Specificity of ARGONAUTE7-miR390 interaction and dual functionality in

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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    • " - 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.
    The Plant Journal 06/2015; 83(3). DOI:10.1111/tpj.12900 · 5.97 Impact Factor
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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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    ABSTRACT: Artificial microRNAs (amiRNAs) are used for selective gene silencing in plants. However, current methods to produce amiRNA constructs for silencing transcripts in monocot species are not suitable for simple, cost-effective and large-scale synthesis. Here, a series of expression vectors based on Oryza sativa MIR390 (OsMIR390) precursor was developed for high-throughput cloning and high expression of amiRNAs in monocots. Four different amiRNA sequences designed to target specifically endogenous genes and expressed from OsMIR390-based vectors were validated in transgenic Brachypodium distachyon plants. Surprisingly, amiRNAs accumulated to higher levels and were processed more accurately when expressed from chimeric OsMIR390-based precursors that include distal stem-loop sequences from Arabidopsis thaliana MIR390a (AtMIR390a). In all cases, transgenic plants displayed the predicted phenotypes induced by target gene repression, and accumulated high levels of amiRNAs and low levels of the corresponding target transcripts. Genome-wide transcriptome profiling combined with 5'-RLM-RACE analysis in transgenic plants confirmed that amiRNAs were highly specific. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    The Plant Journal 03/2015; DOI:10.1111/tpj.12835 · 5.97 Impact Factor
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    • "Its effects on flowering time are because it prolongs the juvenile phase and thus delays the acquisition of the competence to flower (Fahlgren et al., 2006; Rubio-Somoza and Weigel, 2011). MiR390 mediates its effects on flowering time not by directly targeting protein-coding mRNAs but by triggering the production of trans-acting siRNAs from the TAS3 locus, which in turn target and repress mRNA levels of the transcription factors AUXIN RESPONSE FACTORS 3 (ARF3) and ARF4 (Endo et al., 2013; Garcia, 2008; Montgomery et al., 2008; Rubio-Somoza and Weigel, 2011). ARF3 and ARF4 activity promotes the juvenileto-adult vegetative phase transition and mutants defective in tasiRNA biogenesis have a shorter juvenile phase as a result of elevated levels of ARF3 and ARF4. "
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    ABSTRACT: The multiple regulatory pathways controlling flowering and flower development are varied and complex, and they require tight control of gene expression and protein levels. MicroRNAs (miRNAs) act at both the transcriptional and post-transcriptional level to regulate key genes involved in flowering-related processes such as the juvenile–adult transition, the induction of floral competence and flower development. Many different miRNA families are involved in these processes and their roles are summarized in this review, along with potential biotechnological applications for miRNAs in controlling processes related to flowering and flower development.
    Plant Biotechnology Journal 01/2015; DOI:10.1111/pbi.12340 · 5.75 Impact Factor
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