Anthocyanins are a class of antioxidants that scavenge free radicals in cells and play an important role in promoting human health and preventing many diseases. Here, we characterized a maize Bronze gene (BZ1) from the purple colored W22 introgression line, which encodes an anthocyanin 3-O-glucosyltransferase, a key enzyme in the anthocyanin synthesis pathway. Mutation of ZmBZ1 showed bronze-colored seeds and reduced anthocyanins in seeds aleurone layer, seedlings coleoptile, and stem of mature plants by comparison with purple colored W22 (WT). Furthermore, we proved that maize BZ1 is an aleurone layer-specific expressed protein and sub-located in cell nucleus. Real-time tracing of the anthocyanins in developing seeds demonstrated that the pigment was visible from 16 DAP (day after pollination) in field condition, and first deposited in the crown part then spread all over the seed. Additionally, it was transferred along with the embryo cell activity during seed germination, from aleurone layer to cotyledon and coleoptile, as confirmed by microscopy and real-time qRT-PCR. Finally, we demonstrated that the ZmBZ1 contributes to stress tolerance, especially salinity. Further study proved that ZmBZ1 participates in reactive oxygen scavenging (ROS) by accumulating anthocyanins, thereby enhancing the tolerance to abiotic stress.

Anthocyanin, a kind of flavonoid, plays a crucial role in plant resistance to abiotic stress. Salt stress is a kind of abiotic stress that can damage the growth and development of plant seedlings. However, limited research has been conducted on the involvement of maize seedlings in salt stress resistance via anthocyanin accumulation, and its potential molecular mechanism is still unclear. Therefore, it is of great significance for the normal growth and development of maize seedlings to explore the potential molecular mechanism of anthocyanin improving salt tolerance of seedlings via transcriptome analysis. In this study, we identified two W22 inbred lines (tolerant line pur–W22 and sensitive line bro–W22) exhibiting differential tolerance to salt stress during seedling growth and development but showing no significant differences in seedling characteristics under non–treatment conditions. In order to identify the specific genes involved in seedlings’ salt stress response, we generated two recombinant inbred lines (RIL^pur–W22 and RIL^bro–W22) by crossing pur–W22 and bro–W22, and then performed transcriptome analysis on seedlings grown under both non–treatment and salt treatment conditions. A total of 6100 and 5710 differentially expressed genes (DEGs) were identified in RIL^pur–W22 and RIL^bro–W22 seedlings, respectively, under salt–stressed conditions when compared to the non–treated groups. Among these DEGs, 3160 were identified as being present in both RIL^pur–W22 and RIL^bro–W22, and these served as commonly stressed EDGs that were mainly enriched in the redox process, the monomer metabolic process, catalytic activity, the plasma membrane, and metabolic process regulation. Furthermore, we detected 1728 specific DEGs in the salt–tolerant RIL^pur–W22 line that were not detected in the salt–sensitive RIL^bro–W22 line, of which 887 were upregulated and 841 were downregulated. These DEGs are primarily associated with redox processes, biological regulation, and the plasma membrane. Notably, the anthocyanin synthesis related genes in RIL^pur–W22 were strongly induced under salt treatment conditions, which was consistented with the salt tolerance phenotype of its seedlings. In summary, the results of the transcriptome analysis not only expanded our understanding of the complex molecular mechanism of anthocyanin in improving the salt tolerance of maize seedlings, but also, the DEGs specifically expressed in the salt–tolerant line (RIL^pur–W22) provided candidate genes for further genetic analysis.

The germination, seedling growth, and crop productivity of maize seeds are significantly impacted by seed aging. This study investigated the efficacy of silver nanoparticles (AgNPs) as a seed priming agent for maize inbred lines exhibiting varying degrees of aging tolerance. Two inbred lines, aging-sensitive I178 and aging-tolerant X178, were used. AgNP treatment significantly promoted the germination of I178 (from 55% to 85%, compared with water treatment). Notable improvements were observed in root length, shoot length, and lateral root formation after AgNP treatment in I178. However, X178 showed no significant changes in germination and seedling growth after the AgNP treatment. Further transcriptomic analysis was performed on X178 and I178 before (water treatment) and after AgNP treatment to study genes and the expression network of the mechanism induced by AgNP promotion. In I178, AgNP treatment led to a substantial increase in differentially expressed genes (DEGs). A total of 800 DEGs were identified, with 517 being upregulated and 283 downregulated. The DEGs in I178 were mainly involved in metabolic processes, stress responses, and membrane repair. For example, genes related to lipid metabolism and membrane integrity were upregulated, along with seven genes associated with antioxidant action and redox metabolism. This indicates that AgNPs might enhance membrane stability and stress tolerance in I178. In contrast, X178 had a limited transcriptomic response to AgNP treatment. Although 874 DEGs were detected, the number of genes related to key processes like those in I178 did not change significantly, which is in line with its inherent aging tolerance. Overall, these results suggest that AgNPs can effectively improve seed vigor and counteract the negative effects of seed aging, especially in aging-sensitive maize lines. The mechanism likely occurs through regulating gene expression related to stress response, metabolic repair, and membrane stability. This study provides new insights into the molecular basis of AgNP-mediated seed vigor enhancement, which has potential implications for improving seed quality in agricultural production.

Plant seedling morphogenesis is considerably related to photosynthesis, pigment synthesis, and circadian periodicity during seedling development. We identified and cloned a maize zebra or crossbanding leaves mutant wk3735, which produces pale white kernels and was identified and plays a role in the equilibrium of the Redox state the in/out of ETC by active oxygen scavenging. Interestingly, it produces the zebra leaves during the production of the first seven leaves, which is apparently different from the mutation of homologs AtPTOX in Arabidopsis. It is intriguing to investigate how and why yellow crossbands (zebra leaf phenotype) emerge on leaves. As expected, chlorophyll concentration and photosynthetic efficiency both significantly declined in the yellow sector of wk3735 leaves. Meanwhile, we observed the circadian expression pattern of ZmPTOX1, which was further validated by protein interaction assays of the circadian clock protein TIM1 and ZmPTOX1. The transcriptome data of yellow (muW) and green (muG) sectors of knock-out lines and normal leaves of overexpression lines (OE) at the 5th-leaf seedling stage were analyzed. Zebra leaf etiolated sections exhibit a marked defect in the expression of genes involved in the circadian rhythm and rhythmic stress (light and cold stress) responses than green sections. According to the analysis of co-DEGs of muW vs. OE and muG vs. OE, terms linked to cell repair function were upregulated while those linked to environmental adaptability and stress response were downregulated due to the mutation of ZmPTOX1. Further gene expression level analyses of reactive oxygen species (ROS) scavenging enzymes and detection of ROS deposition indicated that ZmPTOX1 played an essential role in plant stress resistance and ROS homeostasis. The pleiotropic roles of ZmPTOX1 in plant ROS homeostasis maintenance, stress response, and circadian rhythm character may collectively explain the phenotype of zebra leaves during wk3735 seedling development.