Understanding Nitrate Transporter two NRT two and Nitrate Assimilation Related two NAR two protein interaction for high affinity nitrate uptake in wheat

Abstract

newline Nitrogen is a primary nutrient for plant growth and development and NO3¯ is a major newlinenitrogen source for many cultivated crops. Nitrate acquisition and transport, newlineare facilitated by transmembrane proteins known as nitrate transporters. Plants utilize newlinetwo main nitrate transport systems i.e., Low-Affinity and High-Affinity Nitrate newlinetransporters, encoded by the NRT1 (NPF) and NRT2 protein families respectively. newlineThese transporter systems enable nitrate uptake from the soil into the roots, and from newlinethe roots to other plant parts. Low-affinity nitrate transporters (NRT1) are active under newlineoptimal nitrate availability, while high-affinity nitrate transporters (NRT2) function newlinewhen nitrate is limited availability. The function of NRT2, requires other proteins i.e., newlineNAR2, to form HATS. In the present study, genome-wide identification of these two newlinefamilies in the bread wheat and investigate their phylogenetic and evolutionary newlinerelationships, to major cereal crops, and their expression analysis. Also, the possible newlineinteractions between selected TaNRT2 and TaNAR2 proteins have been identified by newlineemploying a split-ubiquitin assay and a bimolecular fluorescent complementation newlineassay. 46 NRT2 and 8 NAR2 family genes were identified in the wheat genome. newlineThese genes exhibited tissue- and growth-stage-specific responses to nitrate-limited newlineconditions across major tissues. Additionally, 15N influx studies at three key newlinephysiological stages of four wheat genotypes demonstrated the significant role of newlineHATS in nitrate transport at both tissue and growth stage levels under NO3¯ limited newlineconditions. Our results showed the root-specific and nitrate-limiting expression of the newlineTaNRT2.1-6 gene. All three TaNAR2 proteins interact with TaNRT2.1-6B. The newlinefunctional validation of this root-specific and nitrate-limiting specific TaNRT2.1-6 of newlineB sub-genome was studied in Arabidopsis mutant (atnrt2.1). The transgenic of newlineTaNRT2.1- B6 and TaNAR2 in atnrt2.1 mutant, among three transformed lines newlineTaNRT2.1-B6+TaNAR2.3-D1 combination showed higher recovery of 15N influx, i.e., newline47.28% and 81.20%, while TaNRT2.1-B6+TaNAR2.2-B1 showed lowest recovery, newlinei.e., 13.76% and 48.01%, than that of atnrt2.1 mutant in N-optimum and N-limited newlineconditions, respectively. This study provides a platform for further exploration of newlinewheat nitrate uptake and translocation mechanisms by providing a genomic and newlinefunctional understanding of NRT2 and NAR2. newline