Proceedings of the National Academy of Sciences - PNAS, 2021-08, Vol.118 (34), p.1
2021
Details
- Autor(en) / Beteiligte
- Titel
- Cooperative recruitment of RDR6 by SGS3 and SDE5 during small interfering RNA amplification in Arabidopsis
- Ist Teil von
- Proceedings of the National Academy of Sciences - PNAS, 2021-08, Vol.118 (34), p.1
- Ort / Verlag
- United States: National Academy of Sciences
- Erscheinungsjahr
- 2021
- Link zum Volltext
- Quelle
- EZB Free E-Journals
- Beschreibungen/Notizen
- Small interfering RNAs (siRNAs) are often amplified from transcripts cleaved by RNA-induced silencing complexes (RISCs) containing a small RNA (sRNA) and an Argonaute protein. Amplified siRNAs, termed secondary siRNAs, are important for reinforcement of target repression. In plants, target cleavage by RISCs containing 22-nucleotide (nt) sRNA and Argonaute 1 (AGO1) triggers siRNA amplification. In this pathway, the cleavage fragment is converted into double-stranded RNA (dsRNA) by RNA-dependent RNA polymerase 6 (RDR6), and the dsRNA is processed into siRNAs by Dicer-like proteins. Because nonspecific RDR6 recruitment causes nontarget siRNA production, it is critical that RDR6 is specifically recruited to the target RNA that serves as a template for dsRNA formation. Previous studies showed that Suppressor of Gene Silencing 3 (SGS3) binds and stabilizes 22-nt sRNA-containing AGO1 RISCs associated with cleaved target, but how RDR6 is recruited to targets cleaved by 22-nt sRNA-containing AGO1 RISCs remains unknown. Here, using cell-free extracts prepared from suspension-cultured cells, we established an in vitro system for secondary siRNA production in which 22-nt siRNA-containing AGO1-RISCs but not 21-nt siRNA-containing AGO1-RISCs induce secondary siRNA production. In this system, addition of recombinant Silencing Defective 5 (SDE5) protein remarkably enhances secondary siRNA production. We show that RDR6 is recruited to a cleavage fragment by 22-nt siRNA-containing AGO1-RISCs in coordination with SGS3 and SDE5. The SGS3-SDE5-RDR6 multicomponent recognition system and the poly(A) tail inhibition may contribute to securing specificity of siRNA amplification.
- Sprache
- Englisch
- Identifikatoren
- ISSN: 0027-8424eISSN: 1091-6490DOI: 10.1073/pnas.2102885118Titel-ID: cdi_pubmedcentral_primary_oai_pubmedcentral_nih_gov_8403909
- Format
- –
- Schlagworte
- Amplification, Arabidopsis - genetics, Arabidopsis - metabolism, Arabidopsis Proteins - genetics, Arabidopsis Proteins - metabolism, Biological Sciences, Cleavage, DNA-directed RNA polymerase, Double-stranded RNA, Gene Expression Regulation, Plant, Gene silencing, Nucleotides, Poly(A), Polyadenylation, Proteins, Recruitment, Ribonucleic acid, RNA, RNA polymerase, RNA, Double-Stranded - genetics, RNA, Double-Stranded - metabolism, RNA, Small Interfering - genetics, RNA, Small Interfering - metabolism, RNA-Dependent RNA Polymerase - genetics, RNA-Dependent RNA Polymerase - metabolism, RNA-directed RNA polymerase, RNA-Induced Silencing Complex - genetics, RNA-Induced Silencing Complex - metabolism, RNA-mediated interference, siRNA