H+-pyrophosphatases enhance low nitrogen stress tolerance in transgenic Arabidopsis and wheat by interacting with a receptor-like protein kinase.

Introduction: Nitrogen is a major abiotic stress that affects plant productivity. Previous studies have shown that plant H+-pyrophosphatases (H+-PPases) enhance plant resistance to low nitrogen stress. However, the molecular mechanism underlying H+-PPase-mediated regulation of plant responses to low nitrogen stress is still unknown. In this study, we aimed to investigate the regulatory mechanism of AtAVP1 in response to low nitrogen stress. Methods and Results: AtAVP1 in Arabidopsis thaliana and EdVP1 in Elymus dahuricus belong to the H+-PPase gene family. In this study, we found that AtAVP1 overexpression was more tolerant to low nitrogen stress than was wild type (WT), whereas the avp1-1 mutant was less tolerant to low nitrogen stress than WT. Plant height, root length, aboveground fresh and dry weights, and underground fresh and dry weights of EdVP1 overexpression wheat were considerably higher than those of SHI366 under low nitrogen treatment during the seedling stage. Two consecutive years of low nitrogen tolerance experiments in the field showed that grain yield and number of grains per spike of EdVP1 overexpression wheat were increased compared to those in SHI366, which indicated that EdVP1 conferred low nitrogen stress tolerance in the field. Furthermore, we screened interaction proteins in Arabidopsis; subcellular localization analysis demonstrated that AtAVP1 and Arabidopsis thaliana receptor-like protein kinase (AtRLK) were located on the plasma membrane. Yeast two-hybrid and luciferase complementary imaging assays showed that the AtRLK interacted with AtAVP1. Under low nitrogen stress, the Arabidopsis mutants rlk and avp1-1 had the same phenotypes. Discussion: These results indicate that AtAVP1 regulates low nitrogen stress responses by interacting with AtRLK, which provides a novel insight into the regulatory pathway related to H+-pyrophosphatase function in plants.

Overexpression of V-type H+ pyrophosphatase gene EdVP1 from Elymus dahuricus increases yield and potassium uptake of transgenic wheat under low potassium conditions.

Lack of potassium in soil limits crop yield. Increasing yield and conserving potassium ore requires improving K use efficiency (KUE). Many genes influence KUE in plants, but it is not clear how these genes function in the field. We identified the V-type H+-pyrophosphatase gene EdVP1 from Elymus dahurica. Gene expression analysis showed that EdVP1 was induced by low potassium stress. Protein subcellular localization analysis demonstrated that EdVP1 localized on the plasma membrane. We overexpressed EdVP1 in two wheat varieties and conducted K tolerance experiments across years. Yield per plant, grain number per spike, plant height, and K uptake of four transgenic wheat lines increased significantly compared with WT; results from two consecutive years showed that EdVP1 significantly increased yield and KUE of transgenic wheat. Pot experiments showed that transgenic plants had significantly longer shoots and roots, and higher K accumulation in shoots and roots and H+-PPase activity in shoots than WT under low K. A fluidity assay of potassium ion in EdVP1 transgenic plant roots showed that potassium ion influx and H+ outflow in transgenic plants were higher than WT. Overexpressing EdVP1 significantly improved yield and KUE of transgenic wheat and was related to higher K uptake capacity in root.