Identification and Molecular Characterization of RWP-RK Transcription Factors in Soybean

Abstract

The <i>RWP-RK</i> is a small family of plant-specific transcription factors that are mainly involved in nitrate starvation responses, gametogenesis, and root nodulation. To date, the molecular mechanisms underpinning nitrate-regulated gene expression in many plant species have been extensively studied. However, the regulation of nodulation-specific <i>NIN</i> proteins during nodulation and rhizobial infection under nitrogen starvation in soybean still remain unclear. Here, we investigated the genome-wide identification of <i>RWP-RK</i> transcription factors and their essential role in nitrate-inducible and stress-responsive gene expression in soybean. In total, 28 <i>RWP-RK</i> genes were identified from the soybean genome, which were unevenly distributed on 20 chromosomes from 5 distinct groups during phylogeny classification. The conserved topology of RWP-RK protein motifs, cis-acting elements, and functional annotation has led to their potential as key regulators during plant growth, development, and diverse stress responses. The RNA-seq data revealed that the up-regulation of <i>GmRWP-RK</i> genes in the nodules indicated that these genes might play crucial roles during root nodulation in soybean. Furthermore, qRT-PCR analysis revealed that most GmRWP-RK genes under <i>Phytophthora sojae</i> infection and diverse environmental conditions (such as heat, nitrogen, and salt) were significantly induced, thus opening a new window of possibilities into their regulatory roles in adaptation mechanisms that allow soybean to tolerate biotic and abiotic stress. In addition, the dual luciferase assay indicated that <i>GmRWP-RK1</i> and <i>GmRWP-RK2</i> efficiently bind to the promoters of <i>GmYUC2</i>, <i>GmSPL9</i>, and <i>GmNIN</i>, highlighting their possible involvement in nodule formation. Together, our findings provide novel insights into the functional role of the <i>RWP-RK</i> family during defense responses and root nodulation in soybean.