Unique features of Erwinia chrysanthemi (Dickeya dadantii) RA3B genes involved in the blue indigoidine production.

Erwinia chrysanthemi (Ech) RA3B produces a large amount of blue indigoidine. Using Tn5-induced mutagenesis, three indigoidine-deficient mutants were generated. Followed by library screening, a 5.8kb fragment complemented mutants for indigoidine synthesis was cloned. This fragment contains four complete open-reading frames (ORFs), pecS, pecM, idgA, and idgB, and two partial ORFs, argG, and idgC. These genes are nearly identical to those in strain Ech3937. Primer extension assays demonstrated a clear transcriptional start site prior to idgA, while no promoter preceding idgB and idgC was detected, suggesting that idgA, idgB, and idgC are organized as one transcription unit. In contrast, indAB is separated from indC in Ech3937. Interestingly, an ERIC sequence was present between idgB and idgC in place of the promoter region of the homolog indC, which may contribute to the loss of promoter activity in RA3B. Futhermore, idgB mutant displayed much lighter blue color, while indB mutant appeared white on media. Overexpression of pecS in RA3B resulted in significantly reduced indigoidine production and idgC transcript. Moreover, gel shift and luxAB reporter assays revealed that PecS specifically binds to the sequence preceding idgA and inhibits gene expression, which is consistent with the results observed in Ech3937.

Organization and expression of nitrogen-fixation genes in the aerobic nitrogen-fixing unicellular cyanobacterium Synechococcus sp. strain RF-1.

Sixteen nif and 'nif-associated' genes (expressed only under conditions of nitrogen fixation) in Synechococcus sp. strain RF-1 have been cloned and sequenced. All of the nif and nif-associated genes identified in Synechococcus RF-1 were arranged in a continuous cluster spanning approximately 18 kb and containing seven operons. The nifH operon (nifH-nifD-nifK) has been reported previously. nifB, fdxN, nifS, nifU and nifP were found to be located upstream of the nifH operon. nifB-fdxN-nifS-nifU were expressed as an operon. A nifP-like gene was found to be located just upstream of nifB. nifE, nifN, nifX, nifW and the nif-associated hesA, hesB and 'fdx' were found to be located downstream from nifK. The genes located downstream from nifK are arranged nifE-nifN-nifX-orf-nifW-hesA-hesB-'+ ++fdx' and span approximately 7 kb. The function of the ORF situated between nifX and nifW is not known. However, it was identified as a counterpart of ORF-2 in Anabaena sp. strain PCC 7120 based on the deduced amino acid sequence. Northern hybridization and primer extension analysis indicated that the nif and nif-associated genes are organized in nifE-nifN, nifX-orf, nifW-hesA-hesB and 'fdx'-containing operons, respectively. According to the results of this study and previous reports, the genes are expressed in a rhythmic pattern with peaks during the dark phase when the culture is grown in a 12 h light/12 h dark regimen. The rhythm persisted after the culture was transferred to continuous illumination.