Dataset Information

CemA homologue essential to CO2 transport in the cyanobacterium Synechocystis PCC6803.

ABSTRACT: We have isolated mutants of Synechocystis PCC6803 that grew very slowly in a low-sodium medium, which is unfavorable for HCO3(-) transport, and examined two of these mutants (SC1 and SC2) for their ability to take up CO2 and HCO3(-) in the light. The CO2 transport activity of SC1 and SC2 was much lower than that of the wild type (WT), whereas there was no difference between the mutants and the WT in their activity of HCO3(-) transport. A clone containing a 3.9-kilobase-pair insert DNA that transforms both mutants to the WT phenotype was isolated from a genomic library of WT Synechocystis. Sequencing of the insert DNA in the region of mutations in SC1 and SC2 revealed an open reading frame (designated cotA), which showed significant amino-acid sequence homology to cemA encoding a protein found in the inner envelope membrane of chloroplasts. The cotA gene is present in a single copy and was not cotranscribed with any other gene(s). cotA encodes a protein of 247 amino acids containing four transmembrane domains. There was substitution of a single base in SC1 and two bases in SC2 in their cotA genes. A possible role of the cotA gene product in CO2 transport is discussed.

SUBMITTER: Katoh A

PROVIDER: S-EPMC39476 | biostudies-other | 1996 Apr

REPOSITORIES: biostudies-other

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cemA homologue essential to CO2 transport in the cyanobacterium Synechocystis PCC6803.

Katoh A A Lee K S KS Fukuzawa H H Ohyama K K Ogawa T T

Proceedings of the National Academy of Sciences of the United States of America 19960401 9

We have isolated mutants of Synechocystis PCC6803 that grew very slowly in a low-sodium medium, which is unfavorable for HCO3(-) transport, and examined two of these mutants (SC1 and SC2) for their ability to take up CO2 and HCO3(-) in the light. The CO2 transport activity of SC1 and SC2 was much lower than that of the wild type (WT), whereas there was no difference between the mutants and the WT in their activity of HCO3(-) transport. A clone containing a 3.9-kilobase-pair insert DNA that trans ...[more]

PMID: 8633006

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Project description:BACKGROUND:The production of biofuels in photosynthetic microalgae and cyanobacteria is a promising alternative to the generation of fuels from fossil resources. To be economically competitive, producer strains need to be established that synthesize the targeted product at high yield and over a long time. Engineering cyanobacteria into forced fuel producers should considerably interfere with overall cell homeostasis, which in turn might counteract productivity and sustainability of the process. Therefore, in-depth characterization of the cellular response upon long-term production is of high interest for the targeted improvement of a desired strain. RESULTS:The transcriptome-wide response to continuous ethanol production was examined in Synechocystis sp. PCC6803 using high resolution microarrays. In two independent experiments, ethanol production rates of 0.0338% (v/v) ethanol d-1 and 0.0303% (v/v) ethanol d-1 were obtained over 18 consecutive days, measuring two sets of biological triplicates in fully automated photobioreactors. Ethanol production caused a significant (~40%) delay in biomass accumulation, the development of a bleaching phenotype and a down-regulation of light harvesting capacity. However, microarray analyses performed at day 4, 7, 11 and 18 of the experiment revealed only three mRNAs with a strongly modified accumulation level throughout the course of the experiment. In addition to the overexpressed adhA (slr1192) gene, this was an approximately 4 fold reduction in cpcB (sll1577) and 3 to 6 fold increase in rps8 (sll1809) mRNA levels. Much weaker modifications of expression level or modifications restricted to day 18 of the experiment were observed for genes involved in carbon assimilation (Ribulose bisphosphate carboxylase and Glutamate decarboxylase). Molecular analysis of the reduced cpcB levels revealed a post-transcriptional processing of the cpcBA operon mRNA leaving a truncated mRNA cpcA* likely not competent for translation. Moreover, western blots and zinc-enhanced bilin fluorescence blots confirmed a severe reduction in the amounts of both phycocyanin subunits, explaining the cause of the bleaching phenotype. CONCLUSIONS:Changes in gene expression upon induction of long-term ethanol production in Synechocystis sp. PCC6803 are highly specific. In particular, we did not observe a comprehensive stress response as might have been expected.

Project description:The production of biofuels in photosynthetic microalgae and cyanobacteria is considered a promising alternative to the generation of fuels from fossil resources. However, to be economically competitive, producer strains need to be established that synthesize the targeted product at high yield and over a long time. Engineering cyanobacteria to forced fuel producers should considerably interfere with overall cell homeostasis, which in turn might counteract productivity and sustainability of the process. Therefore, in-depth characterization of the cellular response upon long-term production is of high interest for the targeted improvement of a desired strain. Here we report the results of a monitoring experiment, in which the transcriptome-wide response to continuous ethanol production in the unicellular model cyanobacterium Synechocystis sp. PCC6803 was examined using high resolution microarrays. In two independent experiments, ethanol production rates of 0.0338% (v/v) EtOH d-1 M-BM-1 0.002 and 0.0303% (v/v) d-1 M-BM-1 0.002 were obtained over 18 consecutive days, measuring biological triplicates in fully automated photobioreactors. Ethanol production caused a significant (~40%) delay in biomass accumulation in the producer strains and the development of a bleaching phenotype. Absorption spectroscopy indicated in particular a down-regulation of light harvesting capacity. Microarray analyses performed at day 4, 7, 11 and 18 of the experiment revealed only three mRNAs with a strongly modified accumulation level throughout the course of the experiment. In addition to the overexpressed adhA (slr1192) gene, this was an about 4 fold reduction in cpcB (sll1577) and an about 3 to 6 fold increase in rps8 (sll1809) mRNA levels. Much weaker modifications of expression level or modifications restricted to day 18 of the experiment were observed for genes involved in carbon assimilation (Ribulose bisphosphate carboxylase and Glutamate decarboxylase). Molecular analysis of the reduced cpcB levels revealed a post-transcriptional operon discoordination in the cpcBA operon leaving a truncated mRNA cpcA* likely not competent for translation. Moreover, Western blots and zinc-enhanced bilin fluorescence blots confirmed a severe reduction in the amounts of both phycocyanin subunits, explaining the cause of the bleaching phenotype. We conclude that the changes in gene expression upon induction of long term ethanol production in Synechocystis sp. PCC6803 are highly specific. in particular we did not observe a comprehensive stress response contributing to a complex phenotype as might have been expected. Gene expression of Synechocystis sp. PCC 6803 WT (#621) and the isogenic ethanol producing strain #309 was monitored at 4, 7, 11 and 18 days after induction of ethanol production by copper depletion. Each condition was sampled in biological duplicates

Dataset Information

CemA homologue essential to CO2 transport in the cyanobacterium Synechocystis PCC6803.

Publications

cemA homologue essential to CO2 transport in the cyanobacterium Synechocystis PCC6803.

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