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First published online 22 December 2004
doi: 10.1242/jcs.01616

Journal of Cell Science 118, 323-329 (2005)
Published by The Company of Biologists 2005
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Research Article

Functional analysis of CLPT1, a Rab/GTPase required for protein secretion and pathogenesis in the plant fungal pathogen Colletotrichum lindemuthianum

Piyawan Siriputthaiwan1, Alain Jauneau2, Corentin Herbert1, Daphné Garcin1 and Bernard Dumas1,*

1 UMR 5546 CNRS-Université Paul Sabatier
2 IFR 40 Pôle de Biotechnologie Végétale, 24 Chemin de Borde Rouge, BP17 Auzeville, 31326 Castanet-Tolosan, France

* Author for correspondence (e-mail: dumas{at}scsv.ups-tlse.fr)

Accepted 27 October 2004

In eukaryotic cells, Rab/GTPases are major regulators of vesicular trafficking and are involved in essential processes including exocytosis, endocytosis and cellular differentiation. To investigate the role of these proteins in fungal pathogenicity, a dominant-negative mutant allele of CLPT1, a Rab/GTPase of the bean pathogen Colletotrichum lindemuthianum, was expressed in transgenic strains. This mutated gene encodes the amino-acid substitution N123I analogous to the N133I substitution in a known trans-dominant inhibitor of the Sec4 Rab/GTPase from Saccharomyces cerevisiae. A pectinase gene promoter was used to drive the CLPT1(N123I) allele in C. lindemuthianum, allowing the expression of the foreign gene on pectin medium and during pathogenesis, but not on glucose. The same strategy was used to overexpress the wild-type CLPT1 allele. During growth on pectin medium, production of extracellular pectinases was strongly impaired only in CLPT1(N123I)-expressing strains. Cytological analysis revealed that CLPT1(N123I) strains accumulated intracellular aggregates only on pectin, resulting from the fusion of vesicles containing polysaccharides or glycoproteins. Moreover, these strains showed a severe reduction of pathogenesis and were unable to penetrate the host cells. These results indicated that the Rab/GTPase CLPT1 is essential for fungal pathogenesis by regulating the intracellular transport of secretory vesicles involved in the delivery of proteins to the extracellular medium and differentiation of infectious structures.

Key words: Fungal pathogenicity, Colletotrichum lindemuthianum, Secretion




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