Identification and functional analysis of a chromosome 2D fragment harboring TaFPF1 gene with the potential for yield improvement using a late heading wheat mutant.

KEY MESSAGE: A chromosome fragment influencing wheat heading and grain size was identified using mapping of m406 mutant. The study of TaFPF1 in this fragment provides more insights into wheat yield improvement. In recent years, wheat production has faced formidable challenges driven by rapid population growth and climate change, emphasizing the importance of improving specific agronomic traits such as heading date, spike length, and grain size. To identify potential genes for improving these traits, we screened a wheat EMS mutant library and identified a mutant, designated m406, which exhibited a significantly delayed heading date compared to the wild-type. Intriguingly, the mutant also displayed significantly longer spike and larger grain size. Genetic analysis revealed that a single recessive gene was responsible for the delayed heading. Surprisingly, a large 46.58 Mb deletion at the terminal region of chromosome arm 2DS in the mutant was identified through fine mapping and fluorescence in situ hybridization. Thus, the phenotypes of the mutant m406 are controlled by a group of linked genes. This deletion encompassed 917 annotated high-confidence genes, including the previously studied wheat genes Ppd1 and TaDA1, which could affect heading date and grain size. Multiple genes in this region probably contribute to the phenotypes of m406. We further investigated the function of TaFPF1 using gene editing. TaFPF1 knockout mutants showed delayed heading and increased grain size. Moreover, we identified the direct upstream gene of TaFPF1 and investigated its relationship with other important flowering genes. Our study not only identified more genes affecting heading and grain development within this deleted region but also highlighted the potential of combining these genes for improvement of wheat traits.

ZmNAC17 Regulates Mesocotyl Elongation by Mediating Auxin and ROS Biosynthetic Pathways in Maize.

The mesocotyl is of great significance in seedling emergence and in responding to biotic and abiotic stress in maize. The NAM, ATAF, and CUC2 (NAC) transcription factor family plays an important role in maize growth and development; however, its function in the elongation of the maize mesocotyl is still unclear. In this study, we found that the mesocotyl length in zmnac17 loss-of-function mutants was lower than that in the B73 wild type. By using transcriptomic sequencing technology, we identified 444 differentially expressed genes (DEGs) between zmnac17-1 and B73, which were mainly enriched in the "tryptophan metabolism" and "antioxidant activity" pathways. Compared with the control, the zmnac17-1 mutants exhibited a decrease in the content of indole acetic acid (IAA) and an increase in the content of reactive oxygen species (ROS). Our results provide preliminary evidence that ZmNAC17 regulates the elongation of the maize mesocotyl.

Mediation of Zinc and Iron Accumulation in Maize by ZmIRT2, a Novel Iron-Regulated Transporter.

Iron (Fe) is an essential micronutrient for plant growth. Iron-regulated transporters (IRTs) play important roles in Fe2+ uptake and transport in strategy I plants. Maize (Zea mays) belongs to a strategy II plant, in which mugineic acid (MA)-Fe3+ uptake is mainly carried out by Yellow Stripe 1 (YS1). However, ZmIRT1 was previously identified by our laboratory. In this study, we isolated a novel gene from maize (ZmIRT2), which is highly homologous to OsIRT2 and ZmIRT1. ZmIRT2 was expressed in roots and anther and was induced by Fe and zinc (Zn) deficiencies. ZmIRT2-GFP fusion protein localized to the plasma membrane and endoplasmic reticulum. ZmIRT2 reversed growth defects involving Zn and Fe uptake in mutant yeast. ZmIRT2 overexpression in maize led to elevated Zn and Fe levels in roots, shoots and seeds of transgenic plants. Transcript levels of ZmIRT1 were elevated in roots, while levels of YS1 were reduced in shoots of ZmIRT2 transgenic plants. Our results imply that ZmIRT2 may function solely with ZmIRT1 to mediate Fe uptake in roots. ZmIRT1, ZmIRT2 and ZmYS1 may function in a cooperative manner to maintain Zn and Fe homeostasis in ZmIRT2 overexpressing plants. Furthermore, ZmIRT2 could be used in fortification efforts to elevate Zn and Fe levels in crop plants.

A risk score model based on TGF-ß pathway-related genes predicts survival, tumor microenvironment and immunotherapy for liver hepatocellular carcinoma.

BACKGROUND: Transforming growth factor-beta (TGF-ß) signal is an important pathway involved in all stages of liver hepatocellular carcinoma (LIHC) initiation and progression. Therefore, targeting TGF- ß pathway may be a potential therapeutic strategy for LIHC. Prediction of patients' tumor cells response requires effective biomarkers. METHODS: From 54 TGF-ß-related genes, this research determined the genes showing the greatest relation to LIHC prognosis, and developed a risk score model with 8 TGF-ß-related genes. The model divided LIHC patients from different datasets and platforms into low- and high-risk groups. Multivariate Cox regression analysis confirmed that the model was an independent prognostic factor for LIHC. The differences in genetic mutation, immune cell infiltration, biological pathway, response to immunotherapy or chemotherapy, and tumor microenvironment in LIHC samples showing different risks were analyzed. RESULTS: Compared with low-risk group, in the training set and test set, high-risk group showed shorter survival, lower stromal score and higher M0 macrophages scores, regulatory T cells (Tregs), helper follicular T cells. Moreover, high-risk samples showed higher sensitivity to cisplatin, imatinib, sorafenib and salubrinal and pyrimethamine. High-risk group demonstrated a significantly higher Tumor Immune Dysfunction and Exclusion (TIDE) score, but would significantly benefit less from taking immunotherapy and was less likely to respond to immune checkpoint inhibitors. CONCLUSIONS: In general, this work provided a risk scoring model based on 8 TGF-ß pathway-related genes, which might be a new potential tool for predicting LIHC.

The role of RNA-binding protein, microRNA and alternative splicing in seed germination: a field need to be discovered.

Seed germination is the process through which a quiescent organ reactivates its metabolism culminating with the resumption cell divisions. It is usually the growth of a plant contained within a seed and results in the formation of a seedling. Post-transcriptional regulation plays an important role in gene expression. In cells, post-transcriptional regulation is mediated by many factors, such as RNA-binding proteins, microRNAs, and the spliceosome. This review provides an overview of the relationship between seed germination and post-transcriptional regulation. It addresses the relationship between seed germination and RNA-binding proteins, microRNAs and alternative splicing. This presentation of the current state of the knowledge will promote new investigations into the relevance of the interactions between seed germination and post-transcriptional regulation in plants.

Convergence and Divergence of Sugar and Cytokinin Signaling in Plant Development.

Plants adjust their growth and development through a sophisticated regulatory system integrating endogenous and exogenous cues. Many of them rely on intricate crosstalk between nutrients and hormones, an effective way of coupling nutritional and developmental information and ensuring plant survival. Sugars in their different forms such as sucrose, glucose, fructose and trehalose-6-P and the hormone family of cytokinins (CKs) are major regulators of the shoot and root functioning throughout the plant life cycle. While their individual roles have been extensively investigated, their combined effects have unexpectedly received little attention, resulting in many gaps in current knowledge. The present review provides an overview of the relationship between sugars and CKs signaling in the main developmental transition during the plant lifecycle, including seed development, germination, seedling establishment, root and shoot branching, leaf senescence, and flowering. These new insights highlight the diversity and the complexity of the crosstalk between sugars and CKs and raise several questions that will open onto further investigations of these regulation networks orchestrating plant growth and development.

Activation of ACS7 in Arabidopsis affects vascular development and demonstrates a link between ethylene synthesis and cambial activity.

Ethylene is a gaseous hormone that affects many processes of plant growth and development. During vascular development, ethylene positively regulates cambial cell division in parallel with tracheary element differentiation inhibitory factor (TDIF) peptide signaling. In this study, we identified an ethylene overproducing mutant, acs7-d, exhibiting enhanced cambial activity and reduced wall development in fiber cells. Using genetic analysis, we found that ethylene signaling is necessary for the phenotypes of enhanced cambial cell division as well as defects in stem elongation and fiber cell wall development. Further, the cambial cell proliferation phenotype of acs7-d depends on WOX4, indicating that the two parallel pathways, ethylene and TDIF signaling, converge at WOX4 in regulating cambium activity. Gene expression analysis showed that ethylene impedes fiber cell wall biosynthesis through a conserved hierarchical transcriptional regulation. These results advance our understanding of the molecular mechanisms of ethylene in regulating vascular meristem activity.

Meta-analysis of the effects of overexpression of WRKY transcription factors on plant responses to drought stress.

BACKGROUND: The tryptophan-arginine-lysine-tyrosine (WRKY) transcription factors play important roles in plants, allowing them to adapt to environmental conditions that are not normally conducive to plant growth; in particular, drought. There has been extensive research on WRKY transcription factors and the effects of their overexpression in plants on resistance to drought stress. However, due to the materials (the type and species of donor and receptor, promoters) and treatments (the type and time of stress) used, different and often confounding results have been obtained between studies. Meta-analysis is a powerful statistical tool that can be used to summarize results from numerous independent experiments on the same research topic while accounting for variability across experiments. RESULTS: We carried out a meta-analysis of 16 measured parameters that affect drought resistance in plants overexpressing WRKY transcription factors and wild-type plants. We found that only one of these parameters was significantly different between transgenic and wild-type plants under drought and control conditions at a 95% confidence interval (p = 0.000, p = 0.009, respectively). Eleven of the sixteen parameters were obviously different in WRKY transgenic plants under drought and control conditions (SV, p = 0.023, SSC, p = 0.000, SOD, p = 0.012, SFW, p = 0.000, RL, p = 0.016, Pro, p = 0.000, POD, p = 0.027, MDA, p = 0.000, H2O2, p = 0.003, EL, p = 0.000, CHC, p = 0.000, respectively), seven of the eleven obviously different parameters showed positive effect (SSC, SOD, Pro, POD, MDA, H2O2, EL), four of them revealed negative effect (SV, SFW, RL, CHC). CONCLUSION: We have found that only one of these parameters was significantly different between transgenic and wild-type plants under drought and control conditions respectively, at a 95% confidence interval. And eleven of sixteen parameters showed obviously different of WRKY-overexpressed plants under different conditions (water-stressed and normal), suggesting that WRKY transcription factors play an important role in plant responses to drought stress. These findings also provide a theoretical basis for further study of the role of WRKY transcription factors in the regulation of plant responses to environmental stress.

[A novel assay for analysis of adenovirusmediated endosome lysis of T lymphocytes].

Objective: To analyze adenovirus-mediated endosome lysis of T cells, we developed a novel approach based on pHrodo dextran (pH-sensitive fluorescent dye). Methods: After incubating Jurkat cells (T cell leukemia) with serotype 5 adenovirus (Ad5) and pHrodo dextran, we determined the optimal incubation time and concentration of pHrodo dextran. To assess viral lysis of the endosome, we monitored the ratio changes of mean fluorescence intensity in different time points by laser scanning confocal microscopy. Results: After incubating Jurkat cells with Ad5 and 80 µg/mL pHrodo dextran for 10 minutes, we observed the fluorescence intensity was significantly reduced at 30 minutes compared with that of endosomes at 0 minute. However, we found the mean fluorescence intensity was only slightly reduced by inhibiting V-ATPase with the bafilomycin A1 treatment. Conclusion: The method based on pH-sensitive dye can be used to analyze the adenovirus-mediated endosome lysis of T cells.

Comparative mapping combined with homology-based cloning of the rice genome reveals candidate genes for grain zinc and iron concentration in maize.

BACKGROUND: Grain zinc and iron concentration is a complex trait that is controlled by quantitative trait loci (QTL) and is important for maintaining body health. Despite the substantial effort that has been put into identifying QTL for grain zinc and iron concentration, the integration of independent QTL is useful for understanding the genetic foundation of traits. The number of QTL for grain zinc and iron concentration is relatively low in a single species. Therefore, combined analysis of different genomes may help overcome this challenge. RESULTS: As a continuation of our work on maize, meta-analysis of QTL for grain zinc and iron concentration in rice was performed to identify meta-QTL (MQTL). Based on MQTL in rice and maize, comparative mapping combined with homology-based cloning was performed to identify candidate genes for grain zinc and iron concentration in maize. In total, 22 MQTL in rice, 4 syntenic MQTL-related regions, and 3 MQTL-containing candidate genes in maize (ortho-mMQTL) were detected. Two maize orthologs of rice, GRMZM2G366919 and GRMZM2G178190, were characterized as natural resistance-associated macrophage protein (NRAMP) genes and considered to be candidate genes. Phylogenetic analysis of NRAMP genes among maize, rice, and Arabidopsis thaliana further demonstrated that they are likely responsible for the natural variation of maize grain zinc and iron concentration. CONCLUSIONS: Syntenic MQTL-related regions and ortho-mMQTL are prime areas for future investigation as well as for marker-assisted selection breeding programs. Furthermore, the combined method using the rice genome that was used in this study can shed light on other species and help direct future quantitative trait research. In conclusion, these results help elucidate the molecular mechanism that underlies grain zinc and iron concentration in maize.

Identification and characterization of the zinc-regulated transporters, iron-regulated transporter-like protein (ZIP) gene family in maize.

BACKGROUND: Zinc (Zn) and iron (Fe) are essential micronutrients for plant growth and development, their deficiency or excess severely impaired physiological and biochemical reactions of plants. Therefore, a tightly controlled zinc and iron uptake and homeostasis network has been evolved in plants. The Zinc-regulated transporters, Iron-regulated transporter-like Proteins (ZIP) are capable of uptaking and transporting divalent metal ion and are suggested to play critical roles in balancing metal uptake and homeostasis, though a detailed analysis of ZIP gene family in maize is still lacking. RESULTS: Nine ZIP-coding genes were identified in maize genome. It was revealed that the ZmZIP proteins share a conserved transmembrane domain and a variable region between TM-3 and TM-4. Transiently expression in onion epidermal cells revealed that all ZmZIP proteins were localized to the endoplasmic reticulum and plasma membrane. The yeast complementation analysis was performed to test the Zn or Fe transporter activity of ZmZIP proteins. Expression analysis showed that the ZmIRT1 transcripts were dramatically induced in response to Zn- and Fe-deficiency, though the expression profiles of other ZmZIP changed variously. The expression patterns of ZmZIP genes were observed in different stages of embryo and endosperm development. The accumulations of ZmIRT1 and ZmZIP6 were increased in the late developmental stages of embryo, while ZmZIP4 was up-regulated during the early development of embryo. In addition, the expression of ZmZIP5 was dramatically induced associated with middle stage development of embryo and endosperm. CONCLUSIONS: These results suggest that ZmZIP genes encode functional Zn or Fe transporters that may be responsible for the uptake, translocation, detoxification and storage of divalent metal ion in plant cells. The various expression patterns of ZmZIP genes in embryo and endosperm indicates that they may be essential for ion translocation and storage during differential stages of embryo and endosperm development. The present study provides new insights into the evolutionary relationship and putative functional divergence of the ZmZIP gene family during the growth and development of maize.

The genetic architecture of zinc and iron content in maize grains as revealed by QTL mapping and meta-analysis.

Micronutrient malnutrition, especially zinc (Zn) and iron (Fe) deficiency in diets, has aroused worldwide attention. Biofortification of food crops has been considered as a promising approach for alleviating this deficiency. Quantitative trait locus (QTL) analysis was performed to dissect the genetic mechanism of Zn and Fe content in maize grains using a total of 218 F2:3 families derived from a cross between inbred lines 178 and P53. Meta-analysis was used to integrate genetic maps and detect Meta-QTL (MQTL) across several independent QTL researches for traits related to Zn or Fe content. Five significant QTLs and 10 MQTLs were detected. Two informative genomic regions, bins 2.07 and 2.08, showed a great importance for Zn and Fe content QTLs. The correlation between Zn and Fe level in maize grains was proposed by MQTLs as 8 of the 10 involved both traits. The results of this study suggest that QTL mapping and meta-analysis is an effective approach to understand the genetic basis of Zn and Fe accumulation in maize grains.

ZmCOP1 Regulates Maize Mesocotyl Length and Plant Height through the Phytohormone Pathways.

The morphogenesis of crops is critical to their yield performance. COP1 (constitutively photomorphogenic1) is one of the core regulators in plant morphogenesis and has been deeply studied in Arabidopsis thaliana. However, the function of COP1 in maize is still unclear. Here, we found that the mesocotyl lengths of zmcop1 loss-of-function mutants were shorter than those of wild-type B73 in darkness, while the mesocotyl lengths of lines with ZmCOP1 overexpression were longer than those of wild-type B104. The plant height with zmcop1 was shorter than that of B73 in both short- and long-day photoperiods. Using transcriptome RNA sequencing technology, we identified 33 DEGs (differentially expressed genes) between B73's etiolated seedlings and those featuring zmcop1, both in darkness. The DEGs were mainly enriched in the plant phytohormone pathways. Our results provide direct evidence that ZmCOP1 functions in the elongation of etiolated seedlings in darkness and affects plant height in light. Our data can be applied in the improvement of maize plant architecture.

Research Progress on the Mechanism of Salt Tolerance in Maize: A Classic Field That Needs New Efforts.

Maize is the most important cereal crop globally. However, in recent years, maize production faced numerous challenges from environmental factors due to the changing climate. Salt stress is among the major environmental factors that negatively impact crop productivity worldwide. To cope with salt stress, plants developed various strategies, such as producing osmolytes, increasing antioxidant enzyme activity, maintaining reactive oxygen species homeostasis, and regulating ion transport. This review provides an overview of the intricate relationships between salt stress and several plant defense mechanisms, including osmolytes, antioxidant enzymes, reactive oxygen species, plant hormones, and ions (Na+, K+, Cl-), which are critical for salt tolerance in maize. It addresses the regulatory strategies and key factors involved in salt tolerance, aiming to foster a comprehensive understanding of the salt tolerance regulatory networks in maize. These new insights will also pave the way for further investigations into the significance of these regulations in elucidating how maize coordinates its defense system to resist salt stress.

Transcriptional and Metabolic Responses of Maize Shoots to Long-Term Potassium Deficiency.

Potassium is important for plant growth and crop yield. However, the effects of potassium (K+) deficiency on silage maize biomass yield and how maize shoot feedback mechanisms of K+ deficiency regulate whole plant growth remains largely unknown. Here, the study aims to explore the maize growth, transcriptional and metabolic responses of shoots to long-term potassium deficiency. Under the K+ insufficiency condition, the biomass yield of silage maize decreased. The transcriptome data showed that there were 922 and 1,107 differential expression genes in DH605 and Z58, respectively. In the two varieties, 390 differently expressed overlapping genes were similarly regulated. These genes were considered the fundamental responses to K+ deficiency in maize shoots. Many stress-induced genes are involved in transport, primary and secondary metabolism, regulation, and other processes, which are involved in K+ acquisition and homeostasis. Metabolic profiles indicated that most amino acids, phenolic acids, organic acids, and alkaloids were accumulated in shoots under K+ deficiency conditions and part of the sugars and sugar alcohols also increased. It revealed that putrescine and putrescine derivatives were specifically accumulated under the K+ deficiency condition, which may play a role in the feedback regulation of shoot growth. These results confirmed the importance of K+ on silage maize production and provided a deeper insight into the responses to K+ deficiency in maize shoots.

Identification of quantitative trait loci for related traits of stalk lodging resistance using genome-wide association studies in maize (Zea mays L.).

BACKGROUND: Stalk lodging is one of the main factors affecting maize (Zea mays L.) yield and limiting mechanized harvesting. Developing maize varieties with high stalk lodging resistance requires exploring the genetic basis of lodging resistance-associated agronomic traits. Stalk strength is an important indicator to evaluate maize lodging and can be evaluated by measuring stalk rind penetrometer resistance (RPR) and stalk buckling strength (SBS). Along with morphological traits of the stalk for the third internodes length (TIL), fourth internode length (FIL), third internode diameter (TID), and the fourth internode diameter (FID) traits are associated with stalk lodging resistance. RESULTS: In this study, a natural population containing 248 diverse maize inbred lines genotyped with 83,057 single nucleotide polymorphism (SNP) markers was used for genome-wide association study (GWAS) for six stalk lodging resistance-related traits. The heritability of all traits ranged from 0.59 to 0.72 in the association mapping panel. A total of 85 significant SNPs were identified for the association mapping panel using best linear unbiased prediction (BLUP) values of all traits. Additionally, five candidate genes were associated with stalk strength traits, which were either directly or indirectly associated with cell wall components. CONCLUSIONS: These findings contribute to our understanding of the genetic basis of maize stalk lodging and provide valuable theoretical guidance for lodging resistance in maize breeding in the future.

MiR-495 Inhibits Cisplatin Resistance and Angiogenesis in Esophageal Cancer by Targeting ATP7A.

Background: Cancer resistance to chemotherapy is closely associated with changes in transporter systems. In this study, we investigated the possible regulation of 1 copper ion transporter (ATP7A; ATPase copper transporting alpha) by microRNA miR-495 and its implications in cisplatin resistance and angiogenesis in esophageal cancer. Methods: MiR-495 and ATP7A mRNA expression in clinical tissue samples and 2 cancer cell lines (Eca-109 and TE1) were detected by quantitative real-time polymerase chain reaction. The levels of miR-495 and ATP7A expression in Eca-109 and TE1 cells were increased by transfection with miR-495 mimics and ATP7A-overexpression vectors. Cell proliferation, apoptosis, and angiogenesis were assessed by CCK-8, flow cytometry, and tube formation assays, respectively. The levels of TNF-α and VEGF in cell culture supernatants were detected by enzyme linked immunosorbent assay, and in situ expression of NLRP3 was measured by immunofluorescence. The binding of miR-495 to ATP7A sequences was verified by dual luciferase reporter assays. Results:ATP7A expression was significantly increased, while miR-495 expression was decreased in the cancer tissues of esophageal cancer patients. MiR-495 mimics decreased the proliferation and promoted the apoptosis of cisplatin-resistant Eca-109 and TE1 cells. Furthermore, tube formation by human umbilical vein endothelial cells, TNF-α and VEGF secretion, and the levels of MRP1, ABCG1, ABCA1, and NLRP3 expression in cisplatin-resistant Eca-109 and TE1 cells were all reduced by miR-495 mimics. MiR-495 was shown to directly bind to ATP7A gene sequences to repress ATP7A expression in Eca-109 and TE1 cells. ATP7A overexpression substantially abrogated the changes in proliferation, apoptosis, angiogenesis, and above-mentioned gene expression in cisplatin-resistant Eca-109 and TE1 cells. Conclusions: MiR-495 suppressed cisplatin resistance and angiogenesis in esophageal cancer cells by targeting ATP7A gene expression.

Association mapping for general combining ability with yield, plant height and ear height using F1 population in maize.

General combining ability (GCA) is an important index for inbred lines breeding of maize. To identify the genetic loci of GCA and associated agronomic traits, an association analysis with 195 SSRs was made in phenotypic traits of 240 F1 derived from 120 elite inbred lines containing current breeding resources of maize crossed with 2 testers (Zheng58 and Chang7-2) in two places in 2018. All of the 20 association loci detected for grain yield (GY), plant height (PH), ear height (EH) and GCA for the three traits in two places could explain a phenotypic variation range of 7.31%-9.29%. Among the 20 association loci, 9 (7.31%-9.04%) were associated with GY, 4 (7.22%-8.91%) were related to GCA of GY, 1 (7.56%) was associated with PH, and 3 (7.53%-8.96%) were related to EH. In addition, 3 loci (9.14%-9.29%) were associated with GCA of PH whereas no locus was identified for GCA of EH. In the comparison of the association loci detected in Baoding and Handan, interestingly, one locus (7.69% and 8.11%) was identified in both environments and one locus (7.52% and 7.82%) was identified for yield and GCA of yield. Therefore, the identification of GY-, PH-, EH- and GCA-related association loci could not only provide references for high yield breeding of maize, but also help us comprehend the relationships among GY, agricultural traits and GCA.

Sugar Signaling and Post-transcriptional Regulation in Plants: An Overlooked or an Emerging Topic?

Plants are autotrophic organisms that self-produce sugars through photosynthesis. These sugars serve as an energy source, carbon skeletons, and signaling entities throughout plants' life. Post-transcriptional regulation of gene expression plays an important role in various sugar-related processes. In cells, it is regulated by many factors, such as RNA-binding proteins (RBPs), microRNAs, the spliceosome, etc. To date, most of the investigations into sugar-related gene expression have been focused on the transcriptional level in plants, while only a few studies have been conducted on post-transcriptional mechanisms. The present review provides an overview of the relationships between sugar and post-transcriptional regulation in plants. It addresses the relationships between sugar signaling and RBPs, microRNAs, and mRNA stability. These new items insights will help to reach a comprehensive understanding of the diversity of sugar signaling regulatory networks, and open onto new investigations into the relevance of these regulations for plant growth and development.

Identification and characterization of maize ACD6-like gene reveal ZmACD6 as the maize orthologue conferring resistance to Ustilago maydis.

Enhancing broad-spectrum resistance is a major goal of crop breeding. However, broad-spectrum resistance has not been thoroughly investigated, and its underlying molecular mechanisms remain elusive. In the model plant Arabidopsis (Arabidopsis thaliana), ACCELERATED CELL DEATH6 (ACD6) is a key component of broad-spectrum resistance that acts in a positive feedback loop with salicylic acid (SA) to regulate multiple pattern recognition receptors. However, the role of ACD6 in disease resistance in crop plants is unclear. Here, we show that the transcript of ANK23, one of the 15 ACD6-like genes in maize (Zea mays), is induced by SA and by infection with the pathogenic fungus Ustilago maydis. Heterologous expression of ANK23 restored disease resistance in the Arabidopsis mutant acd6-2. We show that ANK23 is a maize ortholog of ACD6 and therefore rename ANK23 as ZmACD6. Furthermore, using CRISPR/Cas9, we generated ZmACD6 knockout maize plants, which are more susceptible to U. maydis than wild-type plants. We also identified a maize line (SC-9) with relatively high ZmACD6 expression levels from a diverse natural maize population. SC-9 has increased disease resistance to U. maydis and defense activation, suggesting a practical approach to cultivate elite varieties with enhanced disease resistance.