Gene Expression In Maize Research Articles

Effect of the B chromosome-located long non-coding RNAs on gene expression in maize

Using artificial chromosomes in maize breeding allows for site-specific integration of multigene stacks, effectively overcoming the limitations of conventional transgenic approaches. The maize B chromosome, which is dispensable and highly heterochromatic, has minimal impact on phenotypes at low copy numbers, making it a promising platform for engineering artificial chromosomes. However, recent studies have demonstrated that the maize B chromosome can impact gene expression and recombination on the A chromosome. Understanding the genic characteristics of the B chromosomes and their impact on gene expression is essential for their application in artificial chromosome construction. Despite advancements in elucidating how the B chromosome affects A chromosome expression, the role of long non-coding RNAs (lncRNAs) in this context remains unclear. In this study, we analyzed the RNA-seq data from leaf tissue of plants with 0 to 7 B chromosomes, identifying a total of 1614 lncRNAs, including 1516 A chromosome-located and 98 B chromosome-located lncRNAs, 72 of which are specific to the B chromosome. While A-located lncRNAs show greater dependence on the mere presence of the B chromosome, the expression of B-located lncRNAs is significantly affected by the number of B chromosomes present. Regulatory networks constructed in this study suggest that B-located lncRNAs may drive the differential expression of A chromosome-located transcription factors and genes associated with circadian rhythm regulation, indicating their regulatory role in A chromosome gene expression.

Cold mediates maize root hair developmental plasticity via epidermis-specific transcriptomic responses.

Cold stress during early development limits maize (Zea mays L.) production in temperate zones. Low temperatures restrict root growth and reprogram gene expression. Here, we provide a systematic transcriptomic landscape of maize primary roots, their tissues, and cell types in response to cold stress. The epidermis exhibited a unique transcriptomic cold response, and genes involved in root hair formation were dynamically regulated in this cell type by cold. Consequently, activation of genes involved in root hair tip growth contributed to root hair recovery under moderate cold conditions. The maize root hair defective mutants roothair defective 5 (rth5) and roothair defective 6 (rth6) displayed enhanced cold tolerance with respect to primary root elongation. Furthermore, DEHYDRATION RESPONSE ELEMENT-BINDING PROTEIN 2.1 (DREB2.1) was the only member of the dreb subfamily of AP2/EREB transcription factor genes upregulated in primary root tissues and cell types but exclusively downregulated in root hairs upon cold stress. Plants overexpressing dreb2.1 significantly suppressed root hair elongation after moderate cold stress. Finally, the expression of rth3 was regulated by dreb2.1 under cold conditions, while rth6 transcription was regulated by DREB2.1 irrespective of the temperature regime. We demonstrated that dreb2.1 negatively regulates root hair plasticity at low temperatures by coordinating the expression of root hair defective genes in maize.

Analysis of the 5' Untranslated Region Length-Dependent Control of Gene Expression in Maize: A Case Study with the ZmLAZ1 Gene Family.

The untranslated regions (UTRs) within plant mRNAs play crucial roles in regulating gene expression and the functionality of post-translationally modified proteins by various mechanisms. These regions are vital for plants' ability to sense to multiple developmental and environmental stimuli. In this study, we conducted a genome-wide analysis of UTRs and UTR-containing genes in maize (Zea mays). Using the ZmLAZ1 family as a case study, we demonstrated that the length of 5' UTRs could influence gene expression levels by employing GUS reporter gene assays. Although maize and arabidopsis (Arabidopsis thaliana), as well as rice (Oryza sativa), have distinct functional categories of UTR-containing genes, we observed a similar lengthwise distribution of UTRs and a recurring appearance of certain gene ontology (GO) terms between maize and rice. These suggest a potentially conserved mechanism within the Poaceae species. Furthermore, the analysis of cis-acting elements in these 5' UTRs of the ZmLAZ1 gene family further supports the hypothesis that UTRs confer functional specificity to genes in a length-dependent manner. Our findings offer novel insights into the role of UTRs in maize, contributing to the broader understanding of gene expression regulation in plants.

Maize lipid droplet-associated protein 2 is recruited by a virus to enhance viral multiplication and infection through regulating cellular fatty acid metabolism.

Pathogen infection induces massive reprogramming of host primary metabolism. Lipid and fatty acid (FA) metabolism is generally disrupted by pathogens and co-opted for their proliferation. Lipid droplets (LDs) that play important roles in regulating cellular lipid metabolism are utilized by a variety of pathogens in mammalian cells. However, the function of LDs during pathogenic infection in plants remains unknown. We show here that infection by rice black streaked dwarf virus (RBSDV) affects the lipid metabolism of maize, which causes elevated accumulation of C18 polyunsaturated fatty acids (PUFAs) leading to viral proliferation and symptom development. The overexpression of one of the two novel LD-associated proteins (LDAPs) of maize (ZmLDAP1 and ZmLDAP2) induces LD clustering. The core capsid protein P8 of RBSDV interacts with ZmLDAP2 and prevents its degradation through the ubiquitin-proteasome system mediated by a UBX domain-containing protein, PUX10. In addition, silencing of ZmLDAP2 downregulates the expression of FA desaturase genes in maize, leading to a decrease in C18 PUFAs levels and suppression of RBSDV accumulation. Our findings reveal that plant virus may recruit LDAP to regulate cellular FA metabolism to promote viral multiplication and infection. These results expand the knowledge of LD functions and viral infection mechanisms in plants.

Enhancing maize resilience to microplastics and cadmium through nanoparticles intervention

Microplastic (MPs) and heavy metals (HMs) pollution present significant ecological and environmental threats due to their widespread production and release into natural ecosystems. This study aimed to investigate the individual effects of polyvinyl chloride microplastics (PVC-MPs) and cadmium (Cd) on various aspects of plant growth, as well as the potential ameliorative role of titanium dioxide nanoparticles (TiO2 NPs) in mitigating their adverse impacts. The experiment involved exposing maize seedlings to different concentrations of PVC-MPs (0, 2, and 4 mg L−1) and Cd (0, 50, and 100 mg kg−1), along with varying levels of TiO2NPs (50 µg mL−1). Our findings showed that Cd toxicity in the soil significantly reduced growth, gas exchange properties, sugar content, ascorbate-glutathione (ASA-GSH) cycle activity, cellular composition, and proline metabolism in maize. In contrast, Cd contaminating greatly boosted oxidative stress indicators, enzymatic and non-enzymatic antioxidants, and gene expression in maize. Stimulatingly, the use of TiO2NPs caused in a significant development in growth of plant, plant biomass, gas exchange parameters, enzymatic antioxidants and non-enzymatic antioxidant activity, and gene expressions of antioxidants enzymes, lowering oxidative stress (MDA and H2O2) and Cd absorption in maize plants. Furthermore, TiO2NPs increased cellular fractionation (cellulose and hemicellulose), altering proline metabolism and ASA-GSH cycle activity in maize plants. Overall, this work emphasizes the intricate interactions between MPs and Cd, and nanoparticles in plant systems, highlighting TiO2NPs as a possible strategy for increasing plant flexibility and minimizing the negative effects of environmental pollutants. These findings have crucial implications regarding the development of long-term agricultural management approaches in polluted areas.

Genome-wide identification and analyses of ZmAPY genes reveal their roles involved in maize development and abiotic stress responses

Apyrase is a class of enzyme that catalyzes the hydrolysis of nucleoside triphosphates/diphosphates (NTP/NDP), which widely involved in regulation of plant growth and stress responses. However, apyrase family genes in maize have not been identified, and their characteristics and functions are largely unknown. In this study, we identified 16 apyrases (named as ZmAPY1-ZmAPY16) in maize genome, and analyzed their phylogenetic relationships, gene structures, chromosomal distribution, upstream regulatory transcription factors and expression patterns. Analysis of the transcriptome database unveiled tissue-specific and abiotic stress-responsive expression of ZmAPY genes in maize. qPCR analysis further confirmed their responsiveness to drought, heat, and cold stresses. Association analyses indicated that variations of ZmAPY5 and ZmAPY16 may regulate maize agronomic traits and drought responses. Our findings shed light on the molecular characteristics and evolutionary history of maize apyrase genes, highlighting their roles in various biological processes and stress responses. This study forms a basis for further exploration of apyrase functions in maize.

Unveiling the Regulatory Role of miRNAs in Internode Elongation: Integrated Analysis of MicroRNA and mRNA Expression Profiles across Diverse Dwarfing Treatments in Maize (Zea mays L.).

MicroRNAs are crucial regulators of gene expression in maize. However, the mechanisms through which miRNAs control internode elongation remain poorly understood. This study engineered varying levels of internode elongation inhibition, revealing that dwarfing treatments diminished gibberellin levels, curtailed cell longitudinal growth, and slowed the rate of internode elongation. Comprehensive transcriptome and miRNA profiling of the internode elongation zone showed gene expression changes that paralleled the extent of the internode length reduction. We identified 543 genes and 29 miRNAs with significant correlations to internode length, predominantly within families, including miR164 and miR396. By incorporating target gene expression levels, we pinpointed nine miRNA-mRNA pairs that are significantly associated with the regulation of the internode elongation. The inhibitory effects of these miRNAs on their target genes were confirmed through dual-luciferase reporter assays. Overexpression of miR164h in maize resulted in increased internode and cell length, suggesting a novel genetic avenue for manipulating plant stature. These miRNAs may also serve as precise spatiotemporal regulators for in vitro plant development.

Fulvic acid alleviates the stress of low nitrogen on maize by promoting root development and nitrogen metabolism.

The potential of fulvic acid (FA) to improve plant growth has been acknowledged, but its effect on plant growth and nutrient uptake under nutrient stress remains unclear. This study investigated the effects of different FA application rates on maize growth and nitrogen utilization under low nitrogen stress. The results showed that under low nitrogen stress, FA significantly stimulated maize growth, particularly root development, biomass, and nitrogen content. The enhanced activity levels of key enzymes in nitrogen metabolism were observed, along with differential gene expression in maize, which enriched nitrogen metabolism, amino acid metabolism and plant hormone metabolism. The application of FA regulated the hormones' level, reduced abscisic acid content in leaves and Me-JA content in roots, and increased auxin and zeatin ribose content in leaves. This study concludes that, by promoting root development, nitrogen metabolism, and hormone metabolism, an appropriate concentration of FA can enhance plant tolerance to low nitrogen conditions and improve nitrogen use efficiency.

Natural methylation epialleles correlate with gene expression in maize.

DNA methylation in plants is depleted from cis regulatory elements in and near genes but is present in some gene bodies, including exons. Methylation in exons solely in the CG context is called gene body methylation (gbM). Methylation in exons in both CG and non-CG contexts is called TE-like methylation (teM). Assigning functions to both forms of methylation in genes has proven to be challenging. Toward that end, we utilized recent genome assemblies, gene annotations, transcription data, and methylome data to quantify common patterns of gene methylation and their relations to gene expression in maize. We found that gbM genes exist in a continuum of CG methylation levels without a clear demarcation between unmethylated genes and gbM genes. Analysis of expression levels across diverse maize stocks and tissues revealed a weak but highly significant positive correlation between gbM and gene expression except in endosperm. gbM epialleles were associated with an approximately 3% increase in steady-state expression level relative to unmethylated epialleles. In contrast to gbM genes, which were conserved and were broadly expressed across tissues, we found that teM genes, which make up about 12% of genes, are mainly silent, are poorly conserved, and exhibit evidence of annotation errors. We used these data to flag teM genes in the 26 NAM founder genome assemblies. While some teM genes are likely functional, these data suggest that the majority are not, and their inclusion can confound interpretation of whole-genome studies.

ZmWRKY70 activates the expression of hypoxic responsive genes in maize and enhances tolerance to submergence in Arabidopsis

Hypoxic stress due to submergence is a serious threat to the growth and development of maize. WRKY transcription factors are significant regulators of plant responses to various abiotic and biotic stresses. Nevertheless, their function and regulatory mechanisms in the resistance of maize to submergence stress remain unclear. Here we report the cloning of a maize WRKY transcription factor gene, ZmWRKY70, transcripts of which accumulate under submergence stress in maize seedlings. Subcellular localization analysis and yeast transcriptional activation assay indicated that ZmWRKY70 was localized in the nucleus and had transcriptional activation activity. Heterologous overexpression of ZmWRKY70 in Arabidopsis increased the tolerance of seeds and seedlings to submergence stress by upregulating the transcripts of several key genes involved in anaerobic respiration, such as group VII ethylene-responsive factor (ERFVII) (AtRAP2.2), alcohol dehydrogenase (AtADH1), pyruvate decarboxylase (AtPDC1/2), and sucrose synthase (AtSUS4), under submergence conditions. Moreover, the overexpression of ZmWRKY70 in maize mesophyll protoplasts enhanced the expression of ZmERFVII members (ZmERF148, ZmERF179, and ZmERF193), ZmADH1, ZmPDC2/3, and ZmSUS1. Yeast one-hybrid and dual-luciferase activity assays further confirmed that ZmWRKY70 enhanced the expression of ZmERF148 by binding to the W box motif located in the promoter region of ZmERF148. Together, these results indicate that ZmWRKY70 plays a significant role in tolerance of submergence stress. This work provides a theoretical basis, and suggests excellent genes, for biotechnological breeding to improve the tolerance of maize to submergence through the regulation of ZmWRKY genes.

Transcription Factor ZmNAC20 Improves Drought Resistance by Promoting Stomatal Closure and Activating Expression of Stress-Responsive Genes in Maize.

Drought is a major environmental threat that limits crop growth, development, and productivity worldwide. Improving drought resistance with genetic engineering methods is necessary to tackle global climate change. It is well known that NAC (NAM, ATAF and CUC) transcription factors play a critical role in coping with drought stress in plants. In this study, we identified an NAC transcription factor ZmNAC20, which regulates drought stress response in maize. ZmNAC20 expression was rapidly upregulated by drought and abscisic acid (ABA). Under drought conditions, the ZmNAC20-overexpressing plants had higher relative water content and survival rate than the wild-type maize inbred B104, suggesting that overexpression of ZmNAC20 improved drought resistance in maize. The detached leaves of ZmNAC20-overexpressing plants lost less water than those of wild-type B104 after dehydration. Overexpression of ZmNAC20 promoted stomatal closure in response to ABA. ZmNAC20 was localized in the nucleus and regulated the expression of many genes involved in drought stress response using RNA-Seq analysis. The study indicated that ZmNAC20 improved drought resistance by promoting stomatal closure and activating the expression of stress-responsible genes in maize. Our findings provide a valuable gene and new clues on improving crop drought resistance.

The Integration of Metabolomics and Transcriptomics Provides New Insights for the Identification of Genes Key to Auxin Synthesis at Different Growth Stages of Maize.

As a staple food crop, maize is widely cultivated worldwide. Sex differentiation and kernel development are regulated by auxin, but the mechanism regulating its synthesis remains unclear. This study explored the influence of the growth stage of maize on the secondary metabolite accumulation and gene expression associated with auxin synthesis. Transcriptomics and metabonomics were used to investigate the changes in secondary metabolite accumulation and gene expression in maize leaves at the jointing, tasseling, and pollen-release stages of plant growth. In total, 1221 differentially accumulated metabolites (DAMs) and 4843 differentially expressed genes (DEGs) were screened. KEGG pathway enrichment analyses of the DEGs and DAMs revealed that plant hormone signal transduction, tryptophan metabolism, and phenylpropanoid biosynthesis were highly enriched. We summarized the key genes and regulatory effects of the tryptophan-dependent auxin biosynthesis pathways, giving new insights into this type of biosynthesis. Potential MSTRG.11063 and MSTRG.35270 and MSTRG.21978 genes in auxin synthesis pathways were obtained. A weighted gene co-expression network analysis identified five candidate genes, namely TSB (Zm00001d046676 and Zm00001d049610), IGS (Zm00001d020008), AUX2 (Zm00001d006283), TAR (Zm00001d039691), and YUC (Zm00001d025005 and Zm00001d008255), which were important in the biosynthesis of both tryptophan and auxin. This study provides new insights for understanding the regulatory mechanism of auxin synthesis in maize.

Rescue of the first alphanucleorhabdovirus entirely from cloned complementary DNA: An efficient vector for systemic expression of foreign genes in maize and insect vectors.

Recent reverse genetics technologies have enabled genetic manipulation of plant negative-strand RNA virus (NSR) genomes. Here, we report construction of an infectious clone for the maize-infecting Alphanucleorhabdovirus maydis, the first efficient NSR vector for maize. The full-length infectious clone was established using agrobacterium-mediated delivery of full-length maize mosaic virus (MMV) antigenomic RNA and the viral core proteins (nucleoprotein N, phosphoprotein P, and RNA-directed RNA polymerase L) required for viral transcription and replication into Nicotiana benthamiana. Insertion of intron 2 ST-LS1 into the viral L gene increased stability of the infectious clone in Escherichia coli and Agrobacterium tumefaciens. To monitor virus infection in vivo, a green fluorescent protein (GFP) gene was inserted in between the N and P gene junctions to generate recombinant MMV-GFP. Complementary DNA (cDNA) clones of MMV-wild type (WT) and MMV-GFP replicated in single cells of agroinfiltrated N.benthamiana. Uniform systemic infection and high GFP expression were observed in maize inoculated with extracts of the infiltrated N.benthamiana leaves. Insect vectors supported virus infection when inoculated via feeding on infected maize or microinjection. Both MMV-WT and MMV-GFP were efficiently transmitted to maize by planthopper vectors. The GFP reporter gene was stable in the virus genome and expression remained high over three cycles of transmission in plants and insects. The MMV infectious clone will be a versatile tool for expression of proteins of interest in maize and cross-kingdom studies of virus replication in plant and insect hosts.

Heat-Stress-Mitigating Effects of a Protein-Hydrolysate-Based Biostimulant Are Linked to Changes in Protease, DHN, and HSP Gene Expression in Maize

The growth-promoting and heat-mitigating effects of a commercially available protein-hydrolysate-based biostimulant, Kaishi, during the early vegetative stage was investigated by applying it as a foliar spray on soil-grown maize plants or in the nutrient solution of hydroponically grown plants. At 10−3 dilution, the biostimulant inhibited germination and delayed the growth progress, while at 10−6–10−12 dilutions, it promoted shoot and root growth. Heat stress caused biomass reduction, decreased leaf pigment content and the chlorophyll a/chlorophyll b (chl a/b) ratio, caused starch depletion, and increased lipid peroxidation. Kaishi priming resulted in the substantial mitigation of negative stress effects, maintaining growth, stabilizing pigment content and the chl a/b ratio, restoring the leaf starch content, lowering the malondialdehyde (MDA) level, and significantly increasing the free proline content. The expression profiles of a set of genes coding for heat shock proteins (HSPs), dehydrins (DHNs), and proteases were analysed using qRT-PCR after heat stress exposure. The biostimulant-treated plants had higher transcript levels of certain HSPs, DHNs, and protease-coding genes, which remained stable or increased after the applied stress. The results demonstrate that very low concentrations of the biostimulant exerted stress-mitigating effects that could be linked to organ-specific changes in the gene expression of certain stress-inducible proteins.

Effect of aneuploidy of a non-essential chromosome on gene expression in maize.

SUMMARYThe non‐essential supernumerary maize (Zea mays) B chromosome (B) has recently been shown to contain active genes and to be capable of impacting gene expression of the A chromosomes. However, the effect of the B chromosome on gene expression is still unclear. In addition, it is unknown whether the accumulation of the B chromosome has a cumulative effect on gene expression. To examine these questions, the global expression of genes, microRNAs (miRNAs), and transposable elements (TEs) of leaf tissue of maize W22 plants with 0–7 copies of the B chromosome was studied. All experimental genotypes with B chromosomes displayed a trend of upregulated gene expression for a subset of A‐located genes compared to the control. Over 3000 A‐located genes are significantly differentially expressed in all experimental genotypes with the B chromosome relative to the control. Modulations of these genes are largely determined by the presence rather than the copy number of the B chromosome. By contrast, the expression of most B‐located genes is positively correlated with B copy number, showing a proportional gene dosage effect. The B chromosome also causes increased expression of A‐located miRNAs. Differentially expressed miRNAs potentially regulate their targets in a cascade of effects. Furthermore, the varied copy number of the B chromosome leads to the differential expression of A‐located and B‐located TEs. The findings provide novel insights into the function and properties of the B chromosome.

Dot Blot Assay for Assessing Trehalose-6-phosphate Synthase Gene Expression in a Maize Breeding Population under Water Stress

Field maize is an important economic crop grown around the world and it has been mainly used in the animal feed industry. Maize yields have been inadequate for the demand due to drought events. One way to alleviate yield losses is to develop drought tolerant maize varieties for farmers. Trehalose-6-phosphate synthase (TPS) is an important enzyme involved in trehalose biosynthesis which has been found to increase plant tolerance to abiotic stresses. The aim of this research was to screen the levels of TPS gene expression in maize breeding materials under water stress via dot-blot hybridization using cDNA probe. To do so, 34 S2 maize families were grown and subjected to water stress condition. Leave samples were collected at 6 different days after planting (DAP) for a dot blot assay. The results showed that the level of TPS gene expression was highest at 4 days after stress (relative intensity at 64 DAP). However, dot blotting at 6 days after stress (relative intensity at 66 DAP) was effective to differentiate maize families. Furthermore, a moderate negative relationship between relative signal intensity at 66 DAP (RI66) and Smith index based on multi-phenotypic traits was found to be statistically significant. Our study showed that maize with high TPS gene expression tended to be less tolerant to water stress. It is noteworthy that the study of TPS gene expression in mature maize under stress in this study showed results that contrasted with previous reports on seedlings in many plant species. Furthermore, we found that 4 out of 34 S2 maize families may have potential for further use in our breeding program.

Analysis of Global Gene Expression in Maize (Zea mays) Vegetative and Reproductive Tissues That Differ in Accumulation of Starch and Sucrose.

Carbon allocation between vegetative and reproductive tissues impacts cereal grain production. Despite great agricultural importance, sink–source relationships have not been fully characterized at the early reproductive stages in maize. Here, we quantify the accumulation of non-structural carbohydrates and patterns of gene expression in the top internode of the stem and the female inflorescence of maize at the onset of grain filling (reproductive stage R1). Top internode stem and female inflorescence tissues of the Puma maize inbred line were collected at reproductive stage R1 (without pollination) and non-structural carbohydrates were quantified by spectrophotometry. The female inflorescence accumulated starch at higher levels than the top internode of the stem. Global mRNA transcript levels were then evaluated in both tissues by RNA sequencing. Gene expression analysis identified 491 genes differentially expressed between the female inflorescence and the top stem internode. Gene ontology classification of differentially expressed genes showed enrichment for sucrose synthesis, the light-dependent reactions of photosynthesis, and transmembrane transporters. Our results suggest that sugar transporters play a key role in sugar partitioning in the maize stem and reveal previously uncharacterized differences between the female inflorescence and the top internode of the stem at early reproductive stages.

Insight into maize gene expression profiles responses to symbiotic bacteria derived from Helicoverpa armigera and Ostrinia furnacalis.

The insects of Ostrinia furnacalis and Helicoverpa armigera are the two main pests that affect maize growth, which significantly decrease the yield. Plants induce various immune-related pathways to antagonize insect feeding during insect-plant interactions. Moreover, different insect elicitors or effectors participate in the interactions via releasing into plants. While there are many bacteria during insect regurgitation, their roles in insect-plant interaction are unknown. In this study, four bacterial strains were isolated from regurgitation fluid of O. furnacalis and H. armigera, and their cultures were inoculated on maize leaves for response analysis. All the four bacterial strains altered gene expression profiles in maize, and these altered expression profiles included phytohormones, secondary metabolic pathways, transcription factors, MAPK, and plant-pathogen interaction-related genes. A total of 210 genes, such as WRKY54, WRKY62, PIF5, argonaute 1, Xa21, NRR, ubiquitin-proteasome system genes, were co-changed in response to bacterial inoculation. These changes were similar with maize gene profile changes after insect feeding. Symbiotic insect bacteria participate in insect-plant interactions by changing maize gene expression profiles, which might be used to develop anti-pest microbial agents by activating plant defense system with identified microbes. In future, understanding the roles of symbiotic insect bacteria on plant-insect interaction might provide a promising and novel strategy for pest biocontrol using microbes.

Mutation of ZmWRKY86 confers enhanced salt stress tolerance in maize

As one of the largest families of transcription factors in plants, the WRKY proteins play crucial roles in plant growth and development, defense regulation and stress responses. In this research, ZmWRKY86 encoding a WRKY transcription factor was cloned from maize (Zea mays L.). ZmWRKY86 expression was up-regulated by salt stress. ZmWRKY86 is a nuclear protein and has no transcriptional activation ability in yeast. ZmWRKY86 can specifically bind to W-box (TTGACC), which was confirmed by electrophoretic mobility shift assay (EMSA) and dual-LUC experiments. As compared with control, the wrky86 mutants showed enhanced plant tolerance to salt stress with higher survival rate, catalase activity and K+ content, but lower malondialdehyde accumulation, relative electrical leakage level and Na+ content under salt-stress condition. Transcriptome and quantitative real-time PCR analyses indicated that the mutation of ZmWRKY86 led to significant changes in the expression of stress-related genes in maize. Further assays showed that ZmWRKY86 directly interacted with the promoter of two salt stress-related genes Zm00001d020840 and Zm00001d046813. In summary, we identified a WRKY transcription factor ZmWRKY86 that participates in salt stress regulation through controlling the expression of stress-related genes.

Conserved noncoding sequences provide insights into regulatory sequence and loss of gene expression in maize.

Thousands of species will be sequenced in the next few years; however, understanding how their genomes work, without an unlimited budget, requires both molecular and novel evolutionary approaches. We developed a sensitive sequence alignment pipeline to identify conserved noncoding sequences (CNSs) in the Andropogoneae tribe (multiple crop species descended from a common ancestor ∼18 million years ago). The Andropogoneae share similar physiology while being tremendously genomically diverse, harboring a broad range of ploidy levels, structural variation, and transposons. These contribute to the potential of Andropogoneae as a powerful system for studying CNSs and are factors we leverage to understand the function of maize CNSs. We found that 86% of CNSs were comprised of annotated features, including introns, UTRs, putative cis-regulatory elements, chromatin loop anchors, noncoding RNA (ncRNA) genes, and several transposable element superfamilies. CNSs were enriched in active regions of DNA replication in the early S phase of the mitotic cell cycle and showed different DNA methylation ratios compared to the genome-wide background. More than half of putative cis-regulatory sequences (identified via other methods) overlapped with CNSs detected in this study. Variants in CNSs were associated with gene expression levels, and CNS absence contributed to loss of gene expression. Furthermore, the evolution of CNSs was associated with the functional diversification of duplicated genes in the context of maize subgenomes. Our results provide a quantitative understanding of the molecular processes governing the evolution of CNSs in maize.