Transcript analysis of the extended hyp-operon in the cyanobacteria Nostoc sp. strain PCC 7120 and Nostoc punctiforme ATCC 29133

Abstract

BackgroundCyanobacteria harbor two [NiFe]-type hydrogenases consisting of a large and a small subunit, the Hup- and Hox-hydrogenase, respectively. Insertion of ligands and correct folding of nickel-iron hydrogenases require assistance of accessory maturation proteins (encoded by the hyp-genes). The intergenic region between the structural genes encoding the uptake hydrogenase (hupSL) and the accessory maturation proteins (hyp genes) in the cyanobacteria Nostoc PCC 7120 and N. punctiforme were analysed using molecular methods.FindingsThe five ORFs, located in between the uptake hydrogenase structural genes and the hyp-genes, can form a transcript with the hyp-genes. An identical genomic localization of these ORFs are found in other filamentous, N2-fixing cyanobacterial strains. In N. punctiforme and Nostoc PCC 7120 the ORFs upstream of the hyp-genes showed similar transcript level profiles as hupS (hydrogenase structural gene), nifD (nitrogenase structural gene), hypC and hypF (accessory hydrogenase maturation genes) after nitrogen depletion. In silico analyzes showed that these ORFs in N. punctiforme harbor the same conserved regions as their homologues in Nostoc PCC 7120 and that they, like their homologues in Nostoc PCC 7120, can be transcribed together with the hyp-genes forming a larger extended hyp-operon. DNA binding studies showed interactions of the transcriptional regulators CalA and CalB to the promoter regions of the extended hyp-operon in N. punctiforme and Nostoc PCC 7120.ConclusionsThe five ORFs upstream of the hyp-genes in several filamentous N2-fixing cyanobacteria have an identical genomic localization, in between the genes encoding the uptake hydrogenase and the maturation protein genes. In N. punctiforme and Nostoc PCC 7120 they are transcribed as one operon and may form transcripts together with the hyp-genes. The expression pattern of the five ORFs within the extended hyp-operon in both Nostoc punctiforme and Nostoc PCC 7120 is similar to the expression patterns of hupS, nifD, hypF and hypC. CalA, a known transcription factor, interacts with the promoter region between hupSL and the five ORFs in the extended hyp-operon in both Nostoc strains.

A HupS-GFP fusion protein demonstrates a heterocyst-specific localization of the uptake hydrogenase in Nostoc punctiforme

“…After 24 h of combined nitrogen starvation, a homogeneous and weak GFP fluorescence could be observed from proheterocysts. This is in agreement with the observed transcription of hupS in differentiating heterocysts of the same strain of N. punctiforme (Holmqvist, 2010). After approximately 30–34 h of nitrogen starvation, and up to 7 days, several smaller or fewer larger clusters of GFP fluorescence could be observed within the heterocysts.…”

“…Upon deprivation of combined nitrogen, approximately every 10th–20th cell, evenly distributed in a filament of Nostoc , differentiates into a heterocyst. During the heterocyst development, the transcription of the nif genes, encoding the nitrogenase, and the hup genes, encoding the uptake hydrogenase, take place (Elhai & Wolk, 1990; Axelsson et al , 1999; Holmqvist, 2010). The uptake hydrogenase in cyanobacteria consists of a small (HupS) and a large (HupL) subunit, encoded by hupS and hupL , respectively.…”

CO-dependent hydrogen production by the facultative anaerobe Parageobacillus thermoglucosidasius

“…These include a tetratricopeptide-repeat (PhbH) and NHL repeat (PhbK) containing protein, which also occur in the [Ni-Fe] group 2a hydrogenase loci of Nostoc punctiforme ATCC 29133 and Nostoc sp. PCC 7120, where they are co-transcribed with the hydrogenase genes and have been suggested to play a role in protein–protein interactions and Fe–S cluster biogenesis (PhbJ) which may mediate electron transport to redox partners in downstream reactions [ 30 ].…”