Abstract
BackgroundAll archaeal and many bacterial genomes contain Clustered Regularly Interspaced Short Palindrome Repeats (CRISPR) and variable arrays of the CRISPR-associated (cas) genes that have been previously implicated in a novel form of DNA repair on the basis of comparative analysis of their protein product sequences. However, the proximity of CRISPR and cas genes strongly suggests that they have related functions which is hard to reconcile with the repair hypothesis.ResultsThe protein sequences of the numerous cas gene products were classified into ~25 distinct protein families; several new functional and structural predictions are described. Comparative-genomic analysis of CRISPR and cas genes leads to the hypothesis that the CRISPR-Cas system (CASS) is a mechanism of defense against invading phages and plasmids that functions analogously to the eukaryotic RNA interference (RNAi) systems. Specific functional analogies are drawn between several components of CASS and proteins involved in eukaryotic RNAi, including the double-stranded RNA-specific helicase-nuclease (dicer), the endonuclease cleaving target mRNAs (slicer), and the RNA-dependent RNA polymerase. However, none of the CASS components is orthologous to its apparent eukaryotic functional counterpart. It is proposed that unique inserts of CRISPR, some of which are homologous to fragments of bacteriophage and plasmid genes, function as prokaryotic siRNAs (psiRNA), by base-pairing with the target mRNAs and promoting their degradation or translation shutdown. Specific hypothetical schemes are developed for the functioning of the predicted prokaryotic siRNA system and for the formation of new CRISPR units with unique inserts encoding psiRNA conferring immunity to the respective newly encountered phages or plasmids. The unique inserts in CRISPR show virtually no similarity even between closely related bacterial strains which suggests their rapid turnover, on evolutionary scale. Corollaries of this finding are that, even among closely related prokaryotes, the most commonly encountered phages and plasmids are different and/or that the dominant phages and plasmids turn over rapidly.ConclusionWe proposed previously that Cas proteins comprise a novel DNA repair system. The association of the cas genes with CRISPR and, especially, the presence, in CRISPR units, of unique inserts homologous to phage and plasmid genes make us abandon this hypothesis. It appears most likely that CASS is a prokaryotic system of defense against phages and plasmids that functions via the RNAi mechanism. The functioning of this system seems to involve integration of fragments of foreign genes into archaeal and bacterial chromosomes yielding heritable immunity to the respective agents. However, it appears that this inheritance is extremely unstable on the evolutionary scale such that the repertoires of unique psiRNAs are completely replaced even in closely related prokaryotes, presumably, in response to rapidly changing repertoires of dominant phages and plasmids.This article was reviewed by: Eric Bapteste, Patrick Forterre, and Martijn Huynen.Open peer reviewReviewed by Eric Bapteste, Patrick Forterre, and Martijn Huynen.For the full reviews, please go to the Reviewers' comments section.
Highlights
All archaeal and many bacterial genomes contain Clustered Regularly Interspaced Short Palindrome Repeats (CRISPR) and variable arrays of the CRISPR-associated genes that have been previously implicated in a novel form of DNA repair on the basis of comparative analysis of their protein product sequences
The functioning of this system seems to involve integration of fragments of foreign genes into archaeal and bacterial chromosomes yielding heritable immunity to the respective agents. It appears that this inheritance is extremely unstable on the evolutionary scale such that the repertoires of unique prokaryotic siRNAs (psiRNA) are completely replaced even in closely related prokaryotes, presumably, in response to rapidly changing repertoires of dominant phages and plasmids
There are two major, distinct forms of regulatory small RNAs involved in eukaryotic gene silencing: small interfering RNAs and micro RNAs. siRNAs are produced from doublestranded RNAs of viruses and transposable elements, which are processed by the dicer nuclease, one of the essential components of the RNA-Induced Silencing Complexes (RISCs) [7,8,9]
Summary
All archaeal and many bacterial genomes contain Clustered Regularly Interspaced Short Palindrome Repeats (CRISPR) and variable arrays of the CRISPR-associated (cas) genes that have been previously implicated in a novel form of DNA repair on the basis of comparative analysis of their protein product sequences. There are two major, distinct forms of regulatory small RNAs involved in eukaryotic gene silencing: small interfering (si) RNAs and micro (mi) RNAs. siRNAs are produced from doublestranded RNAs of viruses and transposable elements, which are processed by the dicer nuclease, one of the essential components of the RNA-Induced Silencing Complexes (RISCs) [7,8,9]. Unlike the siRNAs, 21–25 nt-long miRNAs are encoded in eukaryotic genomes and are either perfectly (in plants) or imperfectly (in animals) complementary to sequences in the 3'-untranslated regions of specific endogenous mRNAs [6,13]. Base-pairing of miRNAs with the target mRNAs, which is mediated by a distinct form of RISC, results either in RNA cleavage or in down-regulation of translation [8]. Evidence is rapidly accumulating that numerous, probably, thousands of miRNAs in animals and plants are major players in development regulation and chromatin remodeling [6]
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