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Guide-independent DNA cleavage by archaeal Argonaute from Methanocaldococcus jannaschii


Prokaryotic Argonaute proteins acquire guide strands derived from invading or mobile genetic elements, via an unknown pathway, to direct guide-dependent cleavage of foreign DNA. Here, we report that Argonaute from the archaeal organism Methanocaldococcus jannaschii (MjAgo) possesses two modes of action: the canonical guide-dependent endonuclease activity and a non-guided DNA endonuclease activity. The latter allows MjAgo to process long double-stranded DNAs, including circular plasmid DNAs and genomic DNAs. Degradation of substrates in a guide-independent fashion primes MjAgo for subsequent rounds of DNA cleavage. Chromatinized genomic DNA is resistant to MjAgo degradation, and recombinant histones protect DNA from cleavage in vitro. Mutational analysis shows that key residues important for guide-dependent target processing are also involved in guide-independent MjAgo function. This is the first characterization of guide-independent cleavage activity for an Argonaute protein potentially serving as a guide biogenesis pathway in a prokaryotic system.

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Figure 1: Guide-directed target cleavage activity of MjAgo using canonical and non-canonical substrates.
Figure 2: MjAgo processes long linear and circular dsDNAs and genomic DNA in the absence of a guide DNA.
Figure 3: Characterization of DNA degradation products and influence on MjAgo-mediated plasmid degradation.
Figure 4: Mutational analysis of MjAgo guide-independent plasmid cleavage activity.
Figure 5: Putative model of guide-dependent and guide-independent DNA silencing by MjAgo.


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The authors thank all members of the Grohmann laboratory and in particular K. Kramm for cloning of the MjAgo catalytic mutant. The authors thank G. Meister for discussions. Work in the Grohmann RNAP laboratory was funded by the Deutsche Forschungsgemeinschaft (GR 3840/2-1). S.V.A. and M.v.W. acknowledge funding by the European Research Council (starting grant ARCHAELLUM 311523). S.Sc. acknowledges funding by the Deutsche Forschungsgemeinschaft, the excellence cluster CIPSM and Fonds der Chemischen Industrie.

Author information




D.G. conceived the study. A.Z., S.W., M.v.W., L.E., S.St., S.O., A.K., S.B. and D.G. carried out experimental work. A.Z., S.W., M.v.W., S.-V.A., S.Sc., P.T., D.G., A.K. and F.W. performed data analysis. D.G. wrote the manuscript. All authors edited the manuscript.

Corresponding author

Correspondence to Dina Grohmann.

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The authors declare no competing financial interests.

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Supplementary Methods and Supplementary Figures 1–13. (PDF 1676 kb)

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Zander, A., Willkomm, S., Ofer, S. et al. Guide-independent DNA cleavage by archaeal Argonaute from Methanocaldococcus jannaschii. Nat Microbiol 2, 17034 (2017).

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