Metabolomic analysis of the salt-sensitive mutants reveals changes in amino acid and fatty acid composition important to long-term salt stress in Synechocystis sp. PCC 6803.

Early studies in cyanobacteria have found that few genes induced by short-term salt shock (15-60 min) display a stable induction in the long-term (>1 day) salt-acclimated cells; meanwhile, most of the genes responsive to long-term salt stress were different from those by short-term salt shock, suggesting that different regulatory mechanisms may be involved for short-term and long-term salt stress responses. In our previous work using the model cyanobacterium Synechocystis sp. PCC 6803, sll1734 encoding CO2 uptake-related protein (CupA) and three genes encoding hypothetical proteins (i.e., ssr3402, slr1339, and ssr1853) were found induced significantly after a 3-day salt stress, and the corresponding gene knockout mutants were found salt sensitive. To further decipher the mechanisms that these genes may be involved, in this study, we performed a comparative metabolomic analysis of the wild-type Synechocystis and the four salt-sensitive mutants using a gas chromatography-mass spectrometry (GC-MS) approach. A metabolomic data set that consisted of 60 chemically classified metabolites was then subjected to a weighted correlation network analysis (WGCNA) to identify the metabolic modules and hub metabolites specifically related to each of the salt-stressed mutants. The results showed that two, one, zero, and two metabolic modules were identified specifically associated with the knockout events of sll1734, ssr3402, slr1339, and ssr1853, respectively. The mutant-associated modules included metabolites such as lysine and palmitic acid, suggesting that amino acid and fatty acid metabolisms are among the key protection mechanisms against long-term salt stresses in Synechocystis. The metabolomic results were further confirmed by quantitative reverse-transcription PCR analysis, which showed the upregulation of lysine and fatty acid synthesis-related genes. The study provided new insights on metabolic networks involved in long-term salt stress response in Synechocystis.