Co-overexpression of genes for nitrogen transport, assimilation, and utilization boosts rice grain yield and nitrogen use efficiency

Nitrogen (N) fertilization is necessary for obtaining high rice yield. But excessive N fertilizer reduces rice plant N efficiency and causes negative effects such as environmental pollution. In this study, we assem-bled key genes involved in different nodes of N pathways to boost nitrate and ammonium uptake and assimilation, and to strengthen amino acid utilization to increase grain yield and nitrogen use efficiency (NUE) in rice. The combinations OsNPF8.9a x OsNR2, OsAMT1;2 x OsGS1;2 x OsAS1, and OsGS2 x OsAS2 x OsANT3 optimized nitrate assimilation, ammonium conversion, and N reutilization, respectively. In co-overexpressing rice lines obtained by co-transformation, the tiller number, biomass, and grain yield per plant of the OsAMT1;2 x OsGS1;2 x OsAS1-overexpressing line exceeded those of wild-type ZH11, the OsNPF8.9a x OsNR2 x OsGS1;2 x OsAS1-overexpressing line, and the OsGS2 x OsAS2 x OsANT3-overexpressing line. The glutamine synthase activity, free amino acids, and nitrogen utilization efficiency (NUtE) of the OsAMT1;2 x OsGS1;2 x OsAS1-overexpressing line exceeded those of ZH11 and other lines that combined key genes. N influx efficiency was increased in the OsAMT1;2 x OsGS1;2 x OsAS1-overexpressing line and OsNPF8.9a x OsNR2 x OsGS1;2 x OsAS1- overexpressing line under a low ammonium and a low nitrate treatment, respectively. We propose that combining overexpression of OsAMT1;2, OsGS1;2, and OsAS1 is a promising breeding strategy for system-atically increasing rice grain yield and NUE by focusing on key nodes in the N pathway. (c) 2022 Crop Science Society of China and Institute of Crop Science, CAAS. Production and hosting by Elsevier B.V. on behalf of KeAi Communications Co., Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).