Specificity of ARGONAUTE7-miR390 interaction and dual functionality in TAS3 trans-acting siRNA formation

Molecular and Cellular Biology Program, Oregon State University, Corvallis, OR 97331, USA.
Cell (Impact Factor: 32.24). 05/2008; 133(1):128-41. DOI: 10.1016/j.cell.2008.02.033
Source: PubMed


Trans-acting siRNA form through a refined RNAi mechanism in plants. miRNA-guided cleavage triggers entry of precursor transcripts into an RNA-DEPENDENT RNA POLYMERASE6 pathway, and sets the register for phased tasiRNA formation by DICER-LIKE4. Here, we show that miR390-ARGONAUTE7 complexes function in distinct cleavage or noncleavage modes at two target sites in TAS3a transcripts. The AGO7 cleavage, but not the noncleavage, function could be provided by AGO1, the dominant miRNA-associated AGO, but only when AGO1 was guided to a modified target site through an alternate miRNA. AGO7 was highly selective for interaction with miR390, and miR390 in turn was excluded from association with AGO1 due entirely to an incompatible 5' adenosine. Analysis of AGO1, AGO2, and AGO7 revealed a potent 5' nucleotide discrimination function for some, although not all, ARGONAUTEs. miR390 and AGO7, therefore, evolved as a highly specific miRNA guide/effector protein pair to function at two distinct tasiRNA biogenesis steps.

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Available from: Miya D Howell, Oct 02, 2015
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    • " - nt phasiRNAs . However , the pro - portion of 3 0 ' U ' ends is significantly higher for 22 - nt than for 21 - nt phasiRNAs . Knowing that data from Arabidopsis show that 5 0 ' U ' small RNAs are enriched in AGO1 ( Mi et al . , 2008 ) , while 5 0 ' A ' 21 - to 22 - nt small RNAs are enriched in AGO2 , AGO4 , AGO6 , or AGO9 ( Mi et al . , 2008 ; Montgomery et al . , 2008 ; Havecker et al . , 2010 ; McCue et al . , 2015 ) , it is likely that the differentially abundant pha - siRNAs may be sorted to AGOs based on their 5 0 sequences . The function of the 3 0 variation in the 22 - nt phasiRNAs remains to be determined , but could result from 3 0 tailing mediated by HESO1 or related nucleotidyl transferases"
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    ABSTRACT: Small RNAs are a class of noncoding RNAs which are of great importance in gene expression regulatory networks. Different families of small RNAs are generated via distinct biogenesis pathways. One such family specific to plants is that of phased, secondary siRNAs (phasiRNAs); these require RDR6, DCL4, and (typically) a microRNA (miRNA) trigger for their biogenesis. Protein-encoding genes are an important source of phasiRNAs. The model legume Medicago truncatula generates phasiRNAs from many PHAS loci, and we aimed to investigate their biogenesis and mechanism by which miRNAs trigger these molecules. We modulated miRNA abundances in transgenic tissues showing that the abundance of phasiRNAs correlates with the levels of both miRNA triggers and the target, precursor transcripts. We identified sets of phasiRNAs or PHAS loci that predominantly and substantially increase in response to miRNA overexpression. In the process of validating targets from miRNA overexpression tissues, we found that in the miRNA-mRNA target pairing, the 3' terminal nucleotide (the 22(nd) position), but not the 10(th) position, is important for phasiRNA production. Mutating the single 3' terminal nucleotide dramatically diminishes phasiRNA production. Ectopic expression of Medicago NB-LRR-targeting miRNAs in Arabidopsis showed that only a few NB-LRRs are capable of phasiRNA production; our data indicate that this might be due to target inaccessibility determined by sequences flanking target sites. Our results suggest that target accessibility is an important component in miRNA-target interactions that could be utilized in target prediction, and the evolution of mRNA sequences flanking miRNA target sites may be impacted. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
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    • "Control construct pH7WG2-GUS was generated by LR recombination between pENTR- GUS (Life technologies) and pH7GW2-OsUbi. pMDC32-GUS construct was used before (Montgomery et al., 2008). The sequence of all ami- RNA precursors used here are in Appendix S3. "
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    • "This evidence has been provided recently (Minoia et al., 2014). In leaves of PSTVd-infected N. benthamiana , the endogenous AGO1 and distinct epitope-tagged AGOs from A. thaliana that were overexpressed —AGO1, AGO2, AGO3, AGO4, AGO5 and AGO9, but not AGO6, AGO7 and AGO10—associate with vd-sRNAs displaying the same properties (5 -terminal nucleotide and size) reported previously for endogenous and viral sRNAs (Mi et al., 2008; Montgomery et al., 2008; Takeda et al., 2008). The AGO-loaded vd-sRNAs adopt specific distributions along the viroid genome. "
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