Abstract
Rhizobium leguminosarum NodD binds to the nod box of the inducible nod gene nodA as a V-shaped tetramer and bends the nod box. In this work, we show that the nod gene inducer naringenin decreased gel mobility of nod box DNA-NodD complexes by sharpening the NodD-induced DNA bend, which correlated with nodA transcription activation. NodD can induce different DNA bends when the distance between the two half-sites of the nod box was modified, which severely affected NodD-mediated transcriptional control. One or two base pairs were deleted from, or inserted into, the two half-sites of the nod box of nodA. Circular permutation assays showed that such distance modulations allowed NodD to induce relaxed or sharpened DNA bending. In the case of 1 bp deletion, where the DNA bends were more relaxed than in the wild type, nodA transcription was repressed both in the absence and in the presence of inducer naringenin. In the cases of 1 and 2 bp insertion, where the DNA bends were much sharper than in wild type in the absence or presence of the inducer naringenin, nodA transcription was initiated constitutively with no requirement for the inducer naringenin or, even, the NodD regulating protein.