A genome-wide view of the expression and processing patterns of Thermus thermophilus HB8 CRISPR RNAs

Stefan Juranek, Tali Eban, Yael Altuvia*, Miguel Brown, Pavel Morozov, Thomas Tuschl, Hanah Margalit

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

31 Scopus citations

Abstract

The CRISPR-Cas system represents an RNA-based adaptive immune response system in prokaryotes and archaea. CRISPRs (clustered regularly interspaced short palindromic repeats) consist of arrays of short repeat-sequences interspaced by nonrepetitive short spacers, some of which show sequence similarity to foreign phage genetic elements. Their cistronic transcripts are processed to produce the mature CRISPR RNAs (crRNAs), the elements that confer immunity by base-pairing with exogenous nucleic acids. We characterized the expression and processing patterns of Thermus thermophilus HB8 CRISPRs by using differential deep-sequencing, which differentiates between 5′ monophosphate and 5′ non-monophosphate-containing RNAs and/or between 3′ hydroxyl and 3′ non-hydroxyl-containing RNAs. The genome of T. thermophilus HB8 encodes 11 CRISPRs, classified into three distinct repeat-sequence types, all of which were constitutively expressed without deliberately infecting the bacteria with phage. Analysis of the differential deep sequencing data suggested that crRNAs are generated by endonucleolytic cleavage, leaving fragments with 5′ hydroxyl and 3′ phosphate or 2′,3′-cyclic phosphate termini. The 5′ ends of all crRNAs are generated by site-specific cleavage 8 nucleotides upstream of the spacer first position; however, the 3′ ends are generated by two alternative, repeat-sequence-type-dependent mechanisms. These observations are consistent with the operation of multiple crRNA processing systems within a bacterial strain. Published by Cold Spring Harbor Laboratory Press.

Original languageAmerican English
Pages (from-to)783-794
Number of pages12
JournalRNA
Volume18
Issue number4
DOIs
StatePublished - Apr 2012

Keywords

  • Biogenesis
  • CRISPR
  • Deep-sequencing
  • Processing mechanism
  • crRNA

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