Enhancing nitrogen use efficiency (NUE) in crop plants is an important breeding target to reduce excessive use of chemical fertilizers, with substantial benefits to farmers and the environment. In Arabidopsis (Arabidopsis thaliana), allocation of more NO 3 2 to shoots was associated with higher NUE; however, the commonality of this process across plant species have not been sufficiently studied. Two Brassica napus genotypes were identified with high and low NUE. We found that activities of V-ATPase and V-PPase, the two tonoplast proton-pumps, were significantly lower in roots of the high-NUE genotype (Xiangyou15) ] S:R ratios were significantly higher in Xiangyou15. BnNRT1.5 expression was higher in roots of Xiangyou15 compared with 814, while BnNRT1.8 expression was lower. In both B. napus treated with proton pump inhibitors or Arabidopsis mutants impaired in proton pump activity, vacuolar sequestration capacity (VSC) of NO 3 2 in roots substantially decreased. Expression of NRT1.5 was up-regulated, but NRT1.8 was down-regulated, driving greater NO 3 2 long-distance transport from roots to shoots. NUE in Arabidopsis mutants impaired in proton pumps was also significantly higher than in the wild type col-0. Taken together, these data suggest that decrease in VSC of NO 3 2 in roots will enhance transport to shoot and essentially contribute to higher NUE by promoting NO 3 2 allocation to aerial parts, likely through coordinated regulation of NRT1.5 and NRT1.8.
Nitrate, once taken up by plants, can either be stored in vacuoles or reduced by nitrate reductase in the cytoplasm. High accumulation of NO 3 -in the vacuole occurs when assimilation into the cytoplasm is saturated. This study elucidates how proton pumps at the tonoplast (V-ATPase and V-PPase) affect the NO 3 -content of Brassica napus by controlling the distribution of NO 3 -between the cytoplasm and vacuole. Pot experiments were conducted in a greenhouse under normal N (15.0 mM nitrate) conditions using B. napus genotypes that demonstrated either high (Xiangyou15) or low (814) nitrogen use efficiency (NUE). The NO 3 -content of the high NUE genotype was significantly lower than that of the low NUE genotype, whereas the total N per plant of the two genotypes was almost the same, suggesting that the different NUE between the two genotypes is not due to the difference of NO 3 -uptake. The relative expression levels of V-ATPase (vha-a3) and V-PPase (avp1) genes in the high NUE genotype were significantly lower than in the low NUE genotype, resulting in lower V-ATPase and V-PPase activities in the high NUE genotype. The transport of NO 3 -and protons from the cytoplasm to the vacuole is powered by V-ATPase and V-PPase, so their lower activities increase H ? efflux from and reduce NO 3 -influx into the vacuoles of the high NUE genotype. We conclude that the lower activity of proton pumps at the tonoplast is the main reason the high NUE genotype possesses lower NO 3 -content and higher N-use efficiency.
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