Target sequence requirements of a type III-B CRISPR-Cas immune system

Kaitlin Johnson, Brian A. Learn, Michael A. Estrella, Scott Bailey

Research output: Contribution to journalArticle

Abstract

CRISPR-Cas systems are RNA-based immune systems that protect many prokaryotes from invasion by viruses and plasmids. Type III CRISPR systems are unique, as their targeting mechanism requires target transcription. Upon transcript binding, DNA cleavage by type III effector complexes is activated. Type III systems must differentiate between invader and native transcripts to prevent autoimmunity. Transcript origin is dictated by the sequence that flanks the 3 end of the RNA target site (called the PFS). However, how the PFS is recognized may vary among different type III systems. Here, using purified proteins and in vitro assays, we define how the type III-B effector from the hyperthermophilic bacterium Thermotoga maritima discriminates between native and invader transcripts. We show that native transcripts are recognized by base pairing at positions 2 to 5 of the PFS and by a guanine at position 1, which is not recognized by base pairing. We also show that mismatches with the RNA target are highly tolerated in this system, except for those nucleotides adjacent to the PFS. These findings define the target requirement for the type III-B system from T. maritima and provide a framework for understanding the target requirements of type III systems as a whole.

Original languageEnglish (US)
Pages (from-to)10290-10299
Number of pages10
JournalJournal of Biological Chemistry
Volume294
Issue number26
DOIs
StatePublished - Jan 1 2019

Fingerprint

CRISPR-Cas Systems
Clustered Regularly Interspaced Short Palindromic Repeats
Immune system
Thermotoga maritima
Immune System
RNA
Base Pairing
DNA Cleavage
Guanine
Transcription
Autoimmunity
Viruses
Assays
Bacteria
Plasmids
Nucleotides
DNA
Proteins

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Target sequence requirements of a type III-B CRISPR-Cas immune system. / Johnson, Kaitlin; Learn, Brian A.; Estrella, Michael A.; Bailey, Scott.

In: Journal of Biological Chemistry, Vol. 294, No. 26, 01.01.2019, p. 10290-10299.

Research output: Contribution to journalArticle

Johnson, Kaitlin ; Learn, Brian A. ; Estrella, Michael A. ; Bailey, Scott. / Target sequence requirements of a type III-B CRISPR-Cas immune system. In: Journal of Biological Chemistry. 2019 ; Vol. 294, No. 26. pp. 10290-10299.
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