Selection Signatures in the First Exon of Paralogous Receptor Kinase Genes from the Sym2 Region of the Pisum sativum L. Genome.

Anton S Sulima,Ludmila A Lutova,Vladimir A Zhukov,Igor A Tikhonovich,Alexey A Afonin,Aleksandr I Zhernakov

doi:10.3389/fpls.2017.01957

Abstract

During the initial step of the symbiosis between legumes (Fabaceae) and nitrogen-fixing bacteria (rhizobia), the bacterial signal molecule known as the Nod factor (nodulation factor) is recognized by plant LysM motif-containing receptor-like kinases (LysM-RLKs). The fifth chromosome of barrel medic (Medicago truncatula Gaertn.) contains a cluster of paralogous LysM-RLK genes, one of which is known to participate in symbiosis. In the syntenic region of the pea (Pisum sativum L.) genome, three genes have been identified: PsK1 and PsSym37, two symbiosis-related LysM-RLK genes with known sequences, and the unsequenced PsSym2 gene which presumably encodes a LysM-RLK and is associated with increased selectivity to certain Nod factors. In this work, we identified a new gene encoding a LysM-RLK, designated as PsLykX, within the Sym2 genomic region. We sequenced the first exons (corresponding to the protein receptor domain) of PsSym37, PsK1, and PsLykX from a large set of pea genotypes of diverse origin. The nucleotide diversity of these fragments was estimated and groups of haplotypes for each gene were revealed. Footprints of selection pressure were detected via comparative analyses of SNP distribution across the first exons of these genes and their homologs MtLYK2, MtLYK3, and MtLYK4 from M. truncatula retrieved from the Medicago Hapmap project. Despite the remarkable similarity among all the studied genes, they exhibited contrasting selection signatures, possibly pointing to diversification of their functions. Signatures of balancing selection were found in LysM1-encoding parts of PsSym37 and PsK1, suggesting that the diversity of these parts may be important for pea LysM-RLKs. The first exons of PsSym37 and PsK1 displayed signatures of purifying selection, as well as MtLYK2 of M. truncatula. Evidence of positive selection affecting primarily LysM domains was found in all three investigated M. truncatula genes, as well as in the pea gene PsLykX. The data suggested that PsLykX is a promising candidate for PsSym2, which has remained elusive for more than 30 years.

Highlights

One of the hallmark features of legumes (Fabaceae) is their ability to form beneficial symbioses with nitrogen-fixing soil bacteria collectively known as rhizobia
To detect novel genes encoding lysin motifs (LysMs)-receptor-like kinases (RLKs) in the Sym2 region of pea linkage group I (LG I), the pea Psa-B-Cam BAC library was screened at the INRA-CNRGV Plant Genomic Center using quantitative PCR with primer pairs for PsSym37 and PsK1
The PsLykX complete open reading frame was successfully amplified from cDNA generated from RNA extracted from pea roots inoculated with nodule bacteria, and the sequences of PsLykX cDNA ends were verified using 3′- and 5′-rapid amplification of cDNA ends (RACE)

Summary

Introduction

One of the hallmark features of legumes (Fabaceae) is their ability to form beneficial symbioses with nitrogen-fixing soil bacteria collectively known as rhizobia. The establishment of the legume–rhizobial symbiosis begins with mutual recognition of the partners During this initial step, a bacterial lipo-chitooligosaccharide signaling molecule known as the Nod factor (nodulation factor; Denarie et al, 1996) is recognized by plant receptors in the LysM-RLK protein family (Limpens et al, 2003; Madsen et al, 2003; Broghammer et al, 2012). Various LysM-RLKs are required to recognize molecular signals from rhizobia, arbuscular-mycorrhizal fungi (BuendíaClavería et al, 2003; Oldroyd, 2013; Gobbato, 2015; Kawaharada et al, 2015; Buendia et al, 2016; Rasmussen et al, 2016), and some pathogenic microorganisms (Zhang et al, 2015) They occur in non-legumes such as, Arabidopsis thaliana, which is unable to form nitrogen-fixing nodules or mycorrhizal associations (Veiga et al, 2013). LysM-RLKs contribute to the selection of the most effective combinations of micro- and macrosymbionts (Provorov and Vorobyov, 2013)

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