SiLNR1 -Mediated Nitrogen Regulatory Signaling Enhances Nitrogen Use Efficiency and Grain Yield in Foxtail Millet ( Setaria italica L.) under Low-Nitrogen Stress

Foxtail millet (Setaria italica L.), a C4 cereal crop domesticated in China, exhibits exceptional nitrogen use efficiency (NUtE) with marked genotypic variation; however, the molecular basis remains underexplored. Here, we identified candidate genes through a sequential filtering strategy from genetic linkage (bulked segregant analysis sequencing) to transcriptional response (RNA sequencing) within defined quantitative trait locus intervals to dissect NUtE mechanisms in contrasting genotypes: the low-nitrogen-tolerant variety Yugu28 and the low-nitrogen-sensitive variety Qiyehuang, to identify the genetic regulatory mechanisms. Several candidate genes were screened, such as glutamine synthetase (GS2, Seita.3G024100), glutathione S-transferase (GLUS, Seita.3G386000) and nitrogen regulatory protein P-II (Seita.3G051900). Seita.3G051900 was identified as LOW-NITROGEN REGULATORY GENE (SiLNR1), which is highly expressed in different organs. The functional characterization revealed that SiLNR1 overexpression results in strikingly low-N resilience: Compared with the wild-type controls, the transgenic lines presented 106% longer primary roots, 91.3% greater plant height, and 18.9% greater root nitrogen accumulation under low-nitrogen (LN) conditions and boosted grain yield and NUtE by 29.5% and 24.3%, respectively. By contrast, the silnr1 mutant presented markedly shorter primary roots and plant height and lower shoot N accumulation under LN conditions. Field validation of the SiLNR1 allele in the recombinant inbred line population demonstrated that the SiLNR1 allele from Yugu28 confers a substantial and measurable yield gain under LN field conditions. The functions of SiLNR1 associated with enhanced nitrogen uptake and utilization have been revealed, indicating that SiLNR1 contributes to nitrogen metabolism and NUtE positively, thus providing a theoretical basis and application prospect for resource conservation, environmental protection, and sustainable agricultural development.