Genome-Wide Identification of Key Components of RNA Silencing in Two Phaseolus vulgaris Genotypes of Contrasting Origin and Their Expression Analyses in Response to Fungal Infection.

Juan C Alvarez-Diaz,Guillem Rigaill,Vincent Thareau,Ariane Gratias,Stéphanie Pflieger,Valérie Geffroy,Manon M S Richard,Gianluca Teano,Christine Paysant-Le-Roux

doi:10.3390/genes13010064

Abstract

RNA silencing serves key roles in a multitude of cellular processes, including development, stress responses, metabolism, and maintenance of genome integrity. Dicer, Argonaute (AGO), double-stranded RNA binding (DRB) proteins, RNA-dependent RNA polymerase (RDR), and DNA-dependent RNA polymerases known as Pol IV and Pol V form core components to trigger RNA silencing. Common bean (Phaseolus vulgaris) is an important staple crop worldwide. In this study, we aimed to unravel the components of the RNA-guided silencing pathway in this non-model plant, taking advantage of the availability of two genome assemblies of Andean and Meso-American origin. We identified six PvDCLs, thirteen PvAGOs, 10 PvDRBs, 5 PvRDRs, in both genotypes, suggesting no recent gene amplification or deletion after the gene pool separation. In addition, we identified one PvNRPD1 and one PvNRPE1 encoding the largest subunits of Pol IV and Pol V, respectively. These genes were categorized into subgroups based on phylogenetic analyses. Comprehensive analyses of gene structure, genomic localization, and similarity among these genes were performed. Their expression patterns were investigated by means of expression models in different organs using online data and quantitative RT-PCR after pathogen infection. Several of the candidate genes were up-regulated after infection with the fungus Colletotrichum lindemuthianum.

Highlights

Small RNAs have regulatory roles in a multitude of biological processes, including stress responses, development, metabolism, and maintenance of genome integrity, in a sequence-specific manner [1]
RNA silencing is triggered by double-stranded RNA, and the generation and function of the small RNAs depend on key protein families such as Dicer-like (DCLs), Argonautes (AGOs), and RNA-dependent RNA polymerases (RDRs) [4]
In Arabidopsis thaliana, double-stranded RNA (dsRNA) are processed into -sized sRNA duplexes by one of the four DCL (AtDCL1–4) proteins. dsRNA processing, called dicing, is facilitated by one of the six dsRNA-binding proteins (HYPONASTIC1 or AtHYL1, AtDRB2– 5, and AtDRB7) that interact with specific DCLs [7,8]. dsRNA might derive directly from virus replication, inverted repeats, or convergent transcription. dsRNA formation may be genetically programmed at endogenous loci that produce transcripts with internal stemloop structures

Summary

Introduction

Small RNAs have regulatory roles in a multitude of biological processes, including stress responses, development, metabolism, and maintenance of genome integrity, in a sequence-specific manner [1]. Sequence, genomic distribution, biogenesis, and action, most of these small RNA molecules mediate repressive gene regulation through RNA silencing [2]. In A. thaliana, dsRNA may be synthesized by one of the six RDRs (AtRDR1–6) that copy single-stranded RNA (ssRNA) to initiate a new round of RNA silencing. These small RNAs are incorporated into AGO-containing RNA-induced silencing complexes (RISCs) that guide small RNAs to their targets by sequence complementarity resulting in target RNA degradation, translational inhibition, or heterochromatin formation [6]. The A. thaliana genome encodes 10 AGO proteins (AGO1-10), with various functions such as implication in the RdDM pathway (AGO4) or viral defense (AGO2)

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