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COMPARISON OF LATE SYM GENES IN RHIZOBIUM SPECIES AND CONSTRUCTION OF IMPROVED STRAINS

Objective

THE SPECIES CHOSEN FOR THIS WORK, R. MELILOTI, R. LEGUMINOSARUM AND R. PHASEOLI NODULATE IMPORTANT CROPS IN THE EUROPEAN COMMUNITIES, SUCH AS ALFALFA, PEA, BROAD BEAN AND BEAN. THE IMPACT EXPECTED FROM THE IMPROVEMENT OF THESE SYMBIOSES IS A REDUCTION IN THE ENERGY AND PROTEIN DEPENDENCE IN THE COMMUNITY AS WELL AS A LOWERING OF FERTILIZATION COSTS. METHODS ENABLING TO MANIPULATE THE SOIL MICROFLORA, SPECIALLY WITH REFERENCE TO ALFALFA AND PEA, WILL GREATLY ENHANCE THE SIGNFICANCE OF THE ABOVE DESCRIBED RESEARCH.
Rhizobium cell surface components were found to be required for a successful symbiotic interaction with the host plant. Regulatory genes were identified which are involved in the oxygen controlled expression of nitrogen fixation genes. The regulatory pathway differs in Rhizobium meliloti and Rhizobium leguminosarum and may represent an important variable between Rhizobium species.

A laboratory model was created to study the gene transfer between Rhizobium meliloti strains within alfalfa nodules. Nodulation and infection defective strains were used, which are able to complement each other to form functional nodules. Gene transfer between the strains was observed.

Research was carried out in order to gain a deeper insight into the steps and regulatory pathways involved in the development of the Rhizobium legume nitrogen fixing symbiosis, and to identify genes controlling these processes in Rhizobium. This involved: identification and genetic analysis of mutants defective in later stages of symbiotic development; isolation of the relevant genes and investigation of their function and regulation; and establishing the significance of these genes by analysing and comparing different Rhizobium strains and species (mainly R meliloti and R leguminosarum).

A cluster of new genes essential for symbiotic nitrogen fixation was indentified in R meliloti. Most of these genes encode membrane proteins involved, for example, in transferring electrons to the nitrogenase enzyme. Similar genes were present in a wide range of other symbiotic nitrogen fixing species. Lipopolysaccharide, another membrane component, was required for successful symbiotic interaction. Similarly, strains assumed to have surface alterations due to the presence of IncPl plasmids showed defects in symbiotic nitrogen fixation. Regulatory genes were identified which ensure that nitrogen fixation genes are expressed only in low oxygen environments (such as within root nodules) appropriate for nitrogenase function. This regulatory pathway differs in R meliloti and R leguminosarum and may represent an important variable between Rhizobium species.
LATE SYM GENES ARE SUPPOSED TO CONTROL SYMBIOTIC EFFICIENCY. THIS EFFICIENCY MAY BE IMPROVED EITHER BY GENETIC ENGINEERING OF LATE SYM GENES OR BY CREATING MORE EFFICIENT COMBINATIONS OF THESE GENES IN INTERSPECIFIC HYBRIDS. BOTH METHODS NECESSITATE IDENTIFICATION AND STUDY OF THE EXPRESSION OF LATE SYM GENES. THE LABORATORY OF TOULOUSE WILL IDENTIFY LATE SYM GENES ON SYM MEMEGAPLASMID OF R. MELILOTI. LABORATORY OF DUBLIN WILL DETECT THOSE GENES EITHER ON THE CHROMOSOME OR ON PLASMIDS OF R. LEGUMINOSARUM.

LABORATORY OF HARPENDEN WILL ISOLATE MORE PLASMIDS OR CHROMOSOMAL FRAGMENTS ACROSS VARIOUS STRAINS OF R. LEGUMINOSARUM AND R. PHASEOLI. GENETIC AND MOLECULAR ANALYSIS OF THE REGIONS OF INTEREST WILL BE CARRIED OUT IN TOULOUSE, DUBLIN AND BIELEFELD, WHEREAS THE STUDY OF SYMBIOTIC EFFICIENCY OF THE HYBRID STRAINS CONSTRUCTED WILL BE THE TASK OF HARPENDEN. GENE EXPRESSION WILL BE STUDIED USING GENE FUSIONS IN LABORATORY OF TOULOUSE AND BIELFELD.

Funding Scheme

CSC - Cost-sharing contracts

Coordinator

UNIVERSITY OF BIELEFELD
Address
Universitaetstrasse 25
33501 Bielefeld
Germany

Participants (3)

Centre National de la Recherche Scientifique (CNRS)
France
Address
Chemin De Borde-rouge Auzeville
31326 Castanet-tolosan
ROTHAMSTED RESEARCH LIMITED
United Kingdom
Address
West Common
AL5 2JQ Harpenden, Herts
SCHOOL OF BIOLOGICAL SCIENCES
Ireland
Address

Dublin