Pseudomonas syringae pv. phaseolicola genomic clones harboring heterologous DNA sequences suppress the same phaseolotoxin-deficient mutants.
Cosmid cloning and mutagenesis were used to identify genes involved in the production of phaseolotoxin, the chlorosis-inducing phytotoxin of Pseudomonas syringae pv. phaseolicola, the causal agent of halo blight of bean (Phaseolus vulgaris L.). Eight stable clones were isolated from a genomic cosmid library by en masse mating to 10 ethyl methanesulfonate (EMS)-induced Tox- mutants. In cross-matings, each suppressed all 10 mutants as well as an additional 70 EMS-induced Tox- mutants (and one UV-induced Tox- mutant). On the basis of restriction endonuclease analysis and hybridization studies, the clones were grouped into three classes. Clones in a particular class shared common fragments, whereas clones in different classes did not. Clones from class I (but not classes II and III) also suppressed Tn5-induced Tox- mutants. Interposon mutagenesis and marker exchange of a representative clone from class III into the wild-type genome did not alter its Tox+ phenotype, indicating that this clone does not harbor structural or regulatory genes involved in phaseolotoxin production. We suggest that the genome of P. syringae pv. phaseolicola contains a "hot spot" in one of the functions involved in toxin production which is affected by EMS and UV and that heterologous clones are able to suppress the Tox- phenotype because their inserts encode products that are able to substitute for the product of the mutated gene. Alternatively, the inserts may contain sequences which titrate a repressor protein. In either case, the data suggest that suppression of EMS- and UV-induced mutants occurs when heterologous clones are present in multiple copies.
Kamdar
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Department of Plant Pathology, University of Hawaii, Honolulu 96822.
Rowley
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KB
Clements
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Patil
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D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D003360:Cosmids; D004269:DNA, Bacterial / Q000235:genetics*; D005020:Ethyl Methanesulfonate / Q000494:pharmacology; D005098:Exotoxins / Q000096:biosynthesis / Q000235:genetics* / Q000528:radiation effects; D016296:Mutagenesis; D009154:Mutation; D009952:Ornithine / Q000031:analogs & derivatives; D010641:Phenotype; D011549:Pseudomonas / Q000235:genetics*; D015183:Restriction Mapping; D013489:Suppression, Genetic; D014466:Ultraviolet Rays
D004269:DNA, Bacterial; D005098:Exotoxins; C021234:phaseolotoxin; D005020:Ethyl Methanesulfonate; D009952:Ornithine