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Binding of tobamovirus replication protein with small RNA duplexes
Yukio Kurihara1, Naoko Inaba1, Natsumaro Kutsuna2
1Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, Komaba 3-8-1, Meguro-ku, Tokyo 153-8902, Japan.
The Journal of General Virology
|July 12, 2007
Summary
Tobacco mosaic virus (TMV) infection alters small RNA profiles in Arabidopsis. The TMV replication protein suppresses RNA silencing by binding small RNA duplexes, impacting plant defense.
Area of Science:
- Plant Virology
- Molecular Biology
- RNA Silencing
Background:
- RNA silencing is a crucial plant defense mechanism against viruses.
- Tobamoviruses, like tobacco mosaic virus (TMV), encode proteins that suppress RNA silencing.
- The specific mechanisms by which viral proteins interfere with small RNA pathways are not fully understood.
Purpose of the Study:
- To investigate the small interfering RNA (siRNA) profiles in Arabidopsis infected with TMV-Cg.
- To determine if the TMV-Cg 126K replication protein, an RNA-silencing suppressor, affects small RNA abundance and profiles.
- To elucidate the mechanism by which the TMV replication protein suppresses RNA silencing.
Main Methods:
- Small RNA cloning and sequencing to analyze siRNA populations.
- Northern blot analysis to detect specific small RNAs, including microRNAs (miRNAs).
- Transient expression of the TMV replication protein.
- Co-immunoprecipitation assays to identify protein-RNA interactions.
- Gel mobility-shift assays to confirm binding of the replication protein to small RNA duplexes.
Main Results:
- TMV infection altered the size distribution of endogenous small RNAs in Arabidopsis, with a shift towards 21 nt siRNAs.
- The TMV 126K replication protein co-immunoprecipitated with miRNAs, miRNA* duplexes, and hairpin-derived siRNAs.
- Gel mobility-shift assays confirmed that the TMV replication protein directly binds to small RNA duplexes.
Conclusions:
- The TMV-Cg replication protein acts as an RNA-silencing suppressor by binding to small RNA duplexes.
- This binding interferes with the small RNA profile in infected plants, potentially hindering antiviral defense.
- The findings provide insights into the molecular mechanisms of tobamovirus RNA silencing suppression.
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