Overexpression of X Intrinsic Protein 1;1 in Nicotiana tabacum and Arabidopsis reduces boron allocation to shoot sink tissues.

Manuela Desiree Bienert,Beatriz Muries,François Chaumont,Delphine Crappe,Gerd Patrick Bienert

doi:10.1002/pld3.143

Abstract

Major Intrinsic Proteins (MIP) are a family of channels facilitating the diffusion of water and/or small solutes across cellular membranes. X Intrinsic Proteins (XIP) form the least characterized MIP subfamily in vascular plants. XIPs are mostly impermeable to water but facilitate the diffusion of hydrogen peroxide, urea and boric acid when expressed in heterologous expression systems. However, their transport capabilities in planta and their impact on plant physiology are still unknown. Here, we demonstrated that overexpression of NtXIP1;1 in Nicotiana tabacum by the En2pPMA4 or the 35S CaMV promoter and in Arabidopsis, which does not contain any XIP gene, by the 35S CaMV promoter, resulted in boron (B)‐deficiency symptoms such as death of the shoot apical meristem, infertile flowers, and puckered leaves. Leaf B concentrations in symptomatic tissues and B xylem sap concentrations were lower in the overexpressors than in control plants. Importantly, expression of NtXIP1;1 under the control of the AtNIP5;1 promoter complemented the B deficiency phenotype of the Atnip5;1 knockout mutant, defining its ability to act as a boric acid channel in planta. Protein quantification analysis revealed that NtXIP1;1 was predominantly expressed in young B‐demanding tissues and induced under B‐deficient conditions. Our results strongly suggest that NtXIP1;1 plays a role in B homeostasis and its tissue‐specific expression critically contributes to the distribution of B within tobacco.

Highlights

Major Intrinsic Proteins (MIPs) are membrane channels found in all kingdoms of life
Compared to 13 isoforms in humans (Agre & Kozono, 2003), a relatively large number of isoforms are expressed in plants e.g. 35 MIPs in Arabidopsis thaliana (Johanson et al, 2001), 71 in cotton (Park, Scheffler, Bauer, & Campbell, 2010) and even 121 in Brassica napus (Yuan et al, 2017)
While Plasma membrane Intrinsic Proteins (PIPs), Tonoplast Intrinsic Proteins (TIPs), Small basic Intrinsic Proteins (SIPs) and Nodulin26-like Intrinsic Proteins (NIPs) subfamily members occur in all so far published genomes of land plant taxa, X Intrinsic Proteins (XIP) have most likely been evolutionarily lost in monocots and some eudicots such as Brassicacea species (Danielson & Johanson, 2008)

Summary

| INTRODUCTION

Major Intrinsic Proteins (MIPs) are membrane channels found in all kingdoms of life. They assemble as tetramers in various cellular membranes whereas each monomer represents an individual functional unit. Phylogenetic comparisons of XIP isoforms of different land plant species identified two major clusters composed of four (Lopez et al, 2012) or five (Venkatesh, Yu, Gaston, & Park, 2015) distinct XIP groups. This diversity in XIP sequences may hint to heterogeneous functions of various isoforms. Summarizing the current knowledge on plant XIPs, it becomes obvious that molecular characterization efforts for various plant XIPs are confined to developmental- and organ-specific expression analyses and transport selectivity assays in heterologous expression systems Both approaches resulted in a diffuse diversity of identified substrates and expression patterns which pointed neither to a clear functional nor physiological understanding of this uncharacterized protein channel family. Using the X. laevis oocyte system, we quantitatively provided evidence that XIPs do transport boric acid in a direct B uptake assay

| MATERIALS AND METHODS

Findings

| DISCUSSION