TY - JOUR
T1 - Altered stoichiometry Escherichia coli Cascade complexes with shortened CRISPR RNA spacers are capable of interference and primed adaptation
AU - Kuznedelov, Konstantin
AU - Mekler, Vladimir
AU - Lemak, Sofia
AU - Tokmina-Lukaszewska, Monika
AU - Datsenko, Kirill A.
AU - Jain, Ishita
AU - Savitskaya, Ekaterina
AU - Mallon, John
AU - Shmakov, Sergey
AU - Bothner, Brian
AU - Bailey, Scott
AU - Yakunin, Alexander F.
AU - Severinov, Konstantin
AU - Semenova, Ekaterina
N1 - Publisher Copyright:
© The Author(s) 2016.
PY - 2016/12
Y1 - 2016/12
N2 - The Escherichia coli type I-E CRISPR-Cas system Cascade effector is a multisubunit complex that binds CRISPR RNA (crRNA). Through its 32-nucleotide spacer sequence, Cascade-bound crRNA recognizes protospacers in foreign DNA, causing its destruction during CRISPR interference or acquisition of additional spacers in CRISPR array during primed CRISPR adaptation. Within Cascade, the cr-RNA spacer interacts with a hexamer of Cas7 subunits. We show that crRNAs with a spacer length reduced to 14 nucleotides cause primed adaptation, while crRNAs with spacer lengths of more than 20 nucleotides cause both primed adaptation and target interference in vivo. Shortened crRNAs assemble into altered-stoichiometry Cascade effector complexes containing less than the normal amount of Cas7 subunits. The results show that Cascade assembly is driven by crRNA and suggest that multisubunit type I CRISPR effectors may have evolved from much simpler ancestral complexes.
AB - The Escherichia coli type I-E CRISPR-Cas system Cascade effector is a multisubunit complex that binds CRISPR RNA (crRNA). Through its 32-nucleotide spacer sequence, Cascade-bound crRNA recognizes protospacers in foreign DNA, causing its destruction during CRISPR interference or acquisition of additional spacers in CRISPR array during primed CRISPR adaptation. Within Cascade, the cr-RNA spacer interacts with a hexamer of Cas7 subunits. We show that crRNAs with a spacer length reduced to 14 nucleotides cause primed adaptation, while crRNAs with spacer lengths of more than 20 nucleotides cause both primed adaptation and target interference in vivo. Shortened crRNAs assemble into altered-stoichiometry Cascade effector complexes containing less than the normal amount of Cas7 subunits. The results show that Cascade assembly is driven by crRNA and suggest that multisubunit type I CRISPR effectors may have evolved from much simpler ancestral complexes.
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U2 - 10.1093/nar/gkw914
DO - 10.1093/nar/gkw914
M3 - Article
C2 - 27738137
AN - SCOPUS:85010412028
SN - 0305-1048
VL - 44
SP - 10849
EP - 10861
JO - Nucleic acids research
JF - Nucleic acids research
IS - 22
ER -