STUDIES ON HETEROLOGOUS EXPRESSION OF Rhizobium meliloti nifA GENE AND OXYGEN SENSITIVITY OF ITS PRODUCT

When the R. meliloti (Rm) nifA’-lacZ fusion-carrying plasmids were introduced intothe strains of Agrobacterium tumefaciens, R. trifolii and R. astragalus, β-galactosidaseactivity was demonstrated. However, activity was not induced by microaerobiosis. Further-more, R. meliloti nifA’-lacZ fusion was also not expressed in the nodule bacteroids ofR. trifolii and R. astragalus. We speculate that some factor(s) important for the inductionof Rm nifA presumed to be the fixLJ regulatory system would not be operative in thesebacteria. Experiments using R. meliloti nifH’-lacZ/ K. Pneumoniae nifH’-lacZ fusion and theconstitutive Rm nifA system to test the nifA-dependent expression of nifH’-lacZ underaerobic and microaerobic conditions in E. coli were performed. The inhibition of the RmnifA activation of nifH’-lacZ expression in the bacteria grown in the aerobic conditionwas shown. Assays on the Rm nifA-m RNA produced by the constitutive Rm nifA in E.coli under aerobic and microaerobic conditions with the cloned

When the R. meliloti (Rm) nifA'-lacZ fusion-carrying plasmids were introduced into the strains of Agrobacterium tumefaciens, R. trifolii and R. astragalus, β-galactosidase activity was demonstrated. However, activity was not induced by microaerobiosis. Further- more, R. meliloti nifA'-lacZ fusion was also not expressed in the nodule bacteroids of R. trifolii and R. astragalus. We speculate that some factor(s) important for the induction of Rm nifA presumed to be the fixLJ regulatory system would not be operative in these bacteria. Experiments using R. meliloti nifH'-lacZ/ K. Pneumoniae nifH'-lacZ fusion and the constitutive Rm nifA system to test the nifA-dependent expression of nifH'-lacZ under aerobic and microaerobic conditions in E. coli were performed. The inhibition of the Rm nifA activation of nifH'-lacZ expression in the bacteria grown in the aerobic condition was shown. Assays on the Rm nifA-m RNA produced by the constitutive Rm nifA in E. coli under aerobic and microaerobic conditions with the cloned nifA as a probe for dot blot hybridization showed a marked decrease of Rm nifA mRNA when the bacteria were grown under aeration.