Cas9 specifies functional viral targets during CRISPR-Cas adaptation

Autor: Catherine L. Weiner, Poulami Samai, Luciano A. Marraffini, Gregory W. Goldberg, Robert Heler, Joshua W. Modell, David Bikard
Přispěvatelé: Rockefeller University [New York], Biologie de Synthèse - Synthetic biology, Institut Pasteur [Paris] (IP), R.H. is the recipient of a Howard Hughes International Student Research Fellowship. P.S. is supported by a Helmsley Postdoctoral Fellowship for Basic and Translational Research on Disorders of the Digestive System at The Rockefeller University. J.W.M. is a Fellow of The Jane Coffin Childs Memorial Fund for Medical Research. D.B. is supported by a Harvey L. Karp Discovery Award and the Bettencourt Schuller Foundation. L.A.M. is supported by the Rita Allen Scholars Program, an Irma T. Hirschl Award, a Sinsheimer Foundation Award and a NIH Director’s New Innovator Award (1DP2AI104556-01)., Institut Pasteur [Paris]
Jazyk: angličtina
Rok vydání: 2015
Předmět:
Small RNA
MESH: CRISPR-Cas Systems
Streptococcus pyogenes
CRISPR-Associated Proteins
Biology
MESH: Base Sequence
Adaptive Immunity
medicine.disease_cause
Genome
Article
MESH: Protein Structure
Tertiary
stomatognathic system
parasitic diseases
medicine
MESH: Staphylococcus aureus
Escherichia coli
CRISPR
MESH: Protein Binding
Clustered Regularly Interspaced Short Palindromic Repeats
Genetics
Nuclease
[SDV.GEN]Life Sciences [q-bio]/Genetics
Multidisciplinary
MESH: Molecular Sequence Data
Integrases
Cas9
MESH: CRISPR-Associated Proteins
Palindrome
DNA
MESH: DNA
Viral
Protospacer adjacent motif
MESH: Clustered Regularly Interspaced Short Palindromic Repeats
MESH: Nucleotide Motifs
[SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology
embryonic structures
DNA
Viral
biology.protein
MESH: Substrate Specificity
CRISPR-Cas Systems
MESH: Streptococcus pyogenes
Zdroj: Nature
Nature, 2015, 519 (7542), pp.199-202. ⟨10.1038/nature14245⟩
Nature, Nature Publishing Group, 2015, 519 (7542), pp.199-202. ⟨10.1038/nature14245⟩
ISSN: 1476-4687
0028-0836
1476-4679
DOI: 10.1038/nature14245⟩
Popis: International audience; Clustered regularly interspaced short palindromic repeat (CRISPR) loci and their associated (Cas) proteins provide adaptive immunity against viral infection in prokaryotes. Upon infection, short phage sequences known as spacers integrate between CRISPR repeats and are transcribed into small RNA molecules that guide the Cas9 nuclease to the viral targets (protospacers). Streptococcus pyogenes Cas9 cleavage of the viral genome requires the presence of a 5'-NGG-3' protospacer adjacent motif (PAM) sequence immediately downstream of the viral target. It is not known whether and how viral sequences flanked by the correct PAM are chosen as new spacers. Here we show that Cas9 selects functional spacers by recognizing their PAM during spacer acquisition. The replacement of cas9 with alleles that lack the PAM recognition motif or recognize an NGGNG PAM eliminated or changed PAM specificity during spacer acquisition, respectively. Cas9 associates with other proteins of the acquisition machinery (Cas1, Cas2 and Csn2), presumably to provide PAM-specificity to this process. These results establish a new function for Cas9 in the genesis of prokaryotic immunological memory.
Databáze: OpenAIRE
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