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THE MOLECULAR BIOLOGY OF PLANT PATHOGENIC ERWINIA

Objective

BACTERIA OF THE GENUS ERWINIA ARE SIGNIFICANT PLANT PATHOGENS. IN PARTICULAR, THE "SOFT ROT GROUP" ARE THE MAJOR AETIOLOGICAL AGENTS OF VARIOUS DISEASES. A SIGNIFICANT NUMBER OF THE GENES CONTROLLING THE INFECTION PROCESS IN THE PATHOGEN WILL CODE FOR EXTRACELLULAR ENZYMES AND THE APPARATUS REQUIRED FOR THEIR EXPORT.

APART FROM THE OBVIOUS AGRICULTURAL IMPLICATIONS OF UNRAVELLING PATHOGENICITY FACTORS, KNOWLEDGE ON EXTRACELLULAR ENZYMES, IN PARTICULAR, WILL OPEN UP NEW POSSIBILITIES FOR INDUSTRIAL APPLICATIONS, INCLUDING :

- CONSTRUCTION OF NEW ERWINIA-BASED SECRETION VECTORS FOR FOREIGN POLYPEPTIDE PRODUCTION;
- TRANSFER OF ERWINIA PECTINASE AND CELLULASE GENES TO OTHER INDUSTRIAL MICROBES (ZYMOMONAS OR YEAST) TO INCREASE THEIR SUBSTRATE UTILIZATION FOR BIOMASS CONVERSION TO ALCOHOL OR SUGARS;
- GENETIC MANIPULATION OF ERWINIA STRAINS USED FOR COMMERCIAL PRODUCTION OF THE ANTI-LEUKAEMIA ENZYME ASPARAGINASE.
Erwinia chrysanthemi and E carotovora cause bacterial soft rot disease of ornamentals and important crops, causing great economic loss. No chemical control measures exist. The molecular biology of the pathogen has been developed in order to analyse the disease mechanism(s) and provide information which could eventually be exploited for rational disease control.

Genetic and biochemical tools were developed to analyse the pathogen. To characterize the individual cell wall degrading enzymes and assess their role in virulence, the enzyme genes (especially pectinases) were cloned and sequenced, and their regulation and the mechanism of secretion in the pathogen were studied. The role in virulence of each enzyme was assessed using genetically engineered strains and plant pathology assays and previously unidentified pathogenicity factors were searched for, using a combination of genetics, biochemistry and molecular biology.

Soft rot Erwinia species make many forms of extracellular enzymes, especially pectate lysase (Pels). E chrysanthemi carotovora secretes 2 Pels and retains 2 Pels in the periplasm. All Pels are independently expressed, although there is gene clustering. The Pels show differential importance in pathogenicity. The pectin catabolic pathway has been elucidated and many enzymes of this pathway are coordinately regulated by a common repressor. E chrysanthemi makes 2 cellulases (Cel) and E carotovora makes 1. The Cel Y of E chrysanthemi could be involved in induction of a plant defence mechanism. The Cel Y of E chrysanthemi could be involved in induction of a plant defence mechanism. The Cel catabolic pathway was determined and cellulase regulation is affected by environment stimuli. Pels and Cels are secreted by the same pathway. The cell envelope is important for virulence. Genes encoding novel virulence factors, induced by plant extracts have been identified.
THIS COLLABORATIVE PROJECT COMBINES GENETIC, BIOCHEMICAL AND MOLECULAR BIOLOGICAL APPROACHES TO INVESTIGATE THE MECHANISM OF PLANT DISEASE CAUSED BY ERWINIA. POTATO/ECA AND SAINT-PAULIA/ECHY INTERACTIONS WILL BE USED AS PATHOGENICITY MODELS. THE ANALYSIS OF THE SYNTHESIS, REGULATION AND EXPORT OF EXTRACELLULAR ENZYMES WILL BE COMBINED WITH GENETIC AND BIOCHEMICAL STUDIES OF HOST-SPECIFICITY, PO2 AND TEMPERATURE EFFECTS, TO DETERMINE THE RANGE OF FACTORS INVOLVED IN THE PLANT DISEASE.

INTERACTION BETWEEN HOST AND PATHOGEN IS A SEQUENCE OF GENETICALLY-CONTROLLED EVENTS (RECOGNITION, PENETRATION, INDUCTION OF AGGRESSIVE/DEFENSIVE RESPONSES, SYSTEMATIC SPREAD,...). USING MUTAGENESIS, AND MOLECULAR CLONING OF THE RESPECTIVE MUTATIONS, ALL STAGE-SPECIFIC RESPONSIBLE GENES IN THE PATHOGEN WILL BE DEFINED :

- ISOLATION AND CHARACTERIZATION OF NON-PATHOGENIC MUTANTS (WARWICK, BRUSSELS, PARIS);
- STRUCTURE, SYNTHESIS, REGULATION AND EXPORT OF EXTRACELLULAR ENZYMES (ALL 5 LABORATORIES);
- CHARACTERIZATION OF PRODUCTS OF THE SECRETORY APPARATUS, AND OTHER PRODUCTS OF PREVIOUSLY UNIDENTIFIED PATHOGENICITY GENES (ALL 5 LABORATORIES).

SPECIAL EXPERTISE :

ECA MODEL (WARWICK); ECHY (BELGIAN + FRENCH LABS); CHEMICAL/INSERTION MUTANTS (PARIS); PHAGE/BACTERIOCIN RESISTANT MUTANT (BRUSSELS); UNIDENTIFED PATHOGENICITY GENES (PARIS, WARWICK); INTERSPECIFIC COMPLEMENTATION OF ECA, ECHY MUTANTS (ALL LABS).

Funding Scheme

CSC - Cost-sharing contracts

Coordinator

Institut National de la Recherche Agronomique (INRA)
Address
11 Rue Jean Nicot
75007 Paris
France

Participants (4)

Centre National de la Recherche Scientifique (CNRS)
France
Address
31 Chemin Joseph Aiguier
13402 Marseille
Institut National des Sciences Appliquées de Lyon (INSA)
France
Address
20 Avenue Albert Einstein
69621 Villeurbanne
ULB
Belgium
Address

Rhode-st-genese
University of Warwick
United Kingdom
Address
Gibbet Hill Road
CV4 7AL Coventry