Analysis of the expression from Rhizobium meliloti fix-promoters in other Rhizobium backgrounds.

Abstract

Using translational fusions to lacZ, we have measured expression from the promoters of Rhizobium meliloti regulatory genes, nifA and fixK, and structural genes, nifH and fixA, in other fast-growing rhizobia whose nitrogen fixation regulation is less known. Neither nifA nor fixK promoters were activated under both free-living microaerobic and symbiotic conditions, except in R. tropici, where clear symbiotic activation of either nifA or fixK expression could be observed. Both nifH and fixA promoters showed strong heterologous activation during symbiosis and weak activation under free-living nitrogen starvation conditions. Only when the nifH promoter was in R. tropici and R. leguminosarum bv. phaseoli, was clear induction observed in the microaerobic free-living state. Deletion analysis of these promoters suggested that a NifA binding site (UAS) was needed for full heterologous activation of nifHp, either in microaerobiosis or symbiosis. In contrast, the UAS region seemed to be unnecessary for fixA activation. However, a region containing a potential integration host factor (IHF) binding site was observed to be needed for complete heterologous symbiotic induction from fixAp. The moderate induction observed in nitrogen-free medium only required the sigma 54 holoenzyme recognition sequence; this may be indicative of the existence of non-specific activation by NtrC-like proteins. Our results suggest possible common and different features in the control mechanisms of the nitrogen fixation gene expression among Rhizobium species.