The Cas6e ribonuclease is not required for interference and adaptation by the E. coli type I-E CRISPR-Cas system

Ekaterina Semenova; Konstantin Kuznedelov; Kirill A Datsenko; Pierre M Boudry; Ekaterina E Savitskaya; Sofia Medvedeva; Natalia Beloglazova; Maria Logacheva; Alexander F Yakunin; Konstantin Severinov

doi:10.1093/nar/gkv546

The Cas6e ribonuclease is not required for interference and adaptation by the E. coli type I-E CRISPR-Cas system

Nucleic Acids Res. 2015 Jul 13;43(12):6049-61. doi: 10.1093/nar/gkv546. Epub 2015 May 26.

Affiliations

¹ Waksman Institute of Microbiology, Rutgers, the State University of New Jersey, Piscataway, NJ 08854, USA.
² Waksman Institute of Microbiology, Rutgers, the State University of New Jersey, Piscataway, NJ 08854, USA Department of Biological Sciences, Purdue University, West Lafayette, IN 47907, USA.
³ Waksman Institute of Microbiology, Rutgers, the State University of New Jersey, Piscataway, NJ 08854, USA Laboratoire Pathogenèse des Bactéries Anaérobies, Institut Pasteur, Université Paris Diderot, Sorbonne Paris Cité, Cellule Pasteur, 25 rue du Dr Roux, 75724 Paris Cedex 15, France.
⁴ Skolkovo Institute of Science and Technology, Skolkovo 143025, Russia Institute of Molecular Genetics, Russian Academy of Sciences, Moscow 123182, Russia.
⁵ Skolkovo Institute of Science and Technology, Skolkovo 143025, Russia Institute of Gene Biology, Russian Academy of Sciences, Moscow 119334, Russia.
⁶ Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, Ontario M5S 3E5, Canada.
⁷ Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119234 Moscow, Russia Pirogov Russian National Research Medical University, 117997 Moscow, Russia.
⁸ Waksman Institute of Microbiology, Rutgers, the State University of New Jersey, Piscataway, NJ 08854, USA Skolkovo Institute of Science and Technology, Skolkovo 143025, Russia Institute of Molecular Genetics, Russian Academy of Sciences, Moscow 123182, Russia Institute of Gene Biology, Russian Academy of Sciences, Moscow 119334, Russia Peter the Great St. Petersburg Polytechnic University, St. Petersburg 195251, Russia severik@waksman.rutgers.edu.

Abstract

CRISPR-Cas are small RNA-based adaptive prokaryotic immunity systems protecting cells from foreign DNA or RNA. Type I CRISPR-Cas systems are composed of a multiprotein complex (Cascade) that, when bound to CRISPR RNA (crRNA), can recognize double-stranded DNA targets and recruit the Cas3 nuclease to destroy target-containing DNA. In the Escherichia coli type I-E CRISPR-Cas system, crRNAs are generated upon transcription of CRISPR arrays consisting of multiple palindromic repeats and intervening spacers through the function of Cas6e endoribonuclease, which cleaves at specific positions of repeat sequences of the CRISPR array transcript. Cas6e is also a component of Cascade. Here, we show that when mature unit-sized crRNAs are provided in a Cas6e-independent manner by transcription termination, the CRISPR-Cas system can function without Cas6e. The results should allow facile interrogation of various targets by type I-E CRISPR-Cas system in E. coli using unit-sized crRNAs generated by transcription.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Bacteriophages / genetics
CRISPR-Associated Proteins / metabolism
CRISPR-Associated Proteins / physiology*
CRISPR-Cas Systems*
Endoribonucleases / metabolism
Endoribonucleases / physiology*
Escherichia coli / enzymology
Escherichia coli / genetics*
Plasmids / genetics
RNA / metabolism
Transcription Termination, Genetic

Substances

CRISPR-Associated Proteins
RNA
Endoribonucleases

The Cas6e ribonuclease is not required for interference and adaptation by the E. coli type I-E CRISPR-Cas system

Authors

Affiliations

Abstract

Publication types

MeSH terms

Substances

Grants and funding