S; Schalk, T; Schmitz, R E; Schumm, B A; Seiden, A; Turri, M; Walkowiak, W; Williams, D C; Wilson, M G; Albert, J; Chen, E; Dubois-Felsmann, G P; Dvoretskii, A; Hitlin, D G; Narsky, I; Porter, F C; Ryd, A; Samuel, A; Yang, S; Jayatilleke, S; Mancinelli, G; Meadows, B T; Sokoloff, M D; Abe, T; Barillari, T; Blanc, F; Bloom, P; Clark, P J; Ford, W T; Nauenberg, U; Olivas, A; Rankin, P; Roy, J; Smith, J G; van Hoek, W C; Zhang, L; Harton, J L; Hu, T; Soffer, A; Toki, W H; Wilson, R J; Zhang, J; Altenburg, D; Brandt, T; Brose, J; Colberg, T; Dickopp, M; Dubitzky, R S; Hauke, A; Lacker, H M; Maly, E; Müller-Pfefferkorn, R; Nogowski, R; Otto, S; Schubert, K R; Schwierz, R; Spaan, B; Wilden, L; Bernard, D; Bonneaud, G R; Brochard, F; Cohen-Tanugi, J; Thiebaux, Ch; Vasileiadis, G; Verderi, M; Khan, A; Lavin, D; Muheim, F; Playfer, S; Swain, J E; Tinslay, J; Andreotti, M; Bettoni, D; Bozzi, C; Calabrese, R; Cibinetto, G; Luppi, E; Negrini, M; Piemontese, L; Sarti, A; Treadwell, E; Anulli, F; Baldini-Ferroli, R; Calcaterra, A; de Sangro, R; Falciai, D; Finocchiaro, G; Patteri, P; Peruzzi, I M; Piccolo, M; Zallo, A; Buzzo, A; Contri, R; Crosetti, G; Lo Vetere, M; Macri, M; Monge, M R; Passaggio, S; Pastore, F C; Patrignani, C; Robutti, E; Santroni, A; Tosi, S; Bailey, S; Morii, M; Aspinwall, M L; Bhimji, W; Bowerman, D A; Dauncey, P D; Egede, U; Eschrich, I; Morton, G W; Nash, J A; Sanders, P; Taylor, G P; Grenier, G J; Lee, S-J; Mallik, U; Cochran, J; Crawley, H B; Lamsa, J; Meyer, W T; Prell, S; Rosenberg, E I; Yi, J; Davier, M; Grosdidier, G; Höcker, A; Laplace, S; Le Diberder, F; Lepeltier, V; Lutz, A M; Petersen, T C; Plaszczynski, S; Schune, M H; Tantot, L; Wormser, G; Brigljević, V; Cheng, C H; Lange, D J; Wright, D M; Bevan, A J; Coleman, J P; Fry, J R; Gabathuler, E; Gamet, R; Kay, M; Parry, R J; Payne, D J; Sloane, R J; Touramanis, C; Back, J J; Harrison, P F; Shorthouse, H W; Strother, P; Vidal, P B; Brown, C L; Cowan, G; Flack, R L; Flaecher, H U; George, S; Green, M G; Kurup, A; Marker, C E; McMahon, T R; Ricciardi, S; Salvatore, F; Vaitsas, G; Winter, M A; Brown, D; Davis, C L; Allison, J; Barlow, R J; Forti, A C; Hart, P A; Jackson, F; Lafferty, G D; Lyon, A J; Weatherall, J H; Williams, J C; Farbin, A; Jawahery, A; Kovalskyi, D; Lae, C K; Lillard, V; Roberts, D A; Blaylock, G; Dallapiccola, C; Flood, K T; Hertzbach, S S; Kofler, R; Koptchev, V B; Moore, T B; Saremi, S; Staengle, H; Willocq, S; Cowan, R; Sciolla, G; Taylor, F; Yamamoto, R K; Mangeol, D J J; Milek, M; Patel, P M; Lazzaro, A; Palombo, F; Bauer, J M; Cremaldi, L; Eschenburg, V; Godang, R; Kroeger, R; Reidy, J; Sanders, D A; Summers, D J; Zhao, H W; Hast, C; Taras, P; Nicholson, H; Cartaro, C; Cavallo, N; De Nardo, G; Fabozzi, F; Gatto, C; Lista, L; Paolucci, P; Piccolo, D; Sciacca, C; Baak, M A; Raven, G; LoSecco, J M; Gabriel, T A; Brau, B; Pulliam, T; Brau, J; Frey, R; Potter, C T; Sinev, N B; Strom, D; Torrence, E; Colecchia, F; Dorigo, A; Galeazzi, F; Margoni, M; Morandin, M; Posocco, M; Rotondo, M; Simonetto, F; Stroili, R; Tiozzo, G; Voci, C; Benayoun, M; Briand, H; Chauveau, J; David, P; de la Vaissière, Ch; Del Buono, L; Hamon, O; John, M J J; Leruste, Ph; Ocariz, J; Pivk, M; Roos, L; Stark, J; T'Jampens, S; Manfredi, P F; Re, V; Gladney, L; Guo, Q H; Panetta, J; Angelini, C; Batignani, G; Bettarini, S; Bondioli, M; Bucci, F; Calderini, G; Carpinelli, M; Forti, F; Giorgi, M A; Lusiani, A; Marchiori, G; Martinez-Vidal, F; Morganti, M; Neri, N; Paoloni, E; Rama, M; Rizzo, G; Sandrelli, F; Walsh, J; Haire, M; Judd, D; Paick, K; Wagoner, D E; Danielson, N; Elmer, P; Lu, C; Miftakov, V; Olsen, J; Smith, A J S; Varnes, E W; Bellini, F; Cavoto, G; Faccini, R; Ferrarotto, F; Ferroni, F; Gaspero, M; Mazzoni, M A; Morganti, S; Pierini, M; Piredda, G; Safai Tehrani, F; Voena, C; Christ, S; Wagner, G; Waldi, R; Adye, T; De Groot, N; Franek, B; Geddes, N I; Gopal, G P; Olaiya, E O; Xella, S M; Aleksan, R; Emery, S; Gaidot, A; Ganzhur, S F; Giraud, P-F; Hamel de Monchenault, G; Kozanecki, W; Langer, M; London, G W; Mayer, B; Schott, G; Vasseur, G; Yeche, Ch; Zito, M; Purohit, M V; Weidemann, A W; Yumiceva, F X; Aston, D; Bartoldus, R; Berger, N; Boyarski, A M; Buchmueller, O L; Convery, M R; Coupal, D P; Dong, D; Dorfan, J; Dujmic, D; Dunwoodie, W; Field, R C; Glanzman, T; Gowdy, S J; Grauges-Pous, E; Hadig, T; Halyo, V; Hryn'ova, T; Innes, W R; Jessop, C P; Kelsey, M H; Kim, P; Kocian, M L; Langenegger, U; Leith, D W G S; Luitz, S; Luth, V; Lynch, H L; Marsiske, H; Menke, S; Messner, R; Muller, D R; O'Grady, C P; Ozcan, V E; Perazzo, A; Perl, M; Petrak, S; Ratcliff, B N; Robertson, S H; Roodman, A; Salnikov, A A; Schindler, R H; Schwiening, J; Simi, G; Snyder, A; Soha, A; Stelzer, J; Su, D; Sullivan, M K; Tanaka, H A; Va'vra, J; Wagner, S R; Weaver, M; Weinstein, A J R; Wisniewski, W J; Wright, D H; Young, C C; Burchat, P R; Edwards, A J; Meyer, T I; Roat, C; Ahmed, S; Alam, M S; Ernst, J A; Saleem, M; Wappler, F R; Bugg, W; Krishnamurthy, M; Spanier, S M; Eckmann, R; Kim, H; Ritchie, J L; Schwitters, R F; Izen, J M; Kitayama, I; Lou, X C; Ye, S; Bianchi, F; Bona, M; Gallo, F; Gamba, D; Borean, C; Bosisio, L; Della Ricca, G; Dittongo, S; Grancagnolo, S; Lanceri, L; Poropat, P; Vitale, L; Vuagnin, G; Panvini, R S; Banerjee, Sw; Brown, C M; Fortin, D; Jackson, P D; Kowalewski, R; Roney, J M; Band, H R; Dasu, S; Datta, M; Eichenbaum, A M; Hu, H; Johnson, J R; Kutter, P E; Li, H; Liu, R; Di Lodovico, F; Mihalyi, A; Mohapatra, A K; Pan, Y; Prepost, R; Sekula, S J; von Wimmersperg-Toeller, J H; Wu, J; Wu, S L; Yu, Z; Neal, H

    2003-11-14

    We present measurements of branching fractions and CP-violating asymmetries in B0-->rho(+/-)pi(-/+) and B0-->rho-K+ decays. The results are obtained from a data sample of 88.9 x 10(6) Upsilon(4S)-->BB decays collected with the BABAR detector at the SLAC PEP-II asymmetric-energy B Factory. From a time-dependent maximum likelihood fit we measure the branching fractions B(B0-->rho(+/-)pi(-/+))=[22.6+/-1.8 (stat)+/-2.2 (syst)]x10(-6) and B(B0-->rho-K+)=(7.3 -1.2( +1.3)+/-1.3)x10(-6), and the CP-violating charge asymmetries A(rhopi)(CP)=-0.18+/-0.08+/-0.03 and A(rhoK)(CP)=0.28+/-0.17+/-0.08, the direct CP violation parameter C(rhopi)=0.36+/-0.18+/-0.04 and the mixing-induced CP violation parameter S(rhopi)=0.19+/-0.24+/-0.03, and the dilution parameters DeltaC(rhopi)=0.28 -0.19( +0.18)+/-0.04 and DeltaS(rhopi)=0.15+/-0.25+/-0.03.

  1. Role of citron kinase as a target of the small GTPase Rho in cytokinesis.

    PubMed

    Madaule, P; Eda, M; Watanabe, N; Fujisawa, K; Matsuoka, T; Bito, H; Ishizaki, T; Narumiya, S

    1998-07-30

    During mitosis, a ring containing actin and myosin appears beneath the equatorial surface of animal cells. This ring then contracts, forms a cleavage furrow and divides the cell, a step known as cytokinesis. The two daughter cells often remain connected by an intercellular bridge which contains a refringent structure known as the midbody. How the appearance of this ring is regulated is unclear, although the small GTPase Rho, which controls the formation of actin structures, is known to be essential. Protein kinases are also thought to participate in cytokinesis. We now show that a splice variant of a Rho target protein, named citron, contains a protein kinase domain that is related to the Rho-associated kinases ROCK14 and ROK, which regulate myosin-based contractility. Citron kinase localizes to the cleavage furrow and midbody of HeLa cells; Rho is also localized in the midbody. We find that overexpression of citron mutants results in the production of multinucleate cells and that a kinase-active mutant causes abnormal contraction during cytokinesis. We propose that citron kinase regulates cytokinesis at a step after Rho in the contractile process.

  2. CP asymmetries in B{yields}K{pi}, K*{pi}, {rho}K decays

    SciTech Connect

    Gronau, Michael; Pirjol, Dan; Zupan, Jure

    2010-05-01

    We show that ratios of tree and penguin amplitudes in B{yields}K*{pi} and B{yields}{rho}K are 2 to 3 times larger than in B{yields}K{pi}. This allows for considerably larger CP asymmetries in the former processes than the 10% asymmetry measured in B{sup 0{yields}}K{sup +{pi}-}. We study isospin sum rules for rate asymmetries in B{yields}K{pi}, K*{pi}, {rho}K, estimating small violation from interference of tree and electroweak penguin amplitudes. The breaking of the K{pi} asymmetry sum rule is estimated to be 1 to 2% and negative. Violation of K*{pi} and {rho}K sum rules can be estimated from B{yields}{rho}{pi} amplitudes using flavor SU(3), while breaking of a sum rule combining K*{pi} and {rho}K asymmetries can be measured directly in a Dalitz analysis of B{sup 0{yields}}K{sup +{pi}-{pi}0}. The three sum rules can be tested using complete sets of data taken at e{sup +}e{sup -} B factories and in experiments at the LHCb and at a future Super Flavor Factory, providing precision searches for new {Delta}S={Delta}I=1 operators in the low-energy effective Hamiltonian.

  3. Regulation of white and brown adipocyte differentiation by RhoGAP DLC1

    PubMed Central

    Brunmeir, Reinhard; Zhang, Qiongyi; Li, Hongyu; Dharmasegaran, Dharmini; Leong, Carol; Lim, Ying Yan; Han, Weiping

    2017-01-01

    Adipose tissues constitute an important component of metabolism, the dysfunction of which can cause obesity and type II diabetes. Here we show that differentiation of white and brown adipocytes requires Deleted in Liver Cancer 1 (DLC1), a Rho GTPase Activating Protein (RhoGAP) previously studied for its function in liver cancer. We identified Dlc1 as a super-enhancer associated gene in both white and brown adipocytes through analyzing the genome-wide binding profiles of PPARγ, the master regulator of adipogenesis. We further observed that Dlc1 expression increases during differentiation, and knockdown of Dlc1 by siRNA in white adipocytes reduces the formation of lipid droplets and the expression of fat marker genes. Moreover, knockdown of Dlc1 in brown adipocytes reduces expression of brown fat-specific genes and diminishes mitochondrial respiration. Dlc1-/- knockout mouse embryonic fibroblasts show a complete inability to differentiate into adipocytes, but this phenotype can be rescued by inhibitors of Rho-associated kinase (ROCK) and filamentous actin (F-actin), suggesting the involvement of Rho pathway in DLC1-regulated adipocyte differentiation. Furthermore, PPARγ binds to the promoter of Dlc1 gene to regulate its expression during both white and brown adipocyte differentiation. These results identify DLC1 as an activator of white and brown adipocyte differentiation, and provide a molecular link between PPARγ and Rho pathways. PMID:28358928

  4. Regulation of white and brown adipocyte differentiation by RhoGAP DLC1.

    PubMed

    Sim, Choon Kiat; Kim, Sun-Yee; Brunmeir, Reinhard; Zhang, Qiongyi; Li, Hongyu; Dharmasegaran, Dharmini; Leong, Carol; Lim, Ying Yan; Han, Weiping; Xu, Feng

    2017-01-01

    Adipose tissues constitute an important component of metabolism, the dysfunction of which can cause obesity and type II diabetes. Here we show that differentiation of white and brown adipocytes requires Deleted in Liver Cancer 1 (DLC1), a Rho GTPase Activating Protein (RhoGAP) previously studied for its function in liver cancer. We identified Dlc1 as a super-enhancer associated gene in both white and brown adipocytes through analyzing the genome-wide binding profiles of PPARγ, the master regulator of adipogenesis. We further observed that Dlc1 expression increases during differentiation, and knockdown of Dlc1 by siRNA in white adipocytes reduces the formation of lipid droplets and the expression of fat marker genes. Moreover, knockdown of Dlc1 in brown adipocytes reduces expression of brown fat-specific genes and diminishes mitochondrial respiration. Dlc1-/- knockout mouse embryonic fibroblasts show a complete inability to differentiate into adipocytes, but this phenotype can be rescued by inhibitors of Rho-associated kinase (ROCK) and filamentous actin (F-actin), suggesting the involvement of Rho pathway in DLC1-regulated adipocyte differentiation. Furthermore, PPARγ binds to the promoter of Dlc1 gene to regulate its expression during both white and brown adipocyte differentiation. These results identify DLC1 as an activator of white and brown adipocyte differentiation, and provide a molecular link between PPARγ and Rho pathways.

  5. Rho kinase regulates neurite outgrowth of hippocampal neurons via calcium dependent cytoskeleton regulation

    PubMed Central

    Ji, Zhisheng; Cai, Zhenbin; Zhang, Jifeng; Liu, Nannuan; Chen, Jing; Tan, Minghui; Lin, Hongsheng; Guo, Guoqing

    2017-01-01

    Objective: To investigate whether calcium is involved in downstream signal transduction in neurite outgrowth regulated by Rho kinase. Methods: In vitro primary hippocampal neurons were cultured and treated with Rho kinase agonist (LPA) or antagonist (Y-27632). Then, the cytoskeleton and neurite outgrowth were observed. After addition of calcium antagonist BAPTA/AM to reduce intracellular calcium, the cytoskeleton distribution and neurite outgrowth were observed. Results: The activation or inhibition of Rho kinase could significantly alter the number and length of neurites of hippocampal neurons. Rho kinase regulated the cytoskeleton to regulate the neurite outgrowth, and LPA could significantly increase intracellular calcium. After BAPTA/AM treatment, the length and branch number of neurites of neurons reduced markedly. BAPTA/AM was able to reduce intracellular calcium and decrease neuronal cytoskeleton. Treatment with both BAPTA/AM and LPA could stop the retraction of neurites, but the length and branch number of neurites remained unchanged after treatment with Y-27632 and LPA. Conclusion: Calcium may affect the cytoskeleton arrangement to regulate neurite outgrowth, and calcium is involved in the downstream signal transduction of Rho kinase regulated neurite outgrowth of hippocampal neurons. PMID:28337305

  6. Coaching the alpha male.

    PubMed

    Ludeman, Kate; Erlandson, Eddie

    2004-05-01

    Highly intelligent, confident, and successful, alpha males represent about 70% of all senior executives. Natural leaders, they willingly take on levels of responsibility most rational people would find overwhelming. But many of their quintessential strengths can also make alphas difficult to work with. Their self-confidence can appear domineering. Their high expectations can make them excessively critical. Their unemotional style can keep them from inspiring their teams. That's why alphas need coaching to broaden their interpersonal tool kits while preserving their strengths. Drawing from their experience coaching more than 1,000 senior executives, the authors outline an approach tailored specifically for the alpha. Coaches get the alpha's attention by inundating him with data from 360-degree feedback presented in ways he will find compelling--both hard-boiled metrics and vivid verbatim comments from colleagues about his strengths and weaknesses. A 360-degree assessment is a wake-up call for most alphas, providing undeniable proof that their behavior doesn't work nearly as well as they think it does. That paves the way for a genuine commitment to change. In order to change, the alpha must venture into unfamiliar--and often uncomfortable--psychological territory. He must admit vulnerability, accept accountability not just for his own work for others', connect with his underlying emotions, learn to motivate through a balance of criticism and validation, and become aware of unproductive behavior patterns. The goal of executive coaching is not simply to treat the alpha as an individual problem but to improve the entire team dynamic. Initial success creates an incentive to persevere, and the virtuous cycle reverberates throughout the entire organization.

  7. alpha2-Adrenoreceptor antagonists.

    PubMed

    Mayer, P; Imbert, T

    2001-06-01

    A review of the literature relating to the therapeutic potential of alpha2-adrenoceptor antagonists published between 1990 and 2000 is presented. Although extensively studied since the early 1970s in a wide spectrum of therapeutic applications, the distinction of alpha2-adrenoceptor subtypes and some emerging evidence concerning new applications in neurodegenerative disorders, such as Alzheimer's and Parkinson's diseases, obesity and schizophrenia, have refreshed an interest in this class of agents.

  8. Alpha Particle Diagnostic

    SciTech Connect

    Fisher, Ray, K.

    2009-05-13

    The study of burning plasmas is the next frontier in fusion energy research, and will be a major objective of the U.S. fusion program through U.S. collaboration with our international partners on the ITER Project. For DT magnetic fusion to be useful for energy production, it is essential that the energetic alpha particles produced by the fusion reactions be confined long enough to deposit a significant fraction of their initial ~3.5 MeV energy in the plasma before they are lost. Development of diagnostics to study the behavior of energetic confined alpha particles is a very important if not essential part of burning plasma research. Despite the clear need for these measurements, development of diagnostics to study confined the fast confined alphas to date has proven extremely difficult, and the available techniques remain for the most part unproven and with significant uncertainties. Research under this grant had the goal of developing diagnostics of fast confined alphas, primarily based on measurements of the neutron and ion tails resulting from alpha particle knock-on collisions with the plasma deuterium and tritium fuel ions. One of the strengths of this approach is the ability to measure the alphas in the hot plasma core where the interesting ignition physics will occur.

  9. Differential humoral response against alpha- and beta-linked mannose residues associated with tissue invasion by Candida albicans.

    PubMed Central

    Jouault, T; Delaunoy, C; Sendid, B; Ajana, F; Poulain, D

    1997-01-01

    Candida albicans mannan is the major cell wall antigen that elicits antibodies considered to be of little diagnostic value. It comprises epitopes corresponding to sequences of alpha- and beta-1,2-linked mannose residues. Both types of oligomannosidic epitopes may also be present on the glycosidic portions of other C. albicans molecules, i.e., mannoproteins (MP) (either structural or enzymatic) and glycolipids. The human humoral responses against beta-1,2- and alpha-linked oligomannosides were investigated by C. albicans Western blotting by considering the elective distribution of beta-1,2-oligomannosidic epitopes over a 14- to 18-kDa phospholipomannan (PLM) and the presence of alpha-mannosidic epitopes over heavily glycosylated MP. Western blotting of 51 control sera confirmed the presence of antibodies against C. albicans as a commensal member of the indigenous microflora; an immunoglobulin G (IgG) reactivity linked to enzyme-linked immunosorbent assay mannan signals was found for both PLM (beta-1,2-Man residues) and MP (alpha-Man residues). Despite strong reactivities against mannan and MP, IgG from 21 hospitalized patients with mycological evidence of deep-tissue invasion by C. albicans very significantly failed to react or reacted only faintly with PLM. This downregulation of anti-beta-1,2-oligomannosidic epitopes, associated with tissue invasion by C. albicans, was confirmed in 3 of 4 AIDS patients with extended oroesophageal candidosis. The application of a dissociation procedure proved that the absence of PLM reactivity was not due to the presence of immune complexes. These data provide the first evidence for a qualitative modification of the human antimannan antibody response associated with the C. albicans commensal-pathogenic transition. PMID:9144372

  10. Rho1 GTPase and PKC ortholog Pck1 are upstream activators of the cell integrity MAPK pathway in fission yeast.

    PubMed

    Sánchez-Mir, Laura; Soto, Teresa; Franco, Alejandro; Madrid, Marisa; Viana, Raúl A; Vicente, Jero; Gacto, Mariano; Pérez, Pilar; Cansado, José

    2014-01-01

    In the fission yeast Schizosaccharomyces pombe the cell integrity pathway (CIP) orchestrates multiple biological processes like cell wall maintenance and ionic homeostasis by fine tuning activation of MAPK Pmk1 in response to various environmental conditions. The small GTPase Rho2 positively regulates the CIP through protein kinase C ortholog Pck2. However, Pmk1 retains some function in mutants lacking either Rho2 or Pck2, suggesting the existence of additional upstream regulatory elements to modulate its activity depending on the nature of the environmental stimulus. The essential GTPase Rho1 is a candidate to control the activity of the CIP by acting upstream of Pck2, whereas Pck1, a second PKC ortholog, appears to negatively regulate Pmk1 activity. However, the exact regulatory nature of these two proteins within the CIP has remained elusive. By exhaustive characterization of strains expressing a hypomorphic Rho1 allele (rho1-596) in different genetic backgrounds we show that both Rho1 and Pck1 are positive upstream regulatory members of the CIP in addition to Rho2 and Pck2. In this new model Rho1 and Rho2 control Pmk1 basal activity during vegetative growth mainly through Pck2. Notably, whereas Rho2-Pck2 elicit Pmk1 activation in response to most environmental stimuli, Rho1 drives Pmk1 activation through either Pck2 or Pck1 exclusively in response to cell wall damage. Our study reveals the intricate and complex functional architecture of the upstream elements participating in this signaling pathway as compared to similar routes from other simple eukaryotic organisms.

  11. Transcription Elongation Factor NusA Is a General Antagonist of Rho-dependent Termination in Escherichia coli.

    PubMed

    Qayyum, M Zuhaib; Dey, Debashish; Sen, Ranjan

    2016-04-08

    NusA is an essential protein that binds to RNA polymerase and also to the nascent RNA and influences transcription by inducing pausing and facilitating the process of transcription termination/antitermination. Its participation in Rho-dependent transcription termination has been perceived, but the molecular nature of this involvement is not known. We hypothesized that, because both Rho and NusA are RNA-binding proteins and have the potential to target the same RNA, the latter is likely to influence the global pattern of the Rho-dependent termination. Analyses of the nascent RNA binding properties and consequent effects on the Rho-dependent termination functions of specific NusA-RNA binding domain mutants revealed an existence of Rho-NusA direct competition for the overlappingnut(NusA-binding site) andrut(Rho-binding site) sites on the RNA. This leads to delayed entry of Rho at therutsite that inhibits the latter's RNA release process. High density tiling microarray profiles of these NusA mutants revealed that a significant number of genes, together with transcripts from intergenic regions, are up-regulated. Interestingly, the majority of these genes were also up-regulated when the Rho function was compromised. These results provide strong evidence for the existence of NusA-binding sites in different operons that are also the targets of Rho-dependent terminations. Our data strongly argue in favor of a direct competition between NusA and Rho for the access of specific sites on the nascent transcripts in different parts of the genome. We propose that this competition enables NusA to function as a global antagonist of the Rho function, which is unlike its role as a facilitator of hairpin-dependent termination.

  12. Pyrin Inflammasome Activation and RhoA Signaling in the Autoinflammatory Diseases FMF and HIDS

    PubMed Central

    Park, Yong Hwan; Wood, Geryl; Kastner, Daniel L.; Chae, Jae Jin

    2016-01-01

    Mutations of pyrin and mevalonate kinase (MVK) cause distinct interleukin-1β (IL-1β)-mediated autoinflammatory diseases, familial Mediterranean fever (FMF) and hyperimmunoglobulinemia D syndrome (HIDS). Pyrin forms an inflammasome when mutated or in response to bacterial modification of the GTPase RhoA. Here we show that RhoA activates the serine-threonine kinases PKN1 and PKN2 that bind and phosphorylate pyrin. Phosphorylated pyrin binds 14-3-3 proteins, which block the pyrin inflammasome. The binding of 14-3-3 and PKN proteins to FMF-associated mutant pyrin is substantially decreased, and the constitutive IL-1β release from FMF or HIDS patients’ peripheral blood mononuclear cells is attenuated by activating PKN1 and PKN2. Defects in prenylation, seen in HIDS, lead to RhoA inactivation and consequent pyrin inflammasome activation. These data indicate a previously unsuspected fundamental molecular connection between two seemingly distinct autoinflammatory disorders. PMID:27270401

  13. Resveratrol suppresses breast cancer cell invasion by inactivating a RhoA/YAP signaling axis

    PubMed Central

    Kim, Yu Na; Choe, So Ra; Cho, Kyung Hwa; Cho, Do Yeun; Kang, Jaeku; Park, Chang Gyo; Lee, Hoi Young

    2017-01-01

    Hippo/YAP signaling is implicated in tumorigenesis and progression of various cancers. By inhibiting a plethora signaling cascades, resveratrol has strong anti-tumorigenic and anti-metastatic activity. In the present study, we demonstrate that resveratrol decreases the expression of YAP target genes. In addition, our data showed that resveratrol attenuates breast cancer cell invasion through the activation of Lats1 and consequent inactivation of YAP. Strikingly, we also demonstrate that resveratrol inactivates RhoA, leading to the activation of Lats1 and induction of YAP phosphorylation. Further, resveratrol in combination with other agents that inactivate RhoA or YAP showed more marked suppression of breast cancer cell invasion compared with single treatment. Collectively, these findings indicate the beneficial effects of resveratrol on breast cancer patients by suppressing the RhoA/Lats1/YAP signaling axis and subsequently inhibiting breast cancer cell invasion. PMID:28232662

  14. Induction of transient macroapertures in endothelial cells through RhoA inhibition by Staphylococcus aureus factors.

    PubMed

    Boyer, Laurent; Doye, Anne; Rolando, Monica; Flatau, Gilles; Munro, Patrick; Gounon, Pierre; Clément, René; Pulcini, Céline; Popoff, Michel R; Mettouchi, Amel; Landraud, Luce; Dussurget, Olivier; Lemichez, Emmanuel

    2006-06-05

    The GTPase RhoA is a major regulator of the assembly of actin stress fibers and the contractility of the actomyosin cytoskeleton. The epidermal cell differentiation inhibitor (EDIN) and EDIN-like ADP-ribosyltransferases of Staphylococcus aureus catalyze the inactivation of RhoA, producing actin cable disruption. We report that purified recombinant EDIN and EDIN-producing S. aureus provoke large transcellular tunnels in endothelial cells that we have named macroapertures (MAs). These structures open transiently, followed by the appearance of actin-containing membrane waves extending over the aperture. Disruption of actin cables, either directly or indirectly, through rhoA RNAi knockdown also triggers the formation of MAs. Intoxication of endothelial monolayers by EDIN produces a loss of barrier function and provides direct access of the endothelium basement membrane to S. aureus.

  15. New insights into Rho signaling from plant ROP/Rac GTPases.

    PubMed

    Craddock, Christian; Lavagi, Irene; Yang, Zhenbiao

    2012-09-01

    In animal and plant cells, a wide range of key cellular processes that require the establishment of cell polarity are governed by Rho-GTPases. In contrast to animals and yeast, however, plants possess a single Rho-GTPase subfamily called Rho-like GTPases from plants (ROPs). This raises the question of how plants achieve the high level of regulation required for polar cellular processes. It is becoming evident that plants have evolved specific regulators, including ROP-Guanine Exchange Factors (GEFs) and the Rop-interactive CRIB motif-containing protein (RIC) effectors. Recent research shows that the spatiotemporal dynamics of ROPs, the cytoskeleton, endocytosis, and exocytosis are intertwined. This review focuses on the proposed self-organizing nature of ROPs in plants and how ROP-mediated cellular mechanisms compare with those responsible for cell polarity in animals and yeast.

  16. Circadian Rhythms in Rho1 Activity Regulate Neuronal Plasticity and Network Hierarchy.

    PubMed

    Petsakou, Afroditi; Sapsis, Themistoklis P; Blau, Justin

    2015-08-13

    Neuronal plasticity helps animals learn from their environment. However, it is challenging to link specific changes in defined neurons to altered behavior. Here, we focus on circadian rhythms in the structure of the principal s-LNv clock neurons in Drosophila. By quantifying neuronal architecture, we observed that s-LNv structural plasticity changes the amount of axonal material in addition to cycles of fasciculation and defasciculation. We found that this is controlled by rhythmic Rho1 activity that retracts s-LNv axonal termini by increasing myosin phosphorylation and simultaneously changes the balance of pre-synaptic and dendritic markers. This plasticity is required to change clock network hierarchy and allow seasonal adaptation. Rhythms in Rho1 activity are controlled by clock-regulated transcription of Puratrophin-1-like (Pura), a Rho1 GEF. Since spinocerebellar ataxia is associated with mutations in human Puratrophin-1, our data support the idea that defective actin-related plasticity underlies this ataxia.

  17. A METHANE IMAGING SURVEY FOR T DWARF CANDIDATES IN {rho} OPHIUCHI

    SciTech Connect

    Haisch, Karl E.; Barsony, Mary; Tinney, Chris E-mail: mbarsony@stars.sfsu.ed

    2010-08-10

    We report on the results of the first deep, wide-field, near-infrared methane imaging survey of the {rho} Ophiuchi cloud core to search for T dwarfs. Among the 6587 objects detected, 22 were identified as T dwarf candidates. Brown dwarf models indicate that at the age and distance of the {rho} Ophiuchi cloud, these T dwarf candidates have masses between 1 and 2 Jupiter masses. If confirmed as genuine T dwarfs, these objects would be the youngest, lowest mass, and lowest gravity free-floating objects ever directly observed. The existence of these candidates suggests that the initial mass function of the {rho} Ophiuchi cloud extends well into the regime of planetary mass objects. A large fraction (59% {+-} 16%) of our T dwarf candidates appear to be surrounded by circumstellar disks, and thus represents the lowest mass objects yet found to harbor circumstellar disks.

  18. High Throughput Flow Cytometry Bead-based Multiplex Assay for Identification of Rho GTPase Inhibitors

    PubMed Central

    Surviladze, Zurab; Young, Susan M; Sklar, Larry A

    2015-01-01

    Summary Rho family GTPases and their effector proteins regulate a wide range of cell signaling pathways. In normal physiological conditions their activity is tightly controlled and it is not surprising that their aberrant activation contributes to tumorigenesis or other diseases. For this reason, the identification of small, cell permeable molecules capable of inhibition of Rho GTPases can be extraordinarily useful, particularly if they are specific and act reversibly. Herein we describe a flow cytometric assay, which allows us to measure the activity of six small GTPases simultaneously. GST-tagged small GTPases are bound to six glutathione bead sets each set having a different intensity of red fluorescence at a fixed wavelength. The coated bead sets were washed, combined, and dispensed into 384-well plates with test compounds, and fluorescent-GTP binding was used as the read-out. This multiplex bead-based assay was successfully used for to identify both general and selective inhibitors of Rho family GTPases. PMID:22144280

  19. Foci of cyclin A2 interact with actin and RhoA in mitosis

    PubMed Central

    Loukil, Abdelhalim; Izard, Fanny; Georgieva, Mariya; Mashayekhan, Shaereh; Blanchard, Jean-Marie; Parmeggiani, Andrea; Peter, Marion

    2016-01-01

    Cyclin A2 is a key player in the regulation of the cell cycle. Its degradation in mid-mitosis depends primarily on the ubiquitin-proteasome system (UPS), while autophagy also contributes. However, a fraction of cyclin A2 persists beyond metaphase. In this work, we focus on cyclin A2-rich foci detected in mitosis by high resolution imaging and analyse their movements. We demonstrate that cyclin A2 interacts with actin and RhoA during mitosis, and that cyclin A2 depletion induces a dramatic decrease in active RhoA in mitosis. Our data suggest cyclin A2 participation in RhoA activation in late mitosis. PMID:27279564

  20. Identifying the cm-wave continuum emitters in rho Oph W.

    NASA Astrophysics Data System (ADS)

    Casassus, Simon; Burton, Michael; White, Glenn; Dickinson, Clive; Paladini, Roberta; Cleary, Kieran

    2009-04-01

    The young stars in the rho Oph molecular cloud are not hot enough to form a conspicuous HII region. Only faint radiation from the Rayleigh-Jeans tail of ~10-100K dust is expected at wavelengths longwards of ~3mm. Yet we have found with the Cosmic Background Imager (CBI, 6arcmin uniform beam) that the rho Oph W photo-dissociation region (PDR) is surprisingly bright at cm wavelengths. The centimetric emission mechanism probably involves spinning dust, or electric dipole radiation from spinning very small dust grains (VSGs). Spinning dust holds the promise of a new window on VSG and PDR physics. ATCA can resolve the cm-wave continuum in rho Oph W and provide the first well-sampled 6cm - 0.7cm radio spectra and images of PDRs. Models of VSG emission both at cm-waves and in the infrared will bring constraints on physical conditions in the solid and gas states.

  1. The dynamics of Rho GTPase signaling and implications for targeting cancer and the tumor microenvironment

    PubMed Central

    Pajic, Marina; Herrmann, David; Vennin, Claire; Conway, James RW; Chin, Venessa T; Johnsson, Anna-Karin E; Welch, Heidi CE; Timpson, Paul

    2015-01-01

    Numerous large scale genomics studies have demonstrated that cancer is a molecularly heterogeneous disease, characterized by acquired changes in the structure and DNA sequence of tumor genomes. More recently, the role of the equally complex tumor microenvironment in driving the aggressiveness of this disease is increasingly being realized. Tumor cells are surrounded by activated stroma, creating a dynamic environment that promotes cancer development, metastasis and chemoresistance. The Rho family of small GTPases plays an essential role in the regulation of cell shape, cytokinesis, cell adhesion, and cell motility. Importantly, these processes need to be considered in the context of a complex 3-dimensional (3D) environment, with reciprocal feedback and cross-talk taking place between the tumor cells and host environment. Here we discuss the role of molecular networks involving Rho GTPases in cancer, and the therapeutic implications of inhibiting Rho signaling in both cancer cells and the emerging concept of targeting the surrounding stroma. PMID:26103062

  2. The Regulation of Cellular Responses to Mechanical Cues by Rho GTPases

    PubMed Central

    Hoon, Jing Ling; Tan, Mei Hua; Koh, Cheng-Gee

    2016-01-01

    The Rho GTPases regulate many cellular signaling cascades that modulate cell motility, migration, morphology and cell division. A large body of work has now delineated the biochemical cues and pathways, which stimulate the GTPases and their downstream effectors. However, cells also respond exquisitely to biophysical and mechanical cues such as stiffness and topography of the extracellular matrix that profoundly influence cell migration, proliferation and differentiation. As these cellular responses are mediated by the actin cytoskeleton, an involvement of Rho GTPases in the transduction of such cues is not unexpected. In this review, we discuss an emerging role of Rho GTPase proteins in the regulation of the responses elicited by biophysical and mechanical stimuli. PMID:27058559

  3. Evaluation of. nu. -bar/sub rho/ for /sup 239/Pu

    SciTech Connect

    Fort, E. ); Frehaut, J. ); Tellier, H. ); Long, P. )

    1988-08-01

    The average number of prompt neutrons ..nu..-bar/sub rho/ emitted per fission event has been evaluated for /sup 239/Pu with a special emphasis on the fluctuations experimentally observed in the low-energy range. These fluctuations have a significant impact on applications, especially the reactivity coefficient of advanced water reactors. Consequently, the ..nu..-bar/sub rho/ curve has to be defined in the same fine energy mesh as the fission cross section for accurate neutron source calculations. In this range, formalisms are proposed to calculate ..nu..-bar/sub rho/ from the resonance parameters, resolved or averaged. Using the JEF-1 library as a data base, an analysis of several thermal, low-moderated, or fast systems shows a good convergence of the selected microscopic and integral information.

  4. RhoGTPases as Key Players in Mammalian Cell Adaptation to Microgravity

    PubMed Central

    Deroanne, Christophe; Nusgens, Betty; Vico, Laurence; Guignandon, Alain

    2015-01-01

    A growing number of studies are revealing that cells reorganize their cytoskeleton when exposed to conditions of microgravity. Most, if not all, of the structural changes observed on flown cells can be explained by modulation of RhoGTPases, which are mechanosensitive switches responsible for cytoskeletal dynamics control. This review identifies general principles defining cell sensitivity to gravitational stresses. We discuss what is known about changes in cell shape, nucleus, and focal adhesions and try to establish the relationship with specific RhoGTPase activities. We conclude by considering the potential relevance of live imaging of RhoGTPase activity or cytoskeletal structures in order to enhance our understanding of cell adaptation to microgravity-related conditions. PMID:25649831

  5. RhoA Phosphorylation Induces Rac1 Release from Guanine Dissociation Inhibitor α and Stimulation of Vascular Smooth Muscle Cell Migration▿

    PubMed Central

    Rolli-Derkinderen, Malvyne; Toumaniantz, Gilles; Pacaud, Pierre; Loirand, Gervaise

    2010-01-01

    Although overactivation of RhoA is recognized as a common component of vascular disorders, the molecular mechanisms regulating RhoA activity in vascular smooth muscle cells (VSMC) are still unclear. We have previously shown that in VSMC, RhoA is phosphorylated on Ser188 by nitric oxide (NO)-stimulated cGMP-dependent kinase (PKG), which leads to RhoA-Rho kinase pathway inhibition. In this study, we showed that expression of phosphoresistant RhoA mutants prevented the stimulation of VSMC migration and adhesion induced by NO-PKG pathway activation. In contrast, under basal conditions, phosphomimetic RhoA mutants stimulated VSMC adhesion and migration through a signaling pathway requiring Rac1 and the Rho exchange factor Vav3. RhoA phosphorylation or phosphomimetic RhoA mutants induced Rac1 activation but did not activate Vav3. Indeed, phosphorylated RhoA or phosphomimetic mutants trapped guanine dissociation inhibitor α (GDIα), leading to the release of Rac1 and its translocation to the membrane, where it was then activated by the basal Vav3 nucleotide exchange activity. In vivo, RhoA phosphorylation induced by PKG activation in the aortas of rats treated with sildenafil induced dissociation of Rac1 from GDIα and activation of the Rac1 signaling pathway. These results suggest that the phosphorylation of RhoA represents a novel potent and physiological GDIα displacement factor that leads to Rac1 activation and regulation of Rac1-dependent VSMC functions. PMID:20696841

  6. The Bacterial Virulence Factor Lymphostatin Compromises Intestinal Epithelial Barrier Function by Modulating Rho GTPases

    PubMed Central

    Babbin, Brian A.; Sasaki, Maiko; Gerner-Schmidt, Kirsten W.; Nusrat, Asma; Klapproth, Jan-Michael A.

    2009-01-01

    Lymphocyte inhibitory factor A (lifA) in Citrobacter rodentium encodes the large toxin lymphostatin, which contains two enzymatic motifs associated with bacterial pathogenesis, a glucosyltransferase and a protease. Our aim was to determine the effects of each lymphostatin motif on intestinal epithelial-barrier function. In-frame mutations of C. rodentium lifA glucosyltransferase (CrGlM21) and protease (CrPrM5) were generated by homologous recombination. Infection of both model intestinal epithelial monolayers and mice with C. rodentium wild type resulted in compromised epithelial barrier function and mislocalization of key intercellular junction proteins in the tight junction and adherens junction. In contrast, CrGlM21 was impaired in its ability to reduce barrier function and influenced the tight junction proteins ZO-1 and occludin. CrPrM5 demonstrated decreased effects on the adherens junction proteins β-catenin and E-cadherin. Analysis of the mechanisms revealed that C. rodentium wild type differentially influenced Rho GTPase activation, suppressed Cdc42 activation, and induced Rho GTPase activation. CrGlM21 lost its suppressive effects on Cdc42 activation, whereas CrPrM5 was unable to activate Rho signaling. Rescue experiments using constitutively active Cdc42 or C3 exotoxin to inhibit Rho GTPase supported a role of Rho GTPases in the epithelial barrier compromise induced by C. rodentium. Taken together, our results suggest that lymphostatin is a bacterial virulence factor that contributes to the disruption of intestinal epithelial-barrier function via the modulation of Rho GTPase activities. PMID:19286565

  7. APCcdh1 Mediates Degradation of the Oncogenic Rho-GEF Ect2 after Mitosis

    PubMed Central

    Liot, Caroline; Seguin, Laetitia; Siret, Aurélie; Crouin, Catherine; Schmidt, Susanne; Bertoglio, Jacques

    2011-01-01

    Background Besides regulation of actin cytoskeleton-dependent functions, Rho GTPase pathways are essential to cell cycle progression and cell division. Rho, Rac and Cdc42 regulate G1 to S phase progression and are involved in cytokinesis. RhoA GDP/GTP cycling is required for normal cytokinesis and recent reports have shown that the exchange factor Ect2 and the GTPase activating protein MgcRacGAP regulate RhoA activity during mitosis. We previously showed that the transcription factors E2F1 and CUX1 regulate expression of MgcRacGAP and Ect2 as cells enter S-phase. Methodology/Principal Findings We now report that Ect2 is subject to proteasomal degradation after mitosis, following ubiquitination by the APC/C complex and its co-activator Cdh1. A proper nuclear localization of Ect2 is necessary for its degradation. APC-Cdh1 assembles K11-linked poly-ubiquitin chains on Ect2, depending upon a stretch of ∼25 amino acid residues that contain a bi-partite NLS, a conventional D-box and two TEK-like boxes. Site-directed mutagenesis of target sequences generated stabilized Ect2 proteins. Furthermore, such degradation-resistant mutants of Ect2 were found to activate RhoA and subsequent signalling pathways and are able to transform NIH3T3 cells. Conclusions/Significance Our results identify Ect2 as a bona fide cell cycle-regulated protein and suggest that its ubiquitination-dependent degradation may play an important role in RhoA regulation at the time of mitosis. Our findings raise the possibility that the overexpression of Ect2 that has been reported in some human tumors might result not only from deregulated transcription, but also from impaired degradation. PMID:21886810

  8. The assembly of integrin adhesion complexes requires both extracellular matrix and intracellular rho/rac GTPases

    PubMed Central

    1995-01-01

    Interaction of cells with extracellular matrix via integrin adhesion receptors plays an important role in a wide range of cellular: functions, for example cell growth, movement, and differentiation. Upon interaction with substrate, integrins cluster and associate with a variety of cytoplasmic proteins to form focal complexes and with the actin cytoskeleton. Although the intracellular signals induced by integrins are at present undefined, it is thought that they are mediated by proteins recruited to the focal complexes. It has been suggested, for example, that after recruitment to focal adhesions p125FAK can activate the ERK1/2 MAP kinase cascade. We have previously reported that members of the rho family of small GTPases can trigger the assembly of focal complexes when activated in cells. Using microinjection techniques, we have now examined the role of the extracellular matrix and of the two GTP-binding proteins, rac and rho, in the assembly of integrin complexes in both mouse and human fibroblasts. We find that the interaction of integrins with extracellular matrix alone is not sufficient to induce integrin clustering and focal complex formation. Similarly, activation of rho or rac by extracellular growth factors does not lead to focal complex formation in the absence of matrix. Focal complexes are only assembled in the presence of both matrix and functionally active members of the rho family. In agreement with this, the interaction of integrins with matrix in the absence of rho/rac activity is unable to activate the ERK1/2 kinases in Swiss 3T3 cells. In fact, ERK1/2 can be activated fully by growth factors in the absence of matrix and it seems unlikely, therefore, that the adhesion dependence of fibroblast growth is mediated through the ras/MAP kinase pathway. We conclude that extracellular matrix is not sufficient to trigger focal complex assembly and subsequent integrin-dependent signal transduction in the absence of functionally active members of the rho

  9. Aerobic exercise regulates Rho/cofilin pathways to rescue synaptic loss in aged rats

    PubMed Central

    Li, Yan; Zhao, Li; Gu, Boya; Cai, Jiajia; Lv, Yuanyuan; Yu, Laikang

    2017-01-01

    Purpose The role of exercise to prevent or reverse aging-induced cognitive decline has been widely reported. This neuroprotection is associated with changes in the synaptic structure plasticity. However, the mechanisms of exercise-induced synaptic plasticity in the aging brain are still unclear. Thus, the aim of the present study is to investigate the aging-related alterations of Rho-GTPase and the modulatory influences of exercise training. Methods Young and old rats were used in this study. Old rats were subjected to different schedules of aerobic exercise (12 m/min, 60 min/d, 3d/w or 5d/w) or kept sedentary for 12 w. After 12 w of aerobic exercise, the synapse density in the cortex and hippocampus was detected with immunofluorescent staining using synaptophysin as a marker. The total protein levels of RhoA, Rac1, Cdc42 and cofilin in the cortex and hippocampus were detected with Western Blot. The activities of RhoA, Rac1 and Cdc42 were determined using a pull down assay. Results We found that synapse loss occurred in aging rats. However, the change of expression and activity of RhoA, Rac1 and Cdc42 was different in the cortex and hippocampus. In the cortex, the expression and activity of Rac1 and Cdc42 was greatly increased with aging, whereas there were no changes in the expression and activity of RhoA. In the hippocampus, the expression and activity of Rac1 and Cdc42 was greatly decreased and there were no changes in the expression and activity of RhoA. As a major downstream substrate of the Rho GTPase family, the increased expression of cofilin was only observed in the cortex. High frequency exercise ameliorated all aging-related changes in the cortex and hippocampus. Conclusions These data suggest that aerobic exercise reverses synapse loss in the cortex and hippocampus in aging rats, which might be related to the regulation of Rho GTPases. PMID:28152068

  10. Alpha irradiation modeling

    SciTech Connect

    Keeton, S C; Mount, M E

    1999-03-26

    With the end of the Cold War and the associated limitations imposed on the nuclear weapons stockpile by strategic arms treaties, much has changed in the stockpile stewardship program. Weapons that were originally designed for stockpile lives on the order of 15 to 20 years are now being evaluated for much longer periods: in some cases as much as 60 years. As such, issues that were once considered to be of no consequence are being reexamined. Among these is the extent of the radiation dose received by secondary organics over time that results from the intrinsic alpha source of the weapon components. This report describes the results of work performed to estimate the alpha radiation deposition in the organic components of an LLNL system at specific points in its stockpile life. Included are discussions of the development of the intrinsic time- and energy-dependent alpha source term per unit mass, estimation of the effective source and absorber material thicknesses, development of a simplified model for the total intrinsic alpha source term and energy deposition in the absorber, and the alpha radiation deposition in the organic components of a selected LLNL weapon.

  11. Clostridium perfringens TpeL Induces Formation of Stress Fibers via Activation of RhoA-ROCK Signaling Pathway.

    PubMed

    Nagahama, Masahiro; Ohkubo, Akiko; Kinouchi, Yoshihito; Kobayashi, Keiko; Miyamoto, Kazuaki; Takehara, Masaya; Sakurai, Jun

    2015-01-01

    Clostridium perfringens TpeL belongs to a family of large clostridial glucosylating cytotoxins. TpeL modifies Rac1 and Ras subfamily proteins. Herein we report TpeL-induced formation of stress fibers via RhoA-Rho kinase (ROCK) signaling. A recombinant protein (TpeL1-525) derived from the TpeL N-terminal catalytic domain in the presence of streptolysin O (SLO) induced the formation of actin stress fibers in Madin-Darby canine kidney (MDCK) cells in a dose-dependent manner. The RhoA/ROCK pathway is known to control the formation of stress fibers. We examined the role of the RhoA/ROCK pathway in TpeL-induced formation of stress fibers. TpeL1-525-induced formation of stress fibers was inhibited by the ROCK inhibitor, Y27632 and Rho protein inhibitor, C3 transferase. TpeL1-525 activated RhoA and ROCK in a dose-dependent manner. C3 transferase blocked TpeL1-525-induced activation of RhoA and ROCK whereas Y27632 inhibited TpeL-induced activation of ROCK. These results demonstrate for the first time that TpeL induces the formation of stress fibers by activating the RhoA/ROCK signaling pathway.

  12. A High-Throughput Assay for Rho Guanine Nucleotide Exchange Factors Based on the Transcreener GDP Assay.

    PubMed

    Reichman, Melvin; Schabdach, Amanda; Kumar, Meera; Zielinski, Tom; Donover, Preston S; Laury-Kleintop, Lisa D; Lowery, Robert G

    2015-12-01

    Ras homologous (Rho) family GTPases act as molecular switches controlling cell growth, movement, and gene expression by cycling between inactive guanosine diphosphate (GDP)- and active guanosine triphosphate (GTP)-bound conformations. Guanine nucleotide exchange factors (GEFs) positively regulate Rho GTPases by accelerating GDP dissociation to allow formation of the active, GTP-bound complex. Rho proteins are directly involved in cancer pathways, especially cell migration and invasion, and inhibiting GEFs holds potential as a therapeutic strategy to diminish Rho-dependent oncogenesis. Methods for measuring GEF activity suitable for high-throughput screening (HTS) are limited. We developed a simple, generic biochemical assay method for measuring GEF activity based on the fact that GDP dissociation is generally the rate-limiting step in the Rho GTPase catalytic cycle, and thus addition of a GEF causes an increase in steady-state GTPase activity. We used the Transcreener GDP Assay, which relies on selective immunodetection of GDP, to measure the GEF-dependent stimulation of steady-state GTP hydrolysis by small GTPases using Dbs (Dbl's big sister) as a GEF for Cdc42, RhoA, and RhoB. The assay is well suited for HTS, with a homogenous format and far red fluorescence polarization (FP) readout, and it should be broadly applicable to diverse Rho GEF/GTPase pairs.

  13. The role of the RhoA/ROCK pathway in gender-dependent differences in gastric smooth muscle contraction.

    PubMed

    Al-Shboul, Othman

    2016-01-01

    Gender-related differences in various gastric functions and diseases have been reported, with women having a higher prevalence of gastrointestinal disturbances than men. The aim of this study was to investigate sex-dependent differences in activation of the Rho-associated protein kinase (ROCK; RhoA/Rho kinase) pathway and muscle contraction in the stomach using single gastric smooth muscle cells (GSMC) from male and female Sprague-Dawley rats. Expression of ROCK1 and ROCK2 protein and acetylcholine (ACh)-induced activation of RhoA and ROCK were measured using a specifically designed enzyme-linked immunosorbent assay and activity assay kits, respectively. Contraction of a single GSMC was measured by scanning micrometry in the presence or absence of the ROCK inhibitor Y27632 dihydrochloride. ACh-induced activation of RhoA and ROCK and subsequent contraction were greater in male rats than in female rats but neither was related to differences in the expression of ROCK1 or ROCK2 or total RhoA amount. Most important, Y27632 inhibited and abolished differences in ACh-induced contraction in both sexes. In conclusion, increased ACh-induced contraction in the GSMC of male rats is attributable to greater RhoA/ROCK activation independent of differences in the expression of ROCK isoforms or total RhoA.

  14. Viral vector-mediated downregulation of RhoA increases survival and axonal regeneration of retinal ganglion cells

    PubMed Central

    Koch, Jan Christoph; Tönges, Lars; Michel, Uwe; Bähr, Mathias; Lingor, Paul

    2014-01-01

    The Rho/ROCK pathway is a promising therapeutic target in neurodegenerative and neurotraumatic diseases. Pharmacological inhibition of various pathway members has been shown to promote neuronal regeneration and survival. However, because pharmacological inhibitors are inherently limited in their specificity, shRNA-mediated approaches can add more information on the function of each single kinase involved. Thus, we generated adeno-associated viral vectors (AAV) to specifically downregulate Ras homologous member A (RhoA) via shRNA. We found that specific knockdown of RhoA promoted neurite outgrowth of retinal ganglion cells (RGC) grown on the inhibitory substrate chondroitin sulfate proteoglycan (CSPG) as well as neurite regeneration of primary midbrain neurons (PMN) after scratch lesion. In the rat optic nerve crush (ONC) model in vivo, downregulation of RhoA significantly enhanced axonal regeneration compared to control. Moreover, survival of RGC transduced with AAV expressing RhoA-shRNA was substantially increased at 2 weeks after optic nerve axotomy. Compared to previous data using pharmacological inhibitors to target RhoA, its upstream regulator Nogo or its main downstream target ROCK, the specific effects of RhoA downregulation shown here were most pronounced in regard to promoting RGC survival but neurite outgrowth and axonal regeneration were also increased significantly. Taken together, we show here that specific knockdown of RhoA substantially increases neuronal survival after optic nerve axotomy and modestly increases neurite outgrowth in vitro and axonal regeneration after optic nerve crush. PMID:25249936

  15. TGF beta-mediated RhoA expression is necessary for epithelial-mesenchymal transition in the embryonic chick heart.

    PubMed

    Tavares, André Luiz P; Mercado-Pimentel, Melania E; Runyan, Raymond B; Kitten, Gregory T

    2006-06-01

    Endothelia in the atrioventricular canal (AVC) of the embryonic heart undergo an epithelial-mesenchymal transition (EMT) and migrate into the underlying extracellular matrix. We explore here whether RhoA mediates this EMT. RhoA was detected in all cells of the chick heart during the stages studied. Expression was elevated when EMT was actively occurring. Explants treated with C3 exoenzyme in collagen gel cultures showed a significant decrease in mesenchymal cell numbers. siRNA was used to inhibit RhoA mRNA, and both activated endothelial and mesenchymal cells decreased significantly with treatment. Loss of RhoA produced a reduction of RhoB, cyclin-b2, and beta-catenin messages showing that these genes are regulated downstream of RhoA. In contrast, runx-2 was not reduced. Inhibition of TGFbeta3 or TGFbeta2 activity caused a large reduction of RhoA message. These data place RhoA in TGFbeta regulated pathways for both endothelial activation and mesenchymal invasion and demonstrate a functional requirement during EMT.

  16. Identification of Dck1 and Lmo1 as upstream regulators of the small GTPase Rho5 in Saccharomyces cerevisiae.

    PubMed

    Schmitz, Hans-Peter; Jendretzki, Arne; Wittland, Janina; Wiechert, Johanna; Heinisch, Jürgen J

    2015-04-01

    The exact function and regulation of the small GTPase Rho5, a putative homolog of mammalian Rac1, in the yeast Saccharomyces cerevisiae have not yet been elucidated. In a genetic screen initially designed to identify novel regulators of cell wall integrity signaling, we identified the homologs of mammalian DOCK1 (Dck1) and ELMO (Lmo1) as upstream components which regulate Rho5. Deletion mutants in any of the encoding genes (DCK1, LMO1, RHO5) showed hyper-resistance to cell wall stress agents, demonstrating a function in cell wall integrity signaling. Live-cell fluorescence microscopy showed that Dck1, Lmo1 and Rho5 quickly relocate to mitochondria under oxidative stress and cell viability assays indicate a role of Dck1/Lmo1/Rho5 signaling in triggering cell death as a response to hydrogen peroxide treatment. A regulatory role in autophagy/mitophagy is suggested by the colocalization of Rho5 with autophagic markers and the decreased mitochondrial turnover observed in dck1, lmo1 and rho5 deletion mutants. Rho5 activation may thus serve as a central hub for the integration of different signaling pathways.

  17. A Point Mutation in p190A RhoGAP Affects Ciliogenesis and Leads to Glomerulocystic Kidney Defects

    PubMed Central

    Shafer, Maxwell E. R.; Aoudjit, Lamine; Hu, Di; Sharma, Richa; Tremblay, Mathieu; Ishii, Hidetaka; Marcotte, Michael; Stanga, Daniela; Tang, You Chi; Boualia, Sami Kamel; Nguyen, Alana H. T.; Takano, Tomoko; Lamarche-Vane, Nathalie; Vidal, Silvia; Bouchard, Maxime

    2016-01-01

    Rho family GTPases act as molecular switches regulating actin cytoskeleton dynamics. Attenuation of their signaling capacity is provided by GTPase-activating proteins (GAPs), including p190A, that promote the intrinsic GTPase activity of Rho proteins. In the current study we have performed a small-scale ENU mutagenesis screen and identified a novel loss of function allele of the p190A gene Arhgap35, which introduces a Leu1396 to Gln substitution in the GAP domain. This results in decreased GAP activity for the prototypical Rho-family members, RhoA and Rac1, likely due to disrupted ordering of the Rho binding surface. Consequently, Arhgap35-deficient animals exhibit hypoplastic and glomerulocystic kidneys. Investigation into the cystic phenotype shows that p190A is required for appropriate primary cilium formation in renal nephrons. P190A specifically localizes to the base of the cilia to permit axoneme elongation, which requires a functional GAP domain. Pharmacological manipulations further reveal that inhibition of either Rho kinase (ROCK) or F-actin polymerization is able to rescue the ciliogenesis defects observed upon loss of p190A activity. We propose a model in which p190A acts as a modulator of Rho GTPases in a localized area around the cilia to permit the dynamic actin rearrangement required for cilia elongation. Together, our results establish an unexpected link between Rho GTPase regulation, ciliogenesis and glomerulocystic kidney disease. PMID:26859289

  18. GPR116, an adhesion G-protein-coupled receptor, promotes breast cancer metastasis via the Gαq-p63RhoGEF-Rho GTPase pathway.

    PubMed

    Tang, Xiaolong; Jin, Rongrong; Qu, Guojun; Wang, Xiu; Li, Zhenxi; Yuan, Zengjin; Zhao, Chen; Siwko, Stefan; Shi, Tieliu; Wang, Ping; Xiao, Jianru; Liu, Mingyao; Luo, Jian

    2013-10-15

    Adhesion G-protein-coupled receptors (GPCR), which contain adhesion domains in their extracellular region, have been found to play important roles in cell adhesion, motility, embryonic development, and immune response. Because most adhesion molecules with adhesion domains have vital roles in cancer metastasis, we speculated that adhesion GPCRs are potentially involved in cancer metastasis. In this study, we identified GPR116 as a novel regulator of breast cancer metastasis through expression and functional screening of the adhesion GPCR family. We found that knockdown of GPR116 in highly metastatic (MDA-MB-231) breast cancer cells suppressed cell migration and invasion. Conversely, ectopic GPR116 expression in poorly metastatic (MCF-7 and Hs578T) cells promoted cell invasion. We further showed that knockdown of GPR116 inhibited breast cancer cell metastasis in two mammary tumor metastasis mouse models. Moreover, GPR116 modulated the formation of lamellipodia and actin stress fibers in cells in a RhoA- and Rac1-dependent manner. At a molecular level, GPR116 regulated cell motility and morphology through the Gαq-p63RhoGEF-RhoA/Rac1 pathway. The biologic significance of GPR116 in breast cancer is substantiated in human patient samples, where GPR116 expression is significantly correlated with breast tumor progression, recurrence, and poor prognosis. These findings show that GPR116 is crucial for the metastasis of breast cancer and support GPR116 as a potential prognostic marker and drug target against metastatic human breast cancer.

  19. ALPHA MIS: Reference manual

    SciTech Connect

    Lovin, J.K.; Haese, R.L.; Heatherly, R.D.; Hughes, S.E.; Ishee, J.S.; Pratt, S.M.; Smith, D.W.

    1992-02-01

    ALPHA is a powerful and versatile management information system (MIS) initiated and sponsored and by the Finance and Business Management Division of Oak Ridge National Laboratory, who maintain and develop it in concert with the Business Systems Division for its Information Center. A general-purpose MIS, ALPHA allows users to access System 1022 and System 1032 databases to obtain and manage information. From a personal computer or a data terminal, Energy Systems employees can use ALPHA to control their own report reprocessing. Using four general commands (Database, Select, Sort, and Report) they can (1) choose a mainframe database, (2) define subsets within it, (3) sequentially order a subset by one or more variables, and (4) generate a report with their own or a canned format.

  20. AKAP-Lbc anchors protein kinase A and nucleates Galpha 12-selective Rho-mediated stress fiber formation.

    PubMed

    Diviani, D; Soderling, J; Scott, J D

    2001-11-23

    Guanine nucleotide exchange factors of the Dbl family relay signals from membrane receptors to Rho family GTPases. We now demonstrate that a longer transcript of the Lbc gene encodes a chimeric molecule, which we have called AKAP-Lbc, that functions as an A-kinase-anchoring protein (AKAP) and a Rho-selective guanine nucleotide exchange factor. Expression of AKAP-Lbc in fibroblasts favors the formation of stress fibers in a Rho-dependent manner. Application of lysophosphatidic acid or selective expression of Galpha(12) enhances cellular AKAP-Lbc activation. Furthermore, biochemical studies indicate that AKAP-Lbc functions as an adaptor protein to selectively couple Galpha(12) to Rho. Thus, AKAP-Lbc anchors PKA and nucleates the assembly of a Rho-mediated signaling pathway.

  1. Control of developmental networks by Rac/Rho small GTPases: How cytoskeletal changes during embryogenesis are orchestrated

    PubMed Central

    Sáenz‐Narciso, Beatriz; Gómez‐Orte, Eva; Zheleva, Angelina; Gastaca, Irene

    2016-01-01

    Small GTPases in the Rho family act as major nodes with functions beyond cytoskeletal rearrangements shaping the Caenorhabditis elegans embryo during development. These small GTPases are key signal transducers that integrate diverse developmental signals to produce a coordinated response in the cell. In C. elegans, the best studied members of these highly conserved Rho family small GTPases, RHO‐1/RhoA, CED‐10/Rac, and CDC‐42, are crucial in several cellular processes dealing with cytoskeletal reorganization. In this review, we update the functions described for the Rho family small GTPases in spindle orientation and cell division, engulfment, and cellular movements during C. elegans embryogenesis, focusing on the Rho subfamily Rac. Please also see the video abstract here PMID:27790724

  2. The Apollo Alpha Spectrometer.

    NASA Technical Reports Server (NTRS)

    Jagoda, N.; Kubierschky, K.; Frank, R.; Carroll, J.

    1973-01-01

    Located in the Science Instrument Module of Apollo 15 and 16, the Alpha Particle Spectrometer was designed to detect and measure the energy of alpha particles emitted by the radon isotopes and their daughter products. The spectrometer sensor consisted of an array of totally depleted silicon surface barrier detectors. Biased amplifier and linear gate techniques were utilized to reduce resolution degradation, thereby permitting the use of a single 512 channel PHA. Sensor identification and in-flight radioactive calibration were incorporated to enhance data reduction.

  3. Automated NMR fragment based screening identified a novel interface blocker to the LARG/RhoA complex.

    PubMed

    Gao, Jia; Ma, Rongsheng; Wang, Wei; Wang, Na; Sasaki, Ryan; Snyderman, David; Wu, Jihui; Ruan, Ke

    2014-01-01

    The small GTPase cycles between the inactive GDP form and the activated GTP form, catalyzed by the upstream guanine exchange factors. The modulation of such process by small molecules has been proven to be a fruitful route for therapeutic intervention to prevent the over-activation of the small GTPase. The fragment based approach emerging in the past decade has demonstrated its paramount potential in the discovery of inhibitors targeting such novel and challenging protein-protein interactions. The details regarding the procedure of NMR fragment screening from scratch have been rarely disclosed comprehensively, thus restricts its wider applications. To achieve a consistent screening applicable to a number of targets, we developed a highly automated protocol to cover every aspect of NMR fragment screening as possible, including the construction of small but diverse libray, determination of the aqueous solubility by NMR, grouping compounds with mutual dispersity to a cocktail, and the automated processing and visualization of the ligand based screening spectra. We exemplified our streamlined screening in RhoA alone and the complex of the small GTPase RhoA and its upstream guanine exchange factor LARG. Two hits were confirmed from the primary screening in cocktail and secondary screening over individual hits for LARG/RhoA complex, while one of them was also identified from the screening for RhoA alone. HSQC titration of the two hits over RhoA and LARG alone, respectively, identified one compound binding to RhoA.GDP at a 0.11 mM affinity, and perturbed the residues at the switch II region of RhoA. This hit blocked the formation of the LARG/RhoA complex, validated by the native gel electrophoresis, and the titration of RhoA to ¹⁵N labeled LARG in the absence and presence the compound, respectively. It therefore provides us a starting point toward a more potent inhibitor to RhoA activation catalyzed by LARG.

  4. RhoA and Rac1 GTPases Differentially Regulate Agonist-Receptor Mediated Reactive Oxygen Species Generation in Platelets

    PubMed Central

    Akbar, Huzoor; Duan, Xin; Saleem, Saima; Davis, Ashley K.; Zheng, Yi

    2016-01-01

    Agonist induced generation of reactive oxygen species (ROS) by NADPH oxidases (NOX) enhances platelet aggregation and hence the risk of thrombosis. RhoA and Rac1 GTPases are involved in ROS generation by NOX in a variety of cells, but their roles in platelet ROS production remain unclear. In this study we used platelets from RhoA and Rac1 conditional knockout mice as well as human platelets treated with Rhosin and NSC23767, rationally designed small molecule inhibitors of RhoA and Rac GTPases, respectively, to better define the contributions of RhoA and Rac1 signaling to ROS generation and platelet activation. Treatment of platelets with Rhosin inhibited: (a) U46619 induced activation of RhoA; (b) phosphorylation of p47phox, a critical component of NOX; (c) U46619 or thrombin induced ROS generation; (d) phosphorylation of myosin light chain (MLC); (e) platelet shape change; (f) platelet spreading on immobilized fibrinogen; and (g) release of P-selectin, secretion of ATP and aggregation. Conditional deletion of RhoA or Rac1 gene inhibited thrombin induced ROS generation in platelets. Addition of Y27632, a RhoA inhibitor, NSC23766 or Phox-I, an inhibitor of Rac1-p67phox interaction, to human platelets blocked thrombin induced ROS generation. These data suggest that: (a) RhoA/ROCK/p47phox signaling axis promotes ROS production that, at least in part, contributes to platelet activation in conjunction with or independent of the RhoA/ROCK mediated phosphorylation of MLC; and (b) RhoA and Rac1 differentially regulate ROS generation by inhibiting phosphorylation of p47phox and Rac1-p67phox interaction, respectively. PMID:27681226

  5. Automated NMR Fragment Based Screening Identified a Novel Interface Blocker to the LARG/RhoA Complex

    PubMed Central

    Gao, Jia; Ma, Rongsheng; Wang, Wei; Wang, Na; Sasaki, Ryan; Snyderman, David; Wu, Jihui; Ruan, Ke

    2014-01-01

    The small GTPase cycles between the inactive GDP form and the activated GTP form, catalyzed by the upstream guanine exchange factors. The modulation of such process by small molecules has been proven to be a fruitful route for therapeutic intervention to prevent the over-activation of the small GTPase. The fragment based approach emerging in the past decade has demonstrated its paramount potential in the discovery of inhibitors targeting such novel and challenging protein-protein interactions. The details regarding the procedure of NMR fragment screening from scratch have been rarely disclosed comprehensively, thus restricts its wider applications. To achieve a consistent screening applicable to a number of targets, we developed a highly automated protocol to cover every aspect of NMR fragment screening as possible, including the construction of small but diverse libray, determination of the aqueous solubility by NMR, grouping compounds with mutual dispersity to a cocktail, and the automated processing and visualization of the ligand based screening spectra. We exemplified our streamlined screening in RhoA alone and the complex of the small GTPase RhoA and its upstream guanine exchange factor LARG. Two hits were confirmed from the primary screening in cocktail and secondary screening over individual hits for LARG/RhoA complex, while one of them was also identified from the screening for RhoA alone. HSQC titration of the two hits over RhoA and LARG alone, respectively, identified one compound binding to RhoA.GDP at a 0.11 mM affinity, and perturbed the residues at the switch II region of RhoA. This hit blocked the formation of the LARG/RhoA complex, validated by the native gel electrophoresis, and the titration of RhoA to 15N labeled LARG in the absence and presence the compound, respectively. It therefore provides us a starting point toward a more potent inhibitor to RhoA activation catalyzed by LARG. PMID:24505392

  6. Rho kinase acts as a downstream molecule to participate in protein kinase Cε regulation of vascular reactivity after hemorrhagic shock in rats.

    PubMed

    Li, Tao; Zhu, Yu; Zang, Jia-tao; Peng, Xiao-yong; Lan, Dan; Yang, Guang-ming; Xu, Jing; Liu, Liang-ming

    2014-09-01

    Our previous study demonstrated that Rho kinase and protein kinase C (PKC) played important parts in the regulation of vascular reactivity after shock. Using superior mesenteric arteries (SMAs) from hemorrhagic shock rats and hypoxia-treated vascular smooth muscle cells (VSMCs), relationship of PKCε regulation of vascular reactivity to Rho kinase, as well as the signal transduction after shock, was investigated. The results showed that inhibition of Rho kinase with the Rho kinase-specific inhibitor Y-27632 antagonized the PKCε-specific agonist carbachol and highly expressed PKCε-induced increase of vascular reactivity in SMAs and VSMCs, whereas inhibition of PKCε with its specific inhibitory peptide did not antagonize the Rho kinase agonist (U-46619)-induced increase of vascular reactivity in SMAs and VSMCs. Activation of PKCε or highly expressed PKCε upregulated the activity of Rho kinase and the phosphorylation of PKC-dependent phosphatase inhibitor 17 (CPI-17), zipper interacting protein kinase (ZIPK), and integrin-linked kinase (ILK), whereas activation of Rho kinase increased only CPI-17 phosphorylation. The specific neutralization antibodies of ZIPK and ILK antagonized PKCε-induced increases in the activity of Rho kinase, but CPI-17 neutralization antibody did not antagonize this effect. These results suggested that Rho kinase takes part in the regulation of PKCε on vascular reactivity after shock. Rho kinase is downstream of PKCε. Protein kinase Cε activates Rho kinase via ZIPK and ILK; CPI-17 is downstream of Rho kinase.

  7. Table for Conversion of Kendall's Tau to Spearman's Rho within the Context of Measures of Magnitude of Effect for Meta-Analysis.

    ERIC Educational Resources Information Center

    Gilpin, Andrew R.

    1993-01-01

    Kendall's Tau is often considered equivalent to Spearman's Rho as an ordinal measure of correlation in spite of its different metric. Formulas for converting Tau to Rho are reviewed; and a table of corresponding values is presented for Tau, Rho, and several related indices. (SLD)

  8. KIF17 regulates RhoA-dependent actin remodeling at epithelial cell–cell adhesions

    PubMed Central

    Acharya, Bipul R.; Espenel, Cedric; Libanje, Fotine; Raingeaud, Joel; Morgan, Jessica; Jaulin, Fanny; Kreitzer, Geri

    2016-01-01

    ABSTRACT The kinesin KIF17 localizes at microtubule plus-ends where it contributes to regulation of microtubule stabilization and epithelial polarization. We now show that KIF17 localizes at cell–cell adhesions and that KIF17 depletion inhibits accumulation of actin at the apical pole of cells grown in 3D organotypic cultures and alters the distribution of actin and E-cadherin in cells cultured in 2D on solid supports. Overexpression of full-length KIF17 constructs or truncation mutants containing the N-terminal motor domain resulted in accumulation of newly incorporated GFP–actin into junctional actin foci, cleared E-cadherin from cytoplasmic vesicles and stabilized cell–cell adhesions to challenge with calcium depletion. Expression of these KIF17 constructs also increased cellular levels of active RhoA, whereas active RhoA was diminished in KIF17-depleted cells. Inhibition of RhoA or its effector ROCK, or expression of LIMK1 kinase-dead or activated cofilinS3A inhibited KIF17-induced junctional actin accumulation. Interestingly, KIF17 activity toward actin depends on the motor domain but is independent of microtubule binding. Together, these data show that KIF17 can modify RhoA–GTPase signaling to influence junctional actin and the stability of the apical junctional complex of epithelial cells. PMID:26759174

  9. Measurements of B meson decays to (omega)K* and (omega)(rho)

    SciTech Connect

    Aubert, B; Cheng, C H; Lange, D J; Simani, M C; . Wright, D M; Abrams, G S; Borgland, A W; Breon, A B; Brown, D N; Button-Shafer, J; Cahn, R H; Charles, E; Day, C T; Gill, M S; Gritsan, A V; Groysman, Y; Jacobsen, R G; Kadel, R W; Kadyk, J; Kerth, L T; Kolomensky, Y G; Kukartsev, G; Lynch, G; Mir, L M; Oddone, P J; Orimoto, T J; Pripstein, M; Roe, N A; Ronan, M T; Wenzel, W A; Abe, T; Aston, D; Bartoldus, R; Berger, N; Boyarski, A M; Buchmueller, O L; Claus, R; Convery, M R; Cristinziani, M; De Nardo, G; Dong, D; Dorfan, J; Dujmic, D; Dunwoodie, W; Fan, S; Field, R C; Glanzman, T; Gowdy, S J; Hadig, T; Halyo, V; Hast, C; Hryn'ova, T; Innes, W R; Kelsey, M H; Kim, P; Kocian, M L; Keith, D W . S; Libby, J; Luitz, S; Luth, V; Lynch, H L; Marsiske, H; Messner, R; Muller, D R; O'Grady, C P; Ozcan, V E; Perazzo, A; Perl, M; Petrak, S; Ratcliff, B N; Roodman, A; Salnikov, A A; Schindler, R H; Schwiening, J; Snyder, A; Soha, A; Stelzer, H; Strube, J; Su, D; Sullivan, M K; Va'vra, J; Wagner, S R; Weaver, M; Wisniewski, W J; Wittgen, M; Wright, D H; Yarritu, A K; Young, C C; Collaboration, B

    2006-03-14

    The authors describe searches for B meson decays to the charmless vector-vector final states {omega}K* and {omega}{rho} in 89 million B{bar B} pairs produced in e{sup +}e{sup -} annihilation at {radical}s = 10.58 GeV.

  10. Rho0 Photoproduction in Ultra-Peripheral Relativistic Heavy Ion Collisions with STAR

    SciTech Connect

    STAR Coll

    2007-12-20

    Photoproduction reactions occur when the electromagnetic field of a relativistic heavy ion interacts with another heavy ion. The STAR collaboration presents a measurement of {rho}{sup 0} and direct {pi}{sup +}{pi}{sup -} photoproduction in ultra-peripheral relativistic heavy ion collisions at {radical}s{sub NN} = 200 GeV. We observe both exclusive photoproduction and photoproduction accompanied by mutual Coulomb excitation. We find a coherent cross-section of {sigma}(AuAu {yields} Au*Au* {rho}{sup 0}) = 530 {+-} 19 (stat.) {+-} 57 (syst.) mb, in accord with theoretical calculations based on a Glauber approach, but considerably below the predictions of a color dipole model. The {rho}{sup 0} transverse momentum spectrum (p{sub T}{sup 2}) is fit by a double exponential curve including both coherent and incoherent coupling to the target nucleus; we find {sigma}{sub inc}/{sigma}{sub coh} = 0.29 {+-} 0.03 (stat.) {+-} 0.08 (syst.). The ratio of direct {pi}{sup +}{pi}{sup -} production is comparable to that observed in {gamma}p collisions at HERA, and appears to be independent of photon energy. Finally, the measured {rho}{sup 0} spin helicity matrix elements agree within errors with the expected s-channel helicity conservation.

  11. Protein Kinase D Regulates RhoA Activity via Rhotekin Phosphorylation*

    PubMed Central

    Pusapati, Ganesh V.; Eiseler, Tim; Rykx, An; Vandoninck, Sandy; Derua, Rita; Waelkens, Etienne; Van Lint, Johan; von Wichert, Götz; Seufferlein, Thomas

    2012-01-01

    The members of the protein kinase D (PKD) family of serine/threonine kinases are major targets for tumor-promoting phorbol esters, G protein-coupled receptors, and activated protein kinase C isoforms (PKCs). The expanding list of cellular processes in which PKDs exert their function via phosphorylation of various substrates include proliferation, apoptosis, migration, angiogenesis, and vesicle trafficking. Therefore, identification of novel PKD substrates is necessary to understand the profound role of this kinase family in signal transduction. Here, we show that rhotekin, an effector of RhoA GTPase, is a novel substrate of PKD. We identified Ser-435 in rhotekin as the potential site targeted by PKD in vivo. Expression of a phosphomimetic S435E rhotekin mutant resulted in an increase of endogenous active RhoA GTPase levels. Phosphorylation of rhotekin by PKD2 modulates the anchoring of the RhoA in the plasma membrane. Consequently, the S435E rhotekin mutant displayed enhanced stress fiber formation when expressed in serum-starved fibroblasts. Our data thus identify a novel role of PKD as a regulator of RhoA activity and actin stress fiber formation through phosphorylation of rhotekin. PMID:22228765

  12. Role of the Rho GTPase Rac in the activation of the phagocyte NADPH oxidase

    PubMed Central

    Pick, Edgar

    2014-01-01

    The superoxide-generating NADPH oxidase of phagocytes consists of the membrane-associated cytochrome b558 (a heterodimer of Nox2 and p22phox) and 4 cytosolic components: p47phox, p67phox, p40phox, and the small GTPase, Rac, in complex with RhoGDI. Superoxide is produced by the NADPH-driven reduction of molecular oxygen, via a redox gradient located in Nox2. Electron flow in Nox2 is initiated by interaction with cytosolic components, which translocate to the membrane, p67phox playing the central role. The participation of Rac is expressed in the following sequence: (1) Translocation of the RacGDP-RhoGDI complex to the membrane; (2) Dissociation of RacGDP from RhoGDI; (3) GDP to GTP exchange on Rac, mediated by a guanine nucleotide exchange factor; (4) Binding of RacGTP to p67phox; (5) Induction of a conformational change in p67phox, promoting interaction with Nox2. The particular involvement of Rac in NADPH oxidase assembly serves as a paradigm for signaling by Rho GTPases, in general. PMID:24598074

  13. Modelling GTPase dynamics to understand RhoA-driven cancer cell invasion

    PubMed Central

    Hetmanski, Joseph H.R.; Schwartz, Jean-Marc; Caswell, Patrick T.

    2016-01-01

    Metastasis, initially driven by cells migrating and invading through the local environment, leads to most cancer-associated deaths. Cells can use a variety of modes to move in vitro, all of which depend on Rho GTPases at some level. While traditionally it was thought that Rac1 activity drives protrusive lamellipodia at the leading edge of a polarised cell while RhoA drives rear retraction, more recent work in 3D microenvironments has revealed a much more complicated picture of GTPase dynamics. In particular, RhoA activity can dominate the leading edge polymerisation of actin to form filopodial actin-spike protrusions that drive more invasive cell migration. We recently described a potential mechanism to abrogate this pro-invasive localised leading edge Rac1 to RhoA switch via manipulation of a negative feedback loop that was revealed by adopting a logical modelling approach. Both challenging dogma and taking a formal, mathematical approach to understanding signalling involved in motility may be vital to harnessing harmful cell migration and preventing metastasis in future research. PMID:27913679

  14. RhoA and ROCK mediate histamine-induced vascular leakage and anaphylactic shock.

    PubMed

    Mikelis, Constantinos M; Simaan, May; Ando, Koji; Fukuhara, Shigetomo; Sakurai, Atsuko; Amornphimoltham, Panomwat; Masedunskas, Andrius; Weigert, Roberto; Chavakis, Triantafyllos; Adams, Ralf H; Offermanns, Stefan; Mochizuki, Naoki; Zheng, Yi; Gutkind, J Silvio

    2015-04-10

    Histamine-induced vascular leakage is an integral component of many highly prevalent human diseases, including allergies, asthma and anaphylaxis. Yet, how histamine induces the disruption of the endothelial barrier is not well defined. By using genetically modified animal models, pharmacologic inhibitors and a synthetic biology approach, here we show that the small GTPase RhoA mediates histamine-induced vascular leakage. Histamine causes the rapid formation of focal adherens junctions, disrupting the endothelial barrier by acting on H1R Gαq-coupled receptors, which is blunted in endothelial Gαq/11 KO mice. Interfering with RhoA and ROCK function abolishes endothelial permeability, while phospholipase Cβ plays a limited role. Moreover, endothelial-specific RhoA gene deletion prevents vascular leakage and passive cutaneous anaphylaxis in vivo, and ROCK inhibitors protect from lethal systemic anaphylaxis. This study supports a key role for the RhoA signalling circuitry in vascular permeability, thereby identifying novel pharmacological targets for many human diseases characterized by aberrant vascular leakage.

  15. Role of rho kinase in the functional and dysfunctional tonic smooth muscles.

    PubMed

    de Godoy, Márcio A F; Rattan, Satish

    2011-07-01

    Tonic smooth muscles play pivotal roles in the pathophysiology of debilitating diseases of the gastrointestinal and cardiovascular systems. Tonic smooth muscles differ from phasic smooth muscles in the ability to spontaneously develop myogenic tone. This ability has been primarily attributed to the local production of specific neurohumoral substances that can work in conjunction with calcium sensitization via signal transduction events associated with the Ras homolog gene family, member A (RhoA)/Rho-associated, coiled-coil containing protein kinase 2 (ROCK II) pathways. In this article, we discuss the molecular pathways involved in the myogenic properties of tonic smooth muscles, particularly the contribution of protein kinase C vs the RhoA/ROCK II pathway in the genesis of basal tone, pathophysiology and novel therapeutic approaches for certain gastrointestinal and cardiovascular diseases. Emerging evidence suggests that manipulation of RhoA/ROCK II activity through inhibitors or silencing of RNA interface techniques could represent a new therapeutic approach for various gastrointestinal and cardiovascular diseases.

  16. NG2-mediated Rho activation promotes amoeboid invasiveness of cancer cells.

    PubMed

    Paňková, Daniela; Jobe, Njainday; Kratochvílová, Magdalena; Buccione, Roberto; Brábek, Jan; Rösel, Daniel

    2012-01-01

    The aim of this study was to analyze the potential role of NG2 chondroitin sulfate proteoglycan in amoeboid morphology and invasiveness of cancer cells. In the highly metastatic amoeboid cell lines A3 and A375M2, siRNA-mediated down-regulation of NG2 induced an amoeboid-mesenchymal transition associated with decreased invasiveness in 3D collagen and inactivation of the GTPase Rho. Conversely, the expression of NG2 in mesenchymal sarcoma K2 cells as well as in A375M2 cells resulted in an enhanced amoeboid phenotype associated with increased invasiveness and elevated Rho-GTP levels. Remarkably, the amoeboid-mesenchymal transition in A375M2 cells triggered by NG2 down-regulation was associated with increased extracellular matrix-degrading ability, although this was not sufficient to compensate for the decreased invasive capability caused by down-regulated Rho/ROCK signaling. Conversely, in K2 cells with overexpression of NG2, the ability to degrade the extracellular matrix was greatly reduced. Taken together, we suggest that NG2-mediated activation of Rho leading to effective amoeboid invasiveness is a possible mechanism through which NG2 could contribute to tumor cell invasion and metastasis.

  17. Migration of epithelial cells on laminins: RhoA antagonizes directionally persistent migration.

    PubMed

    Zhang, Zhigang; Chometon, Gretel; Wen, Tingting; Qu, Haiyan; Mauch, Cornelia; Krieg, Thomas; Aumailley, Monique

    2011-01-01

    Spatial and temporal expression of laminin isoforms is assumed to provide specific local information to neighboring cells. Here, we report the remarkably selective presence of LM-111 at the very tip of hair follicles where LM-332 is absent, suggesting that epithelial cells lining the dermal-epidermal junction at this location may receive different signals from the two laminins. This hypothesis was tested in vitro by characterizing with functional and molecular assays the comportment of keratinocytes exposed to LM-111 and LM-332. The two laminins induced morphologically distinct focal adhesions, and LM-332, but not LM-111, elicited persistent migration of keratinocytes. The different impact on cellular behavior was associated with distinct activation patterns of Rho GTPases and other signaling intermediates. In particular, while LM-111 triggered a robust activation of Cdc42, LM-332 provoked a strong and sustained activation of FAK. Interestingly, activation of Rac1 was necessary but not sufficient to promote migration because there was no directed migration on LM-111 despite Rac1 activation. In contrast, RhoA antagonized directional migration, since silencing of RhoA by RNA interference boosted unidirectional migration on LM-332. Molecular analysis of the role of RhoA strongly suggested that the mechanisms involve disassembly of cell-cell contacts, loss of the cortical actin network, mobilization of α6β4 integrin out of stable adhesions, and displacement of the integrin from its association with the insoluble pool of intermediate filaments.

  18. Opposing roles for distinct LINC complexes in regulation of the small GTPase RhoA

    PubMed Central

    Thakar, Ketan; May, Christopher K.; Rogers, Anna; Carroll, Christopher W.

    2017-01-01

    Linker of Nucleoskeleton and Cytoskeleton (LINC) complexes span the nuclear envelope and transduce force from dynamic cytoskeletal networks to the nuclear lamina. Here we show that LINC complexes also signal from the nuclear envelope to critical regulators of the actin cytoskeleton. Specifically, we find that LINC complexes that contain the inner nuclear membrane protein Sun2 promote focal adhesion assembly by activating the small GTPase RhoA. A key effector in this process is the transcription factor/coactivator complex composed of SRF/Mkl1. A constitutively active form of SRF/Mkl1 was not sufficient to induce focal adhesion assembly in cells lacking Sun2, however, suggesting that LINC complexes support RhoA activity through a transcription-independent mechanism. Strikingly, we also find that the inner nuclear membrane protein Sun1 antagonizes Sun2 LINC complexes and inhibits RhoA activation and focal adhesion assembly. Thus different LINC complexes have opposing roles in the transcription-independent control of the actin cytoskeleton through the small GTPase RhoA. PMID:28035049

  19. p190-B, A Novel RhoGAP, In Mammary Gland Development and Breast Cancer Progression

    DTIC Science & Technology

    2006-09-01

    the membranes (data not shown). Acknowledgments We thank Shirley Small for her help with animal husbandry and colony management; Maria Gonzalez -Rimbau...buds and breast cancer. Cell Growth Differ 2000 11:343–354 13. Chakravarty G, Hadsell D, Buitrago W, Settleman J, Rosen JM 2003 p190-B RhoGAP regulates

  20. AKAP-Lbc: a molecular scaffold for the integration of cyclic AMP and Rho transduction pathways.

    PubMed

    Diviani, Dario; Baisamy, Laurent; Appert-Collin, Aline

    2006-07-01

    A Kinase-anchoring proteins (AKAPs) are a family of functionally related proteins involved in the targeting of the PKA holoenzyme towards specific physiological substrates. We have recently identified a novel anchoring protein expressed in cardiomyocytes, called AKAP-Lbc, that functions as a PKA-targeting protein as well as a guanine nucleotide exchange factor (GEF) that activates the GTPase RhoA. Here, we discuss the most recent findings elucidating the molecular mechanisms and the transduction pathways involved in the regulation of the AKAP-Lbc signaling complex inside cells. We could show that AKAP-Lbc is regulated in a bi-directional manner by signals that activate or deactivate its Rho-GEF activity. Activation of AKAP-Lbc occurs in response to agonists that stimulate G proteins coupled receptors linked to the heterotrimeric G protein G12, whereas inactivation occurs through mechanisms that require phosphorylation of AKAP-Lbc by anchored PKA and subsequent recruitment of the regulatory protein 14-3-3. Interestingly, we could demonstrate that AKAP-Lbc can form homo-oligomers inside cells and that 14-3-3 can inhibit the Rho-GEF activity of AKAP-Lbc only when the anchoring protein adopts an oligome