NIN-LIKE PROTEIN3.2 inhibits repressor Aux/IAA14 expression and enhances root biomass in maize seedlings under low nitrogen.

Plants generally enhance their root growth in the form of greater biomass and/or root length to boost nutrient uptake in response to short-term low nitrogen (LN). However, the underlying mechanisms of short-term LN-mediated root growth remain largely elusive. Our genome-wide association study, haplotype analysis, and phenotyping of transgenic plants showed that the crucial nitrate signaling component NIN-LIKE PROTEIN3.2 (ZmNLP3.2), a positive regulator of root biomass, is associated with natural variations in root biomass of maize (Zea mays L.) seedlings under LN. The monocot-specific gene AUXIN/INDOLE-3-ACETIC ACID14 (ZmAux/IAA14) exhibited opposite expression patterns to ZmNLP3.2 in ZmNLP3.2 knockout and overexpression lines, suggesting that ZmNLP3.2 hampers ZmAux/IAA14 transcription. Importantly, ZmAux/IAA14 knockout seedlings showed a greater root dry weight (RDW), whereas ZmAux/IAA14 overexpression reduced RDW under LN compared with wild-type plants, indicating that ZmAux/IAA14 negatively regulates the RDW of LN-grown seedlings. Moreover, in vitro and vivo assays indicated that AUXIN RESPONSE FACTOR19 (ZmARF19) binds to and transcriptionally activates ZmAux/IAA14, which was weakened by the ZmNLP3.2-ZmARF19 interaction. The zmnlp3.2 ZmAux/IAA14-OE seedlings exhibited further reduced RDW compared to ZmAux/IAA14 overexpression lines when subjected to LN treatment, corroborating the ZmNLP3.2-ZmAux/IAA14 interaction. Thus, our study reveals a ZmNLP3.2-ZmARF19-ZmAux/IAA14 module regulating root biomass in response to nitrogen limitation in maize.

ZmNRT1.1B (ZmNPF6.6) determines nitrogen use efficiency via regulation of nitrate transport and signalling in maize.

Nitrate (NO3 - ) is crucial for optimal plant growth and development and often limits crop productivity under low availability. In comparison with model plant Arabidopsis, the molecular mechanisms underlying NO3 - acquisition and utilization remain largely unclear in maize. In particular, only a few genes have been exploited to improve nitrogen use efficiency (NUE). Here, we demonstrated that NO3 - -inducible ZmNRT1.1B (ZmNPF6.6) positively regulated NO3 - -dependent growth and NUE in maize. We showed that the tandem duplicated proteoform ZmNRT1.1C is irrelevant to maize seedling growth under NO3 - supply; however, the loss of function of ZmNRT1.1B significantly weakened plant growth under adequate NO3 - supply under both hydroponic and field conditions. The 15 N-labelled NO3 - absorption assay indicated that ZmNRT1.1B mediated the high-affinity NO3 - -transport and root-to-shoot NO3 - translocation. Transcriptome analysis further showed, upon NO3 - supply, ZmNRT1.1B promotes cytoplasmic-to-nuclear shuttling of ZmNLP3.1 (ZmNLP8), which co-regulates the expression of genes involved in NO3 - response, cytokinin biosynthesis and carbon metabolism. Remarkably, overexpression of ZmNRT1.1B in modern maize hybrids improved grain yield under N-limiting fields. Taken together, our study revealed a crucial role of ZmNRT1.1B in high-affinity NO3 - transport and signalling and offers valuable genetic resource for breeding N use efficient high-yield cultivars.

OsAMT1;1 and OsAMT1;2 Coordinate Root Morphological and Physiological Responses to Ammonium for Efficient Nitrogen Foraging in Rice.

Optimal plant growth and development rely on morphological and physiological adaptions of the root system to forage heterogeneously distributed nitrogen (N) in soils. Rice grows mainly in the paddy soil where ammonium (NH4+) is present as the major N source. Although root NH4+ foraging behaviors are expected to be agronomically relevant, the underlying mechanism remains largely unknown. Here, we showed that NH4+ supply transiently enhanced the high-affinity NH4+ uptake and stimulated lateral root (LR) branching and elongation. These synergistic physiological and morphological responses were closely related to NH4+-induced expression of NH4+ transporters OsAMT1;1 and OsAMT1;2 in roots. The two independent double mutants (dko) defective in OsAMT1;1 and OsAMT1;2 failed to induce NH4+ uptake and stimulate LR formation, suggesting that OsAMT1s conferred the substrate-dependent root NH4+ foraging. In dko plants, NH4+ was unable to activate the expression of OsPIN2, and the OsPIN2 mutant (lra1) exhibited a strong reduction in NH4+-triggered LR branching, suggesting that the auxin pathway was likely involved in OsAMT1s-dependent LR branching. Importantly, OsAMT1s-dependent root NH4+ foraging behaviors facilitated rice growth and N acquisition under fluctuating NH4+ supply. These results revealed an essential role of OsAMT1s in synergizing root morphological and physiological processes, allowing for efficient root NH4+ foraging to optimize N capture under fluctuating N availabilities.

Genome-wide analysis of zinc- and iron-regulated transporter-like protein family members in apple and functional validation of ZIP10.

Deficiency of zinc (Zn) and iron (Fe) is common in apple grown in orchards, which affects fruit yield and quality. However, the mechanisms of absorption and transport of Zn and Fe in apples are still unclear. In the present study, we aimed to identify MdZIP genes and explore the mechanism of response of MdZIPs to Zn and Fe deficiencies. Eighteen Zn- and Fe-regulated transporter-like protein (ZIP) family members were identified in apple (Malus domestica L.) and named according to their chromosomal location. Phylogenetic analysis divided MdZIPs into four groups, and the most closely related MdZIPs in the phylogenetic tree showed similar gene structures and protein motifs. Expression pattern analysis indicated that ZIP genes in apple were differentially expressed among tissues and developmental stages under Zn and Fe deficiency. The overexpression of MdZIP10 increased the content of Zn and Fe in Arabidopsis thaliana L. and MdZIP10 played crucial roles in the uptake and transport of Zn and Fe. MdZIP10 was able to rescue growth of Zn2+ and Fe2+ uptake defective yeast mutants under Zn2+ and Fe2+ deficient conditions, respectively. Symptoms of Zn and Fe deficiency were alleviated in the MdZIP10 transgenic plants. The expression of genes related to Fe and Zn uptake and transport was induced in the MdZIP10 transgenic plants, thereby stimulating endogenous Fe and Zn uptake and transport mechanisms. The present study lays the foundation for future functional analysis of ZIP genes in apple.

Overexpression of the Maize ZmNLP6 and ZmNLP8 Can Complement the Arabidopsis Nitrate Regulatory Mutant nlp7 by Restoring Nitrate Signaling and Assimilation.

Nitrate is a key nutrient that affects maize growth and yield, and much has yet to be learned about nitrate regulatory genes and mechanisms in maize. Here, we identified nine ZmNLP genes in maize and analyzed the functions of two ZmNLP members in nitrate signaling. qPCR results revealed a broad pattern of expression for ZmNLP genes in different stages and organs with the highest levels of transcript expression of ZmNLP6 and ZmNLP8. When ZmNLP6 and ZmNLP8 were overexpressed in the Arabidopsis nitrate regulatory gene mutant nlp7-4, nitrate assimilation and induction of nitrate-responsive genes in the transgenic plants were recovered to WT levels, indicating that ZmNLP6 and ZmNLP8 can replace the essential roles of the master nitrate regulatory gene AtNLP7 in nitrate signaling and metabolism. ZmNLP6 and ZmNLP8 are localized in the nucleus and can bind candidate nitrate-responsive cis-elements in vitro. The biomass and yield of transgenic Arabidopsis lines overexpressing ZmNLP6 and ZmNLP8 showed significant increase compared with WT and nlp7-4 mutant line in low nitrate conditions. Thus, ZmNLP6 and ZmNLP8 regulate nitrate signaling in transgenic Arabidopsis plants and may be potential candidates for improving nitrogen use efficiency of maize.