The genetic dissection of chemotaxis in agrobacterium tumefaciens

Abstract

A range of sugars, many of them characteristic of plant extracts were tested as potential chemoattractants for Agrobacierium .The results divided the sugars into 4 groups of attrcictants and indicated the presence of a highly sensitive chemotaxis system in A. tumefaciens. Motility in Agrobactewrium consisted of long straight runs, with relatively few tumbles or stops. The propulsive mechanism seemed to resemble that of Rhizobium. Methionine-starved methionine auxotrophs of A. tumefaciens , although fully motile, were non-chemotactic to sucrose or acetosyringone, unless supplemented with exogenous methionine. Neither ethionine nor a-methyl-DL-methionine could correct the non-chemotactic phenotype, while seleno- DL-methionine partially restored taxis. Pulse-labelling of A.tumefaciens with L- [methyl-(^3) H] - methionine in the presence of chloramphenicol, and an attractant resulted in the appearance of 2 radio-labelled proteins of approximately 55KDa. Thus, in A. tumefaciens, chemotactic responses may be associated with the transfer of methyl groups from methionine via S-eidenosyl methionine to MCPs. Using transposon mutagenesis a battery of A.tumefaciens chemotaxis mutants were generated and characterized. A number of mutated behavioural genes were isolated using the kanamycin resistant determinant of Tn5 as a positive selectable marker. Tn5 flanking sequences were used as probes to recover wild-type behavioural genes from a gene library constructed in the cosmid pLAFR3. Behavioural genes were found to be clustered on the A. tumefaciens chromosome and to possess similarity with behavioural genes from R.meliloti.