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Rho1 and other GTP-binding proteins are associated with vesicles carrying glucose oxidase activity from Fusarium oxysporum f. sp. lycopersici

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Abstract

In eucaryotic cells, the delivery of a secreted protein to the plasma membrane via vesicles must include transport, recognition, and fusion events. Proteins exposed on the cytoplasmic face of the secretory vesicles play a role in these events; these include the GTP-binding proteins, which are crucial components in this process. Fractions enriched with vesicles carrying glucose oxidase (GOX) activity from Fusarium oxysporum f. sp. lycopersici, a soilborne fungal pathogen causing vascular wilt on tomato plants, were obtained using two successive sucrose gradients, the first a linear-log and the second an isopycnic gradient. In this study, we used the following Fusarium strains: a wild-type and a strain carrying a Δrho1 loss-of-function mutation (presenting dramatically reduced virulence). By ADP-ribosylation with C3 exotoxin, and Western blot analysis with specific antibodies, we identified the small GTPases Rho1, Rho4, Cdc42 and Rab8, and a heterotrimeric Gα protein associated with vesicles carrying GOX activity. This was done for both strains, with the exception of Rho1, which was absent in the mutant strain; in addition, the levels of the Cdc42 protein were observed to be higher in the Δrho1 strain. These data indicate that three Rho proteins, Rho1, Rho4, and Cdc42, are present in secretory vesicles carrying GOX activity in F. oxysporum, and that Rho1 is not essential for the transport and secretion of, at least, cargo proteins carried in secretory vesicles, or Cdc42/Rho4 can fulfill its role in these events.

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References

  • Adamo JE, Rossi G, Brennwald P (1999) The Rho GTPase Rho3 has a direct role in exocytosis that is distinct from its role in actin polarity. Mol Biol Cell 10:4121–4133

    CAS  PubMed  Google Scholar 

  • Adamo JE, Moskow JJ, Gladfelter AS, Viterbo D, Lew DJ, Brennwald PJ (2001) Yeast Cdc42 functions at a late step in exocytosis, specifically during polarized growth of the emerging bud. J Cell Biol 155:581–592

    Article  CAS  PubMed  Google Scholar 

  • Ayscough KR, Stryker J, Pokala N, Sanders M, Crews P, Drubin DG (1997) High rates of actin filament turnover in budding yeast and roles for actin in establishment and maintenance of cell polarity revealed using the actin inhibitor latrunculin-A. J Cell Biol 137:399–416

    Article  CAS  PubMed  Google Scholar 

  • Barr FA, Leyte A, Mollner S, Pfeuffer T, Tooze SA, Huttner WB (1991) Trimeric G-proteins of the trans-Golgi network are involved in the formation of constitutive secretory vesicles and immature secretory granules. FEBS Lett 294:239–243

    Article  CAS  PubMed  Google Scholar 

  • Beckman CH (1987) The nature of wilt diseases of plants. The American Phytopatological Society, St. Paul, MN

    Google Scholar 

  • Boyd C, Hughes T, Pypaert M, Novick P (2004) Vesicles carry most exocyst subunits to exocytic sites marked by the remaining two subunits, Sec3p and Exo70p. J Cell Biol 167:889–901

    Article  CAS  PubMed  Google Scholar 

  • Brakke MK, Van Pelt N (1970) Linear-log sucrose gradients for estimating sedimentation coefficients of plan viruses and nucleic acids. Anal Biochem 38:56–64

    Article  CAS  PubMed  Google Scholar 

  • Cabib E, Drgonova J, Drgon T (1998) Role of small G proteins in yeast cell polarization and wall biosynthesis. Annu Rev Biochem 67:307–333

    Article  CAS  PubMed  Google Scholar 

  • Di Pietro AP, Madrid M, Caracuel Z, Delgado-Jarana J, Roncero MIG (2003) Fusarium oxysporum: exploring the molecular arsenal of a vascular wilt fungus. Mol Plant Pathol 4:315–325

    Article  CAS  Google Scholar 

  • Erickson JW, Cerione RA (2001) Multiple roles for Cdc42 in cell regulation. Curr Opin Cell Biol 13:153–157

    Article  CAS  PubMed  Google Scholar 

  • Govindan B, Bowser R, Novick P (1995) The role of Myo2, a yeast class V myosin, in vesicular transport. J Cell Biol 128:1055–1068

    Article  CAS  PubMed  Google Scholar 

  • Gronover CS, Kasulke D, Tudzynski P, Tudzynski B (2001) The role of G protein alpha subunits in the infection process of the gray mold fungus Botrytis cinerea. Mol Plant-Microbe Interact 14:1293–1302

    Article  CAS  PubMed  Google Scholar 

  • Guo W, Grant A, Novick P (1999) Exo84p is an exocyst protein essential for secretion. J Biol Chem 274:23558–23564

    Article  CAS  PubMed  Google Scholar 

  • Guo W, Sacher M, Barrowman J, Ferro-Novick S, Novick P (2000) Protein complexes in transport vesicle targeting. Trends Cell Biol 10:251–255

    Article  CAS  PubMed  Google Scholar 

  • Guo W, Tamanoi F, Novick P (2001) Spatial regulation of the exocyst complex by Rho1 GTPase. Nat Cell Biol 3:353–360

    Article  CAS  PubMed  Google Scholar 

  • Houterman PM, Speijer D, Dekker HL, De Koster CG, Cornelissen BJC, Rep M (2007) The mixed xylem sap proteome of Fusarium oxysporum-infected tomato plants. Mol Plant Pathol 8:215–221

    Article  CAS  PubMed  Google Scholar 

  • Jain S, Akiyama K, Mae K, Ohguchi T, Takata R (2002) Targeted disruption of a G protein α subunit gene results in reduced pathogenicity in Fusarium oxysporum. Curr Genet 41:407–413

    Article  CAS  PubMed  Google Scholar 

  • Jain S, Akiyama K, Takata R, Ohguchi T (2005) Signaling via the G protein alpha subunit FGA2 necessary for pathogenesis in Fusarium oxysporum. FEMS Microbiol Lett 243:165–172

    Article  CAS  PubMed  Google Scholar 

  • Johnson DI (1999) Cdc42: An essential Rho-type GTPase controlling eukaryotic cell polarity. Microbiol Mol Biol Rev 63:54–105

    CAS  PubMed  Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685

    Article  CAS  PubMed  Google Scholar 

  • Leiter E, Marx F, Pusztahelyi T, Haas H, Pocsi I (2004) Penicillium chrysogenum glucose oxidase a study on its antifungal effects. J Appl Microbiol 97:1201–1209

    Article  CAS  PubMed  Google Scholar 

  • Leyte A, Barr FA, Kehlenbach RH, Huttner WB (1992) Multiple trimeric G-proteins on the trans-Golgi network exert stimulatory and inhibitory effects on secretory vesicle formation. EMBO J 11:4795–4804

    CAS  PubMed  Google Scholar 

  • Lipschutz JH, Mostov KE (2002) Exocytosis: The many masters of the exocyst. Curr Biol 12:R212–R214

    Article  CAS  PubMed  Google Scholar 

  • Martínez-Rocha AL, Roncero MIG, López-Ramirez A, Mariné M, Guarro J, Martínez-Cadena G, Di Pietro A (2008) Rho1 has distinct functions in morphogenesis, cell wall biosynthesis and virulence of Fusarium oxysporum. Cell Microbiol 10:1339–1351

    Article  PubMed  Google Scholar 

  • Matsui Y, Toh-e A (1992) Yeast RHO3 and RHO4 ras superfamily genes are necessary for bud growth, and their defect is suppressed by a high dose of bud formation genes CDC42 and BEM1. Mol Cell Biol 12:5690–5699

    CAS  PubMed  Google Scholar 

  • Moreno-Jiménez R, García-Soto J, Martínez-Cadena G (2008) Small GTP-binding proteins are associated to chitosomes and vesicles carrying glucose oxidase from Mucor circinelloides. Microbiology 154:842–851

    Article  PubMed  Google Scholar 

  • Novick P, Guo W (2002) Ras family therapy: Rab, Rho and talk to the exocyst. Trends Cell Biol 12:247–249

    Article  CAS  PubMed  Google Scholar 

  • Park HO, Bi E (2007) Central roles of small GTPases in the development of cell polarity in yeast and beyond. Microbiol Mol Biol Rev 71:48–96

    Article  CAS  PubMed  Google Scholar 

  • Pruyne D, Bretscher A (2000) Polarization of cell growth in yeast. II. The role of the cortical actin organization. J Cell Sci 113:571–585

    CAS  PubMed  Google Scholar 

  • Ramasamy K, Kelley RL, Reddy CA (1985) Lack of lignin degradation by glucose oxidase-negative mutants of Phanerochaete chrysosporium. Biochem Biophys Res Commun 131:436–441

    Article  CAS  PubMed  Google Scholar 

  • Sekine A, Fujiwara M, Narumiya S (1989) Asparagine residue in the rho gene product is the modification site for botulinum ADP-ribosiltransferase. J Biol Chem 264:8602–8605

    CAS  PubMed  Google Scholar 

  • Solomon PS, Tan K-C, Sanchez P, Cooper RM, Oliver RP (2004) The disruption of a G alpha subunit sheds new light on the pathogenicity of Stagonospora nodorum on wheat. Mol Plant-Microbe Interact 17:456–466

    Article  CAS  PubMed  Google Scholar 

  • TerBush DR, Maurice T, Roth D, Novick P (1996) The exocyst is a multiprotein complex required for exocytosis in Saccharomyces cerevisiae. EMBO J 15:6483–6494

    CAS  PubMed  Google Scholar 

  • Wennerberg K, Rossman KL, Der CJ (2005) The Ras superfamily at a glance. J Cell Sci 118:843–846

    Article  CAS  PubMed  Google Scholar 

  • Wong CM, Wong KH, Chen XD (2008) Glucose oxidase: natural occurrence, function, properties and industrial applications. Appl Microbiol Biotechnol 78:927–938

    Article  CAS  PubMed  Google Scholar 

  • Wu H, Rossi G, Brennwald P (2008) The ghost in the machine: small GTPases as spatial regulators of exocytosis. Trends Cell Biol 18:397–404

    Article  CAS  PubMed  Google Scholar 

  • Zhang X, Bi E, Novick P, Du L, Kozminski KG, Lipschutz JH, Guo W (2001) Cdc42 Interacts with the exocyst and regulates polarized secretion. J Biol Chem 276:46745–46750

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This study was supported by the Mexican National Council of Science and Technology (CONACYT) under Grant 62568, and the Universidad de Guanajuato. K.M.S. is a doctoral student with a scholarship from CONACYT and the Universidad de Guanajuato.

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Authors and Affiliations

  1. División de Ciencias Naturales y Exactas, Departamento de Biología, Campus Guanajuato, Universidad de Guanajuato, Apdo. postal 187, 36000, Guanajuato, Gto, Mexico

    Karla Macías-Sánchez, Jesús García-Soto, Adriana López-Ramírez & Guadalupe Martínez-Cadena

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  1. Karla Macías-Sánchez
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  2. Jesús García-Soto
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  3. Adriana López-Ramírez
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  4. Guadalupe Martínez-Cadena
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Correspondence to Guadalupe Martínez-Cadena.

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Macías-Sánchez, K., García-Soto, J., López-Ramírez, A. et al. Rho1 and other GTP-binding proteins are associated with vesicles carrying glucose oxidase activity from Fusarium oxysporum f. sp. lycopersici . Antonie van Leeuwenhoek 99, 671–680 (2011). https://doi.org/10.1007/s10482-010-9543-0

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