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Analysis of a genomic DNA region from the cyanobacterium Synechococcus sp. strain PCC7942 involved in carboxysome assembly and function.

ABSTRACT: We report on the sequencing and analysis of a 3,557-bp genomic DNA clone that is located between 4.8 and 1.2 kilobase pairs (kb) upstream of the rbcL gene and is capable of complementing a class of cyanobacterium Synechococcus sp. strain PCC7942 mutants requiring a high level of CO2. The upstream 2,704 bp of this sequence is novel, the remaining 852 bp having been reported by other workers. Four new open reading frames (ORFs) have been identified along with putative promoter elements. These ORFs, which could code for proteins of 7, 10.9, 11, and 58 kDa in size, have been named ORF 64, ccmK, ccmL, and ccmM, respectively. The last three have been named ccm genes on the basis that insertional mutagenesis of each produces a phenotype requiring a high level of CO2 (i.e., each produces a lesion in the CO2 concentrating mechanism). The putative gene product for the large ccmM ORF has three internally repeated regions and also has two possible DNA binding motifs. Two defined mutants in the 3,557-bp region, mutants PVU and P-N, have been more fully characterized. The PVU mutant has a drug marker inserted into the ccmL gene, and it possesses abnormal rod-shaped carboxysomes. The P-N mutant is a 2.64-kb deletion of DNA from the same position in ccmL to a region closer to rbcL. This mutant, which has previously been shown to lack carboxysomes and have soluble ribulosebiphosphate carboxylase/oxygenase activity, has now been shown to have a predominantly soluble carboxysomal carbonic anhydrase activity. Both mutants were found to possess carboxysomal carbonic anhydrase activities which are below wild-type levels, and in the P-N mutant this activity appears to be unstable. The results are discussed in terms of the possible interactions of putative ccm gene products in the process of carboxysome assembly and function.

SUBMITTER: Price GD

PROVIDER: S-EPMC204604 | biostudies-other | 1993 May

REPOSITORIES: biostudies-other

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Analysis of a genomic DNA region from the cyanobacterium Synechococcus sp. strain PCC7942 involved in carboxysome assembly and function.

Price G D GD Howitt S M SM Harrison K K Badger M R MR

Journal of bacteriology 19930501 10

We report on the sequencing and analysis of a 3,557-bp genomic DNA clone that is located between 4.8 and 1.2 kilobase pairs (kb) upstream of the rbcL gene and is capable of complementing a class of cyanobacterium Synechococcus sp. strain PCC7942 mutants requiring a high level of CO2. The upstream 2,704 bp of this sequence is novel, the remaining 852 bp having been reported by other workers. Four new open reading frames (ORFs) have been identified along with putative promoter elements. These ORFs ...[more]

PMID: 8491708

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Project description:RNA degradation is an important process that influences the ultimate concentration of individual proteins inside cells. While the main enzymes that facilitate this process have been identified, global maps of RNA turnover are available for only a few species. Even in these cases, there are few sequence elements that are known to enhance or destabilize a native transcript; even fewer confer the same effect when added to a heterologous transcript. To address this knowledge gap, we assayed genome-wide RNA degradation in the cyanobacterium Synechococcus sp. strain PCC 7002 by collecting total RNA samples after stopping nascent transcription with rifampin. We quantified the abundance of each position in the transcriptome as a function of time using RNA-sequencing data and later analyzed the global mRNA decay map using machine learning principles. Half-lives, calculated on a per-ORF (open reading frame) basis, were extremely short, with a median half-life of only 0.97 min. Despite extremely rapid turnover of most mRNA, transcripts encoding proteins involved in photosynthesis were both highly expressed and highly stable. Upon inspection of these stable transcripts, we identified an enriched motif in the 3' untranslated region (UTR) that had similarity to Rho-independent terminators. We built statistical models for half-life prediction and used them to systematically identify sequence motifs in both 5' and 3' UTRs that correlate with stabilized transcripts. We found that transcripts linked to a terminator containing a poly(U) tract had a longer half-life than both those without a poly(U) tract and those without a terminator.IMPORTANCE RNA degradation is an important process that affects the final concentration of individual mRNAs, affecting protein expression and cellular physiology. Studies of how RNA is degraded increase our knowledge of this fundamental process as well as enable the creation of genetic tools to manipulate RNA stability. By studying global transcript turnover, we searched for sequence elements that correlated with transcript (in)stability and used these sequences to guide tool design. This study probes global RNA turnover in a cyanobacterium, Synechococcus sp. strain PCC 7002, that both has a unique array of RNases that facilitate RNA degradation and is an industrially relevant strain that could be used to convert CO2 and sunlight into useful products.

Dataset Information

Analysis of a genomic DNA region from the cyanobacterium Synechococcus sp. strain PCC7942 involved in carboxysome assembly and function.

Publications

Analysis of a genomic DNA region from the cyanobacterium Synechococcus sp. strain PCC7942 involved in carboxysome assembly and function.

Similar Datasets

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