Identification and characterization of two nitrogen-regulated genes of the cyanobacterium Synechococcus sp. strain PCC7942 required for maximum efficiency of nitrogen assimilation.

Two nitrogen-regulated genes were found in the genomic DNA region upstream of the nirA operon involved in uptake and utilization of nitrate in Synechococcus sp. strain PCC7942. The two genes (nirB and ntcB) are transcribed divergently from nirA and encode proteins of 349 and 309 amino acid residues, respectively. The levels of nirB and ntcB transcripts were low in cells growing on ammonium and increased upon transfer of ammonium-grown cells to nitrate-containing medium. The deduced NirB protein sequence has no similarities to other known proteins, whereas the deduced NtcB protein sequence is homologous to bacterial transcriptional activators of the LysR family. Defined mutants constructed by interrupting nirB or ntcB with a drug resistance marker grew as fast as the wild-type strain on ammonium but grew slower than the wild-type strain on nitrate or nitrite. The nirB mutant had higher activities of nitrate reductase, glutamine synthetase, and glutamate synthase than the wild-type strain, but its nitrite reductase activity was 40% of the wild-type levels. The mutant excreted nitrite into the medium during growth on nitrate, showing that nitrite reductase limits nitrate assimilation. These findings suggested that nirB is required for expression of maximum nitrite reductase activity. When grown on ammonium, the nirB mutant grew normally but cultures of the ntcB mutant still showed a yellowish-green color typical of nitrogen-limited cells. NtcB seems to regulate utilization of fixed nitrogen by controlling the expression of a certain gene(s) involved in nitrogen metabolism.