Analysis of Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) systems in the cyanobacterium Synechocystis sp. PCC 6803

Cyanobacteria are oxygenic phototrophs which inhabit both terrestrial and aquatic environments, where they encounter a high concentration of bacteriophages. This gives rise to an evolutionary pressure for effective defense mechanisms. The genome of the freshwater model organism Synechocystis sp. PCC 6803 consists of a circular chromosome and 7 plasmids. We have characterized the primary transcriptome of Synechocystis and in the course of this study it turned out that a particular class of abundant transcripts originates from the 103 kb plasmid pSYSA.

pSYSA contains three Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) loci. This class of genetic element constitute an adaptive and inheritable defense system which protects bacteria and archaea from infection by phages.

We show that for each of the three CRISPRs, long precursor RNAs are transcribed and processed by independent endoribonucleases, leading to the accumulation of very characteristic sRNAs. The three CRISPRs differ in their nature: CRISPR3 is the transcriptionally most active locus, whereas CRISPR2 responds to environmental stimuli and CRISPR1 may be considered as an archetypical copy. Rifampicin experiments revealed distinct differences between the half-lifes of transcripts from the first two CRISPRs and CRISPR3. We have characterized individual mutants for each of the three CRISPR regions and 13 CRISPR-associated proteins.

As there are profound differences in the CRISPR-associated protein apparatus between archaea and bacterial species studied as models thus far, it is interesting that the cyanobacterial CRISPRs are unique in combining features known only from archaea with an otherwise bacterial CRISPR system.

The biological relevance of plasmid pSYSA seems to be associated with an important defense mechanism, making the plasmid indispensable for viability. Furthermore toxin-antitoxin-systems encoded on the plasmid ensure maintenance of the plasmid in the cell.