Disruption of the Lotus japonicus transporter LjNPF2.9 increases shoot biomass and nitrate content without affecting symbiotic performances

Abstract

Background After uptake from soil into the root tissue, distribution and allocation of nitrate throughout the whole plant body, is a critical step of nitrogen use efficiency (NUE) and for modulation of plant growth in response to various environmental conditions. In legume plants nitrate distribution is also important for the regulation of the nodulation process that allows to fix atmospheric N (N 2 ) through the symbiotic interaction with rhizobia (symbiotic nitrogen fixation, SNF). Results Here we report the functional characterization of the Lotus japonicus gene LjNPF2.9, which is expressed mainly in the root vascular structures, a key localization for the control of nitrate allocation throughout the plant body. LjNPF2.9 expression in Xenopus laevis oocytes induces 15 NO 3 accumulation indicating that it functions as a nitrate importer. The phenotypic characterization of three independent knock out mutants indicates an increased shoot biomass in the mutant backgrounds. This phenotype is associated to an increased/decreased nitrate content detected in the shoots/roots. Furthermore, our analysis indicates that the accumulation of nitrate in the shoot does not affect the nodulation and N-Fixation capacities of the knock out mutants. Conclusions This study shows that LjNPF2.9 plays a crucial role in the downward transport of nitrate to roots, occurring likely through a xylem-to-phloem loading-mediated activity. The increase of the shoot biomass and nitrate accumulation might represent a relevant phenotype in the perspective of an improved NUE and this is further reinforced in legume plants by the reported lack of effects on the SNF efficiency. Electronic supplementary material The online version of this article (10.1186/s12870-019-1978-5) contains supplementary material, which is available to authorized users.

Molecular Biology in the Improvement of Biological Nitrogen Fixation by Rhizobia and Extending the Scope to Cereals

“…In plants, the nitrogen flux across different organs is controlled by transporters that belong to mainly four protein families, namely chloride channels, slowly activating anion channels, nitrate/peptide transporters (NPF) and nitrate transporters [ 92 ]. By disrupting L. japonicus transporter LjNPF2.9, Sol et al [ 93 ] demonstrated an increase in shoot biomass without affecting symbiotic nitrogen fixation. In summary, the strains were genetically modified targeting various genes and different processes of host–rhizobial symbiosis.…”

The functional characterization of LjNRT2.4 indicates a novel, positive role of nitrate for an efficient nodule N2‐fixation activity

“…In the case of nitrate, a primary role in the networks governing nitrate assimilation, storage and distribution among different plant tissues and organs, is played by two protein families – the low‐affinity Nitrate Transporter Peptide (NPF) and the high‐affinity Nitrate Transporter (NRT2; Wang et al ., 2018). NPF is a large family of 53, 80 and 86 members in Arabidopsis thaliana , Oryza sativa and Lotus japonicus , respectively (Tsay et al ., 2007; Léran et al ., 2014; Sol et al ., 2019). The NPF members are divided into eight subfamilies and able to transport different substrates (Léran et al ., 2014).…”

The Lotus japonicus NPF3.1 Is a Nodule-Induced Gene That Plays a Positive Role in Nodule Functioning

“…We have recently reported the identification of the 86 members of the L. japonicus NPF family (accession MG20; Criscuolo et al, 2012;Sol et al, 2019)…”

“…Frontiers in Plant Science | www.frontiersin.org was assigned based on the two-letter code established in Léran et al (2014). The name LjNPF3.1 is assigned to the MG20 gene Lj2g3v1155500.1 (Sol et al, 2019) and the identical copy LotjaGi2g1v0278100.1, identified in the L. japonicus accession Gifu (Supplementary Table S2; Kamal et al, 2020). 4 LjNPF3.1 is one of the three members of the clade 3 (Supplementary Table S2) coding for a 580-amino acid protein with a molecular mass of 64.4 kDa.…”

“…A primary role in the network governing nitrate uptake, assimilation, storage, and distribution among different plant tissues and organs is played by two protein families, the low-affinity nitrate transporter peptide family (NPF; LATS > 0.5 mM) and the high-affinity nitrate transporter system (NRT2; HATS < 0.5 mM). NPF is a large plant family consisting of 53, 93, and 86 members in Arabidopsis thaliana, Oryza sativa, and Lotus japonicus, respectively (Tsay et al, 2007;Léran et al, 2014;Sol et al, 2019). To date, nitrate transport activity has been reported for 17 A. thaliana NPF proteins (Corratgé-Faillie and Lacombe, 2017), with AtNPF6.3 being the only exception as this displays a switching dual nitrate affinity, controlled via the phosphorylation of the threonine residue at position 101, in response to fluctuating external concentrations (Liu et al, 1999;Liu and Tsay, 2003;Parker and Newstead, 2014;Sun et al, 2014).…”