What question did this study set out to answer?

To establish and validate a cell-free plant system capable of recapitulating Argonaute- and RISC-mediated antiviral RNA silencing during active virus replication.

March 27, 2013Nucleic Acids ResearchOpen Access

AGO/RISC-mediated antiviral RNA silencing in a plant in vitro system

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Authors

JSJana SchuckMartin Luther University Halle-Wittenberg TGTorsten GursinskyLuther University VPVitantonio PantaleoInstitute for Sustainable Plant Protection

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Implication

In vitro study demonstrates reconstitution of Argonaute-mediated antiviral RNA silencing in plant extracts, indicating structural vulnerabilities in replicating viral RNA genomes.

Key Points

To establish and validate a cell-free plant system capable of recapitulating Argonaute- and RISC-mediated antiviral RNA silencing during active virus replication.
Used cytoplasmic extracts derived from tobacco protoplasts that support Tombusvirus RNA replication.
Reconstituted RNA-induced silencing complexes (RISC) using recombinant plant ARGONAUTE proteins (AGO1, AGO2, AGO3, and AGO5) programmed with exogenous synthetic siRNAs or viral siRNAs.
Assessed endonucleolytic cleavage susceptibility of replicating (+)RNA versus (-)RNA viral strands, target RNA secondary structure constraints, and the inhibitory effect of viral silencing suppressor p19.
RISC assembled with AGO1, AGO2, AGO3, or AGO5 successfully catalyzed target-specific endonucleolytic cleavage of viral RNA and suppressed viral replication in vitro.
Replicating viral (+)RNA strands were directly accessible to RISC cleavage, whereas (-)RNA replication intermediates remained completely protected.
Viral silencing suppressor p19 effectively blocked antiviral silencing when present early during RISC assembly, and effective viral siRNAs for AGO1 and AGO2 targeted shared accessible secondary structure sites.

Cite This Study

Schuck et al. (2013) studied this question.

synapsesocial.com/papers/6a95ab80eae36b2f6ea8ea13 https://doi.org/10.1093/nar/gkt193

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Implication

In vitro study demonstrates reconstitution of Argonaute-mediated antiviral RNA silencing in plant extracts, indicating structural vulnerabilities in replicating viral RNA genomes.

AGO/RISC-mediated antiviral RNA silencing in a plant in vitro system

Discussion

Member takes

Implication

Key Points

Cite This Study

AGO/RISC-mediated antiviral RNA silencing in a plant in vitro system

Discussion

Member takes

Implication

Key Points

Cite This Study

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