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

The ability of cyanobacteria to capture solar energy, via oxygenic photosynthesis, and convert that energy to molecular hydrogen (H 2 ) has made them an interesting group of organisms with potential as future energy producers. There are three types of enzymes directly involved in the cyanobacterial hydrogen metabolism; nitrogenases that produce H 2 as a by-product when fixating atmospheric nitrogen, uptake hydrogenases that catalyze the oxidation of H 2 ,thereby preventing energy losses from the cells, and bidirectional hydrogenases that has the capacity to both oxidize and reduce H 2 . Hydrogenases are complex metalloenzymes, and the insertion of ligands and correct folding of the proteins require assistance of accessory proteins, the Hyp proteins. Cyanobacterial hydrogenases are NiFe-type hydrogenases and consist of a large and a small subunit. Today, the maturation process of the large subunit has been uncovered to a large extent in cyanobacteria, mostly by analogy assumptions from studies done in other bacteria such as Escherichia coli but also from mutational analyses in cyanobacteria, while the maturation process of the small subunit is still unknown. Recently a set of genes, putatively involved in the maturation process of the small subunit, was discovered in Nostoc sp. PCC 7120 and Nostoc punctiforme ATCC 29133. These five ORFs, encoding unknown proteins, are located in between the uptake hydrogenase structural genes and the hyp -genes were shown to be transcribed together with the hyp -genes in Nostoc PCC 7120. The ORFs upstream the hyp -genes can be found in the same genomic arrangement in other filamentous, nitrogen fixing cyanobacterial strains but are interestingly missing in strains incapable of nitrogen fixation. In this study we have further investigated the function of the ORFs upstream the hyp -genes by studying their transcription pattern after nitrogen depletion in the filamentous, nitrogen fixing strains Nostoc PCC 7120 and N. punctiforme . The transcription pattern were compared to the transcription pattern of hupS and hoxY , encoding the uptake and bidirectional hydrogenase small subunits, nifD , encoding a nitrogenase subunit and hypC and hypF , encoding the maturation process accessory proteins HypC and HypF. All the five ORFs upstream the hyp -genes, in both organisms, were upregulated after nitrogen step down in accordance with the transcription pattern for hupS , nifD , hypC and hypF which support the theory that these genes mig