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Alpha-tocopherol is essential for acquired chill-light tolerance in the cyanobacterium Synechocystis sp. strain PCC 6803.

ABSTRACT: Unlike Escherichia coli, the cyanobacterium Synechocystis sp. strain PCC 6803 is insensitive to chill (5 degrees C) in the dark but rapidly losses viability when exposed to chill in the light (100 micromol photons m(-2) s(-1)). Preconditioning at a low temperature (15 degrees C) greatly enhances the chill-light tolerance of Synechocystis sp. strain PCC 6803. This phenomenon is called acquired chill-light tolerance (ACLT). Preconditioned wild-type cells maintained a substantially higher level of alpha-tocopherol after exposure to chill-light stress. Mutants unable to synthesize alpha-tocopherol, such as slr1736, slr1737, slr0089, and slr0090 mutants, almost completely lost ACLT. When exposed to chill without light, these mutants showed no or a slight difference from the wild type. When complemented, the slr0089 mutant regained its ACLT. Copper-regulated expression of slr0090 from P(petE) controlled the level of alpha-tocopherol and ACLT. We conclude that alpha-tocopherol is essential for ACLT of Synechocystis sp. strain PCC 6803. The role of alpha-tocopherol in ACLT may be based largely on a nonantioxidant activity that is not possessed by other tocopherols or pathway intermediates.

SUBMITTER: Yang Y

PROVIDER: S-EPMC2258665 | biostudies-literature | 2008 Mar

REPOSITORIES: biostudies-literature

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Alpha-tocopherol is essential for acquired chill-light tolerance in the cyanobacterium Synechocystis sp. strain PCC 6803.

Yang Yang Y Yin Chuntao C Li Weizhi W Xu Xudong X

Journal of bacteriology 20071228 5

Unlike Escherichia coli, the cyanobacterium Synechocystis sp. strain PCC 6803 is insensitive to chill (5 degrees C) in the dark but rapidly losses viability when exposed to chill in the light (100 micromol photons m(-2) s(-1)). Preconditioning at a low temperature (15 degrees C) greatly enhances the chill-light tolerance of Synechocystis sp. strain PCC 6803. This phenomenon is called acquired chill-light tolerance (ACLT). Preconditioned wild-type cells maintained a substantially higher level of ...[more]

PMID: 18165303

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Project description:Two cAMP receptor proteins (CRPs), Sycrp1 (encoded by sll1371) and Sycrp2 (encoded by sll1924), exist in the cyanobacterium Synechocystis sp. strain PCC 6803. Previous studies have demonstrated that Sycrp1 has binding affinity for cAMP and is involved in motility by regulating the formation of pili. However, the function of Sycrp2 remains unknown. Here, we report that sycrp2 disruption results in the loss of motility of Synechocystis sp. PCC 6803, and that the phenotype can be recovered by reintroducing the sycrp2 gene into the genome of sycrp2-disrupted mutants. Electron microscopy showed that the sycrp2-disrupted mutant lost the pilus apparatus on the cell surface, resulting in a lack of cell motility. Furthermore, the transcript level of the pilA9-pilA11 operon (essential for cell motility and regulated by the cAMP receptor protein Sycrp1) was markedly decreased in sycrp2-disrupted mutants compared with the wild-type strain. Moreover, yeast two-hybrid analysis and a pulldown assay demonstrated that Sycrp2 interacted with Sycrp1 to form a heterodimer and that Sycrp1 and Sycrp2 interacted with themselves to form homodimers. Gel mobility shift assays revealed that Sycrp1 specifically binds to the upstream region of pilA9 Together, these findings indicate that in Synechocystis sp. PCC 6803, Sycrp2 regulates the formation of pili and cell motility by interacting with Sycrp1.IMPORTANCE cAMP receptor proteins (CRPs) are widely distributed in cyanobacteria and play important roles in regulating gene expression. Although many cyanobacterial species have two cAMP receptor-like proteins, the functional links between them are unknown. Here, we found that Sycrp2 in the cyanobacterium Synechocystis sp. strain PCC 6803 is essential for twitching motility and that it interacts with Sycrp1, a known cAMP receptor protein involved with twitching motility. Our findings indicate that the two cAMP receptor-like proteins in cyanobacteria do not have functional redundancy but rather work together.

Project description:BackgroundCyanobacteria are phototrophic prokaryotes that convert inorganic carbon as CO2 into organic compounds at the expense of light energy. They need only inorganic nutrients and can be cultivated to high densities using non-arable land and seawater. This has made cyanobacteria attractive organisms for the production of biofuels and chemical feedstock. Synechocystis sp. PCC 6803 is one of the most widely used cyanobacterial model strains. Based on its available genome sequence and genetic tools, Synechocystis has been genetically modified to produce different biotechnological products. Efficient isoprene production is an attractive goal because this compound is widely used as chemical feedstock.ResultsHere, we report on our attempts to generate isoprene-producing strains of Synechocystis using a plasmid-based strategy. As previously reported, a codon-optimized plant isoprene synthase (IspS) was expressed under the control of different Synechocystis promoters that ensure strong constitutive or light-regulated ispS expression. The expression of the ispS gene was quantified by qPCR and Western blotting, while the amount of isoprene was quantified using GC-MS. In addition to isoprene measurements in the headspace of closed culture vessels, single photon ionization time-of-flight mass spectrometry (SPI-MS) was applied, which allowed online measurements of isoprene production in open-cultivation systems under various conditions. Under standard conditions, a good correlation existed between ispS expression and isoprene production rate. The cultivation of isoprene production strains under NaCl-supplemented conditions decreased isoprene production despite enhanced ispS mRNA levels. The characterization of the metabolome of isoprene-producing strains indicated that isoprene production might be limited by insufficient precursor levels. Transcriptomic analysis revealed the upregulation of mRNA and regulatory RNAs characteristic of acclimation to metabolic stress.ConclusionsOur best production strains produced twofold higher isoprene amounts in the presence of low NaCl concentrations than previously reported strains. These results will guide future attempts to establish isoprene production in cyanobacterial hosts.

Project description:Every cyanobacterial species contains gene encoding site-2-protease (S2P) homolog. The studied prokaryotic S2P homologs play essential roles in regulating stress response through intramembrane proteolysis of membrane-bound anti-sigma factors. Here, gene of Slr0643, one of four S2P homologs in Synechocystis sp. PCC 6803, was insertionally disrupted to explore its physiological role. Only a partially segregated mutant was obtained, indicating its indispensability for growth. The partially disrupted mutant could not grow at pH 6.5, while wild type acclimated to pH 6.5 quickly. The slr0643 gene expression was transiently induced after pH transfer from 7.5 to 6.5. Both evidences demonstrated the pivotal role of fully functional Slr0643 in acid acclimation. DNA microarray and quantitative RT-PCR analyses decoded genes involved in early acid acclimation and revealed differentially expressed genes due to slr0643 disruption at both pH conditions. Early acid acclimation to pH 6.5 included upregulation of sigH, hik16 and hik35, and downregulation of pcrR and sigG; as well as downregulation of porins and upregulation of inorganic carbon and nitrogen transporters. Defective photosynthesis and excess expression of NADH dehydrogenase, together with over upregulation of carbon transporter and repression of nitrogen transporter and metabolism gene contributed to the acid lethality of mutant at pH 6.5. Most interestingly, analysis of microarray data revealed the close relationship between slr0643 disruption and expression of sigH operon. Therefore it was implied that Slr0643/Sll0857/SigH might acts through S2P/anti-Sigma factor/Sigma factor mechanism to play a role in acid acclimation. M-bM-^@M-^C Loop design was used to compare differential expression of wild type and mutant at pH 7.5 and pH 6.5 respectively, including dye-swape. Biological variation was sampled by extracting RNA from three independent experiments and pooling them together before hybridizations. Two or four replicate chips for each comparison and two replicate printings per chip represent technical repeats.

Dataset Information

Alpha-tocopherol is essential for acquired chill-light tolerance in the cyanobacterium Synechocystis sp. strain PCC 6803.

Publications

Alpha-tocopherol is essential for acquired chill-light tolerance in the cyanobacterium Synechocystis sp. strain PCC 6803.

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