Physiological and Molecular Characterization of a Synechocystis sp. PCC 6803 Mutant Lacking Histidine Kinase Slr1759 and Response Regulator Slr1760

Abstract

The hybrid sensory histidine kinase Slr1759 of the cyanobacterium Synechocystis sp. strain PCC 6803 contains multiple sensory domains and a multi-step phosphorelay system. Immuno blot analysis provided evidence that the histidine kinase Slr1759 is associated with the cytoplasmic membrane. The gene slr1759 is part of an operon together with slr1760, encoding a response regulator. A comparative investigation was performed on Synechocystis sp. strain PCC 6803 wild type (WT) and an insertionally inactivated slr1759-mutant (Hik14) which also lacks the transcript for the response regulator Slr1760. The mutant Hik14 grew significantly slower than WT in the early growth phase, when both were inoculated with a low cell density into BG11 medium without additional buffer and when aerated with air enriched with 2% CO 2 . Since the aeration with CO 2 -enriched air results in a decrease of the pH value in the medium, the growth experiments indicated that Hik14 is not able to adjust its metabolic activities as rapidly as WT to compensate for a larger decrease of the pH value in the medium. No significant differences in growth between Hik14 and WT were observed when cells were inoculated with a higher cell density in BG11 medium or when the BG11 medium contained 50 mm Epps-NaOH, pH 7.5, to prevent the pH drop. This Hik14 phenotype has so far only been seen under the above defined growth condition. Results of photosynthetic activity measurements as well as Northern blot-, immuno blot-, and metabolite analyses suggest that the two-component system Slr1759/Slr1760 has a function in the coordination of several metabolic activities which is in good agreement with the complex domain structure of Slr1759. The direct targets of this two-component system have so far not been identified.

The hybrid histidine kinase Slr1759 of the cyanobacterium Synechocystis sp. PCC 6803 contains FAD at its PAS domain

“…Indeed, upon subcellular fractionation of Synechocystis sp. PCC 6803 cells Slr1759 was found associated with the cytoplasmic membrane consistent with a potential involvement in pH sensing (Nodop et al 2006).…”

“…Thus, the mutant presumably is less able to cope with acidiWcation of the medium and the more energy-demanding metabolisation of CO 2 . Further characterization of the mutant pointed to an enhanced respiratory activity and a slightly reduced photosynthetic activity, implicating Slr1759 in the coordination of carbon assimilation, pH homeostasis, respiration and photosynthesis (Nodop et al 2006).…”

“…PCC 6803 harbours 47 genes encoding predicted Hiks many of which are implicated in stress reactions (Mizuno et al 1996;Murata and Suzuki 2006) including responses to cold stress , high salt (Marin et al 2003), high osmolarity (Mikami et al 2002), heavy metals (LopezMaury et al 2002) or low phosphate (Hirani et al 2001), as well as in manganese homeostasis (Ogawa et al 2002;Yamaguchi et al 2002) or phototaxis (Shin et al 2008). We have previously characterized the slr1759 mutant constructed in the Murata laboratory (Suzuki et al 2000;Nodop et al 2006). It is insertionally inactivated in slr1759 encoding the hybrid histidine kinase Hik14 and also does not express slr1760 encoding a response regulator that is part of an operon with slr1759 (Nodop et al 2006).…”

“…We have previously characterized the slr1759 mutant constructed in the Murata laboratory (Suzuki et al 2000;Nodop et al 2006). It is insertionally inactivated in slr1759 encoding the hybrid histidine kinase Hik14 and also does not express slr1760 encoding a response regulator that is part of an operon with slr1759 (Nodop et al 2006). Thus, Slr1759 and Slr1760 presumably represent a two-component system encoded by linked genes.…”

Detection of an L-amino acid dehydrogenase activity in Synechocystis sp. PCC 6803

“…Growth was performed with a slightly modified BG11 medium ( Stephan et al , 2000 ) with nitrate as the N-source (17.7 mM sodium nitrate) or with nitrate-free BG11 medium containing 5 mM L -arginine-HCl to which 50 mM EPPS-NaOH pH 7.5 was added to prevent acidification. After 48 h of growth with nitrate or L -arginine the pH of the CO 2 -aerated BG11 medium was between 7.0 and 7.5 ( Nodop et al , 2006 ). Growth of the PsbO-free mutant was performed as described above, except that the growth medium contained kanamycin sulphate (7.5 mg l −1 ).…”