Nitrate alleviates ammonium toxicity in Brassica napus by coordinating rhizosphere and cell pH and ammonium assimilation.

In natural and agricultural situations, ammonium ( NH 4 + $$ {\mathrm{NH}}_4^{+} $$ ) is a preferred nitrogen (N) source for plants, but excessive amounts can be hazardous to them, known as NH 4 + $$ {\mathrm{NH}}_4^{+} $$ toxicity. Nitrate ( NO 3 - $$ {\mathrm{NO}}_3^{-} $$ ) has long been recognized to reduce NH 4 + $$ {\mathrm{NH}}_4^{+} $$ toxicity. However, little is known about Brassica napus, a major oil crop that is sensitive to high NH 4 + $$ {\mathrm{NH}}_4^{+} $$ . Here, we found that NO 3 - $$ {\mathrm{NO}}_3^{-} $$ can mitigate NH 4 + $$ {\mathrm{NH}}_4^{+} $$ toxicity by balancing rhizosphere and intracellular pH and accelerating ammonium assimilation in B. napus. NO 3 - $$ {\mathrm{NO}}_3^{-} $$ increased the uptake of NO 3 - $$ {\mathrm{NO}}_3^{-} $$ and NH 4 + $$ {\mathrm{NH}}_4^{+} $$ under high NH 4 + $$ {\mathrm{NH}}_4^{+} $$ circumstances by triggering the expression of NO 3 - $$ {\mathrm{NO}}_3^{-} $$ and NH 4 + $$ {\mathrm{NH}}_4^{+} $$ transporters, while NO 3 - $$ {\mathrm{NO}}_3^{-} $$ and H + efflux from the cytoplasm to the apoplast was enhanced by promoting the expression of NO 3 - $$ {\mathrm{NO}}_3^{-} $$ efflux transporters and genes encoding plasma membrane H + -ATPase. In addition, NO 3 - $$ {\mathrm{NO}}_3^{-} $$ increased pH in the cytosol, vacuole, and rhizosphere, and down-regulated genes induced by acid stress. Root glutamine synthetase (GS) activity was elevated by NO 3 - $$ {\mathrm{NO}}_3^{-} $$ under high NH 4 + $$ {\mathrm{NH}}_4^{+} $$ conditions to enhance the assimilation of NH 4 + $$ {\mathrm{NH}}_4^{+} $$ into amino acids, thereby reducing NH 4 + $$ {\mathrm{NH}}_4^{+} $$ accumulation and translocation to shoot in rapeseed. In addition, root GS activity was highly dependent on the environmental pH. NO 3 - $$ {\mathrm{NO}}_3^{-} $$ might induce metabolites involved in amino acid biosynthesis and malate metabolism in the tricarboxylic acid cycle, and inhibit phenylpropanoid metabolism to mitigate NH 4 + $$ {\mathrm{NH}}_4^{+} $$ toxicity. Collectively, our results indicate that NO 3 - $$ {\mathrm{NO}}_3^{-} $$ balances both rhizosphere and intracellular pH via effective NO 3 - $$ {\mathrm{NO}}_3^{-} $$ transmembrane cycling, accelerates NH 4 + $$ {\mathrm{NH}}_4^{+} $$ assimilation, and up-regulates malate metabolism to mitigate NH 4 + $$ {\mathrm{NH}}_4^{+} $$ toxicity in oilseed rape.