An Active Type I-E CRISPR-Cas System Identified in Streptomyces avermitilis.

Yi Qiu,Shiwei Wang,Zhi Chen,Yuan Song,Yajie Guo,Erika Kothe

doi:10.1371/journal.pone.0149533

Abstract

CRISPR-Cas systems, the small RNA-dependent immune systems, are widely distributed in prokaryotes. However, only a small proportion of CRISPR-Cas systems have been identified to be active in bacteria. In this work, a naturally active type I-E CRISPR-Cas system was found in Streptomyces avermitilis. The system shares many common genetic features with the type I-E system of Escherichia coli, and meanwhile shows unique characteristics. It not only degrades plasmid DNA with target protospacers, but also acquires new spacers from the target plasmid DNA. The naive features of spacer acquisition in the type I-E system of S. avermitilis were investigated and a completely conserved PAM 5’-AAG-3’ was identified. Spacer acquisition displayed differential strand bias upstream and downstream of the priming spacer, and irregular integrations of new spacers were observed. In addition, introduction of this system into host conferred phage resistance to some extent. This study will give new insights into adaptation mechanism of the type I-E systems in vivo, and meanwhile provide theoretical foundation for applying this system on the genetic modification of S. avermitilis.

Highlights

Bacteria can be found nearly everywhere even in some harsh environments with the risk of predatory viruses and potentially harmful plasmids
Intense investigations have been performed to reveal the molecular mechanisms of the type I-E CRISPR-Cas system in E. coli
Activity of this system is repressed in wild-type E. coli, hindering studies of the inherent function of type I-E CRISPRCas systems in this species

Summary

Introduction

Bacteria can be found nearly everywhere even in some harsh environments with the risk of predatory viruses and potentially harmful plasmids. CRISPR loci typically comprise clustered, noncontiguous direct repeats interspaced by variable sequences called spacers, and are frequently flanked by CRISPR-associated (cas) genes. CRISPR-Cas systems are widespread in bacteria and archaea, and are classified into three major types (type I, II and III) and 12 subtypes (I-A, I-B, etc.) according to the difference of cas gene contents and defense pathways across species [3]. Mechanisms of adaptive immunity mediated by CRISPR-Cas systems are intriguing and the subtype I-E CRISPR-Cas system in E. coli has been extensively studied [4, 5]. CRISPR loci in E. coli comprise multiple palindromic repeats of 29 nucleotides separated by variable spacers of 32 or 33 nucleotides [6]. Leader sequences flanking one side of each CRISPR locus appear to PLOS ONE | DOI:10.1371/journal.pone.0149533 February 22, 2016

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