Molecular identification and physiological functional analysis of NtNRT1.1B that mediated nitrate long-distance transport and improved plant growth when overexpressed in tobacco

Frontiers in Plant Science

Plant Science

Fengchun Cao¹  Yucheng Xiang¹  Changzheng Wu¹  Wenrui Li¹  Weiqin Yang¹  Lai-Hua Liu¹  Shuhui Duan²  Mingfa Zhang²  Ming Fang²  Wenxuan Pu³  Pingjun Huang³ 

[1] College of Resources and Environmental Sciences, Department of Plant Nutrition, Key Lab of Plant-Soil Interaction of Ministry of Education, China Agricultural University, Beijing, China;Hunan Tobacco Research Institute (Changsha, Chenzhou, Xiangxi), China National Tobacco Corporation Hunan Company, Changsha, China;Tobacco Research Institute of Technology Centre, China Tobacco Hunan Industrial Corporation, Changsha, China;

关键词: nitrate transporter NtNRT1.1B;    yeast complementation;    long-distance transport;    promoter activity;    overexpression in tobacco;    nitrogen use efficiency;   

DOI  :  10.3389/fpls.2023.1078978

 received in 2022-10-24, accepted in 2023-01-20,  发布年份 2023

来源: Frontiers

【 摘 要 】

Although recent physiological studies demonstrate that flue-cured tobacco preferentially utilizes nitrate (NO3−) or ammonium nitrate (NH4NO3), and possesses both high- and low-affinity uptake systems for NO3−, little is known about the molecular component(s) responsible for acquisition and translocation in this crop. Here we provide experimental data showing that NtNRT1.1B with a 1,785-bp coding sequence exhibited a function in mediating NO3− transport associated with tobacco growth on NO3− nutrition. Heterologous expression of NtNRT1.1B in the NO3− uptake-defective yeast Hp△ynt1 enabled a growth recovery of the mutant on 0.5 mM NO3−, suggesting a possible molecular function of NtNRT1.1B in the import of NO3− into cells. Transient expression of NtNRT1.1B::green fluorescent protein (GFP) in tobacco leaf cells revealed that NtNRT1.1B targeted mainly the plasma membrane, indicating the possibility of NO3− permeation across cell membranes via NtNRT1.1B. Furthermore, promoter activity assays using a GFP marker clearly indicated that NtNRT1.1B transcription in roots may be down-regulated by N starvation and induced by N resupply, including NO3−, after 3 days’ N depletion. Significantly, constitutive overexpression of NtNRT1.1B could remarkably enhance tobacco growth by showing a higher accumulation of biomass and total N, NO3−, and even NH4+ in plants supplied with NO3−; this NtNRT1.1B-facilitated N acquisition/accumulation could be strengthened by short-term 15N-NO3− root influx assays, which showed 15%–20% higher NO3− deposition in NtNRT1.1B-overexpressors as well as a high affinity of NtNRT1.1B for NO3− at a Km of around 30–45 µM. Together with the detection of NtNRT1.1B promoter activity in the root stele and shoot–stem vascular tissues, and higher NO3− in both xylem exudate and the apoplastic washing fluid of NtNRT1.1B-transgenic lines, NtNRT1.1B could be considered as a valuable molecular breeding target aiming at improving crop N-use efficiency by manipulating the absorption and long-distance distribution/transport of nitrate, thus adding a new functional homolog as a nitrate permease to the plant NRT1 family.

【 授权许可】

Unknown   
Copyright © 2023 Wu, Xiang, Huang, Zhang, Fang, Yang, Li, Cao, Liu, Pu and Duan