Activation of the Anabaena nir operon promoter requires both NtcA (CAP family) and NtcB (LysR family) transcription factors

A region of the genome of the heterocyst-forming cyanobacterium Anabaena sp. PCC 7120 containing the ntcB gene was identified. This region is located upstream from the nir operon involved in nitrate assimilation in this cyanobacterium. An Anabaena ntcB mutant was able to use ammonium and dinitrogen as sources of nitrogen for growth but was unable to assimilate nitrate. Enzymes of the nitrate reduction system were not synthesized in the ntcB mutant under derepression conditions. The transcription start-point of the Anabaena nir operon, which has been shown to be subjected to ammonium-stimulated repression and whose expression requires the global nitrogen regulator NtcA, was only weakly used in the ntcB mutant. The expression of the ntcB gene in strain PCC 7120 was also subjected to repression by ammonium and was found to take place from an NtcA-activated promoter located 31 bp upstream from the start of the ntcB gene. NtcB binds to the nir promoter region in vitro and protects a region localized just upstream from the NtcA-binding site in footprinting assays. These results showed that NtcB, a LysR-family protein, is required in addition to NtcA, a CAP-family protein, for the expression of genes encoding proteins specifically involved in nitrate assimilation in Anabaena sp. PCC 7120.