Transcriptional Analysis of Hydrogenase Genes in the Cyanobacteria Anacystis nidulans and Anabaena variabilis Monitored by RT-PCR

Abstract

Diverse cyanobacteria express an uptake hydrogenase, encoded by the genes hupSL, and a bidirectional, NAD(P)(+)-reducing hydrogenase with the genes hox(E)FUYH. In the unicellular Anacystis nidulans, the hox genes are organized on two separate loci, whereas they are contiguous in one cluster, though interspersed with two unidentified reading frames, ORF 3 and 8, in the heterocystous Anabaena variabilis. The hox gene clusters of these two cyanobacteria have now been transcriptionally analyzed by RT-PCR. A polycistronic transcript was identified in both cyanobacteria. In A. nidulans, one message for each locus has been detected, the dicistronic hoxEF unit, and the polycistronic hoxUYHWhypAB one. In A. variabilis, the transcript consists of the hox genes hoxFUYH as well as the unidentified ORFs. Previous enzyme determinations on the occurrence of the uptake hydrogenase in vegetative cells and thus outside of heterocysts gave ambiguous results. Therefore, transcription of both hup and hox genes has been analyzed in both heterocysts and vegetative cells of A. variabilis. A hupL transcript is detectable in heterocysts and also, though less extensive but clearly discernible, in vegetative cells of NH(4)(+)-grown A. variabilis.

Hydrogenases and Hydrogen Metabolism of Cyanobacteria

“…4) (17). In contrast to these results, a low level of hupL expression (detected by RT-PCR) has been found in A. variabilis ATCC 29413 vegetative cells grown with the addition of ammonia (28). Interest- ingly, when exposed to anaerobic conditions A. variabilis synthesizes a second Mo nitrogenase cellularly localized in the vegetative cells (199).…”

“…In contrast, in the unicellular Synechococcus strain PCC 6301 two transcripts were detected, the dicistronic hoxEF and the polycistronic hoxUYHWhypAB (28). Further promoter activities were identified in the hox locus by using ␤-galactosidase and bacterial luciferase as reporters (28). With the exception of a shift from non-nitrogen-fixing to nitrogen-fixing conditions, which was associated with no significant change in the transcript level of hoxH in N. muscorum (Fig.…”

“…An ORF positioned upstream of hoxF and termed hoxE has been identified and sequenced in Synechococcus strain PCC 6301 and Synechocystis strain PCC 6803, and is also present in Anabaena strain PCC 7120 (http://www.kazusa.or.jp./cyano/anabaena). It has been suggested to encode a third diaphorase subunit (28). Moreover, in Synechococcus strain PCC 6301, an ORF termed hoxW is localized downstream of hoxH, putatively encoding a protease ( Fig.…”

“…Transcriptional studies (using RT-PCR) indicate that the structural genes form a single transcriptional unit in A. variabilis ATCC 29413 (28). In contrast, in the unicellular Synechococcus strain PCC 6301 two transcripts were detected, the dicistronic hoxEF and the polycistronic hoxUYHWhypAB (28). Further promoter activities were identified in the hox locus by using ␤-galactosidase and bacterial luciferase as reporters (28).…”

Characterization and transcriptional analysis of hupSLW in Gloeothece sp. ATCC 27152: an uptake hydrogenase from a unicellular cyanobacterium

“…Subsequent studies, in other filamentous strains, have confirmed the induction of an hupL transcript under nitrogen-fixing conditions only (Axelsson et al, 1999;Happe et al, 2000;Hansel et al, 2001). One exception is Anabaena variabilis ATCC 29413, where a low level of hupL expression has been detected in vegetative cells grown with the addition of ammonia (Boison et al, 2000). In all the cyanobacterial strains examined so far, transcriptional studies have shown that the hupSL genes constitute a single transcript, containing no additional ORFs (Happe et al, 2000;Lindberg et al, 2000).…”