The Mechanism of Exogenous Salicylic Acid and 6-Benzylaminopurine Regulating the Elongation of Maize Mesocotyl.

The elongation of the mesocotyl plays an important role in the emergence of maize deep-sowing seeds. This study was designed to explore the function of exogenous salicylic acid (SA) and 6-benzylaminopurine (6-BA) in the growth of the maize mesocotyl and to examine its regulatory network. The results showed that the addition of 0.25 mmol/L exogenous SA promoted the elongation of maize mesocotyls under both 3 cm and 15 cm deep-sowing conditions. Conversely, the addition of 10 mg/L exogenous 6-BA inhibited the elongation of maize mesocotyls. Interestingly, the combined treatment of exogenous SA-6-BA also inhibited the elongation of maize mesocotyls. The longitudinal elongation of mesocotyl cells was the main reason affecting the elongation of maize mesocotyls. Transcriptome analysis showed that exogenous SA and 6-BA may interact in the hormone signaling regulatory network of mesocotyl elongation. The differential expression of genes related to auxin (IAA), jasmonic acid (JA), brassinosteroid (BR), cytokinin (CTK) and SA signaling pathways may be related to the regulation of exogenous SA and 6-BA on the growth of mesocotyls. In addition, five candidate genes that may regulate the length of mesocotyls were screened by Weighted Gene Co-Expression Network Analysis (WGCNA). These genes may be involved in the growth of maize mesocotyls through auxin-activated signaling pathways, transmembrane transport, methylation and redox processes. The results enhance our understanding of the plant hormone regulation of mesocotyl growth, which will help to further explore and identify the key genes affecting mesocotyl growth in plant hormone signaling regulatory networks.

WGCNA analysis of the effect of exogenous BR on leaf angle of maize mutant lpa1.

Leaf angle, as one of the important agronomic traits of maize, can directly affect the planting density of maize, thereby affecting its yield. Here we used the ZmLPA1 gene mutant lpa1 to study maize leaf angle and found that the lpa1 leaf angle changed significantly under exogenous brassinosteroid (BR) treatment compared with WT (inbred line B73). Transcriptome sequencing of WT and lpa1 treated with different concentrations of exogenous BR showed that the differentially expressed genes were upregulated with auxin, cytokinin and brassinosteroid; Genes associated with abscisic acid are down-regulated. The differentially expressed genes in WT and lpa1 by weighted gene co-expression network analysis (WGCNA) yielded two gene modules associated with maize leaf angle change under exogenous BR treatment. The results provide a new theory for the regulation of maize leaf angle by lpa1 and exogenous BR.

Phenotypic Investigation and RNA-seq of KN1 Involved in Leaf Angle Formation in Maize (Zea mays L.).

Leaf angle (LA) is one of the core agronomic traits of maize, which controls maize yield by affecting planting density. Previous studies have shown that the KN1 gene is closely related to the formation of maize LA, but its specific mechanism has not been fully studied. In this study, phenotype investigation and transcriptomic sequencing were combined to explore the mechanism of LA changes in wild type maize B73 and mutant kn1 under exogenous auxin (IAA) and abscisic acid (ABA) treatment. The results showed that the effect of exogenous phytohormones had a greater impact on the LA of kn1 compared to B73. Transcriptome sequencing showed that genes involved in IAA, gibberellins (GAs) and brassinosteroids (BRs) showed different differential expression patterns in kn1 and B73. This study provides new insights into the mechanism of KN1 involved in the formation of maize LA, and provides a theoretical basis for breeding maize varieties with suitable LA.

Corrigendum: Morphological characterization and transcriptome analysis of leaf angle mutant bhlh112 in maize [Zea mays L.].

[This corrects the article DOI: 10.3389/fpls.2022.995815.].

Morphological characterization and transcriptome analysis of leaf angle mutant bhlh112 in maize [Zea mays L.].

Leaf angle is an important agronomic trait in maize [Zea mays L.]. The compact plant phenotype, with a smaller leaf angle, is suited for high-density planting and thus for increasing crop yields. Here, we studied the ethyl methane sulfonate (EMS)-induced mutant bhlh112. Leaf angle and plant height were significantly decreased in bhlh112 compared to the wild-type plants. After treatment of seedlings with exogenous IAA and ABA respectively, under the optimal concentration of exogenous hormones, the variation of leaf angle of the mutant was more obvious than that of the wild-type, which indicated that the mutant was more sensitive to exogenous hormones. Transcriptome analysis showed that the ZmbHLH112 gene was related to the biosynthesis of auxin and brassinosteroids, and involved in the activation of genes related to the auxin and brassinosteroid signal pathways as well as cell elongation. Among the GO enrichment terms, we found many differentially expressed genes (DEGs) enriched in the cell membrane and ribosomal biosynthesis, hormone biosynthesis and signaling pathways, and flavonoid biosynthesis, which could influence cell growth and the level of endogenous hormones affecting leaf angle. Therefore, ZmbHLH112 might regulate leaf angle development through the auxin signaling and the brassinosteroid biosynthesis pathways. 12 genes related to the development of leaf were screened by WGCNA; In GO enrichment and KEGG pathways, the genes were mainly enriched in rRNA binding, ribosome biogenesis, Structural constituent of ribosome; Arabidopsis ribosome RNA methyltransferase CMAL is involved in plant development, likely by modulating auxin derived signaling pathways; The free 60s ribosomes and polysomes in the functional defective mutant rice minute-like1 (rml1) were significantly reduced, resulting in plant phenotypic diminution, narrow leaves, and growth retardation; Hence, ribosomal subunits may play an important role in leaf development. These results provide a foundation for further elucidation of the molecular mechanism of the regulation of leaf angle in maize.

Mutant lpa1 Analysis of ZmLPA1 Gene Regulates Maize Leaf-Angle Development through the Auxin Pathway.

Maize plant type is one of the main factors determining maize yield, and leaf angle is an important aspect of plant type. The rice Loose Plant Architecture1 (LPA1) gene and Arabidopsis AtIDD15/SHOOT GRAVITROPISM5 (SGR5) gene are related to their leaf angle. However, the homologous ZmLPA1 in maize has not been studied. In this study, the changing of leaf angle, as well as gene expression in leaves in maize mutant lpa1 and wild-type 'B73' under different IAA concentrations were investigated. The regulation effect of IAA on the leaf angle of lpa1 was significantly stronger than that of the wild type. Transcriptome analysis showed that different exogenous IAA treatments had a common enrichment pathway-the indole alkaloid biosynthesis pathway-and among the differentially expressed genes, four genes-AUX1, AUX/IAA, ARF and SAUR-were significantly upregulated. This study revealed the regulation mechanism of ZmLPA1 gene on maize leaf angle and provided a promising gene resource for maize breeding.

Transcriptome expression profiles reveal response mechanisms to drought and drought-stress mitigation mechanisms by exogenous glycine betaine in maize.

Drought stress is one of the major abiotic stresses that limit growth, development and yield of maize crops. To better understand the responses of maize inbred lines with different levels of drought resistance and the molecular mechanism of exogenous glycine betaine (GB) in alleviating drought stress, the responses of two maize inbred lines to drought stress and to the stress-mitigating effects of exogenous GB were investigated. Seedling morphology, physiological and biochemical indexes, root cell morphology and root transcriptome expression profiles were compared between a drought-tolerant inbred line Chang 7-2 and drought-sensitive inbred line TS141. Plants of both lines were subjected to treatments of drought stress alone and drought stress with application of exogenous GB. The results showed that with the increase of drought treatment time, the growth and development of TS141 were inhibited, while those of Chang 7-2 were not significantly different from those of the control (no drought stress and GB). Compared with the corresponding data of the drought-stress group, every index measured from the two inbred lines indicated mitigating effects from exogenous GB, and TS141 produced stronger mitigating responses due to the GB. Transcriptome analysis showed that 562 differentially expressed genes (DEGs) were up-regulated and 824 DEGs were down-regulated in both inbred lines under drought stress. Due to the exogenous GB, 1061 DEGs were up-regulated and 424 DEGs were down-regulated in both lines. In addition, quantitative real-time polymerase chain reaction (qRT-PCR) was used to verify 10 DEGs screened from the different treatments. These results showed that the expression of 9 DEGs were basically consistent with their respective transcriptome expression profiles. The results of this study provide models of potential mechanisms of drought tolerance in maize as well as potential mechanisms of how exogenous GB may regulate drought tolerance.

Study on ZmRPN10 Regulating Leaf Angle in Maize by RNA-Seq.

Ubiquitin/proteasome-mediated proteolysis (UPP) plays a crucial role in almost all aspects of plant growth and development, proteasome subunit RPN10 mediates ubiquitination substrate recognition in the UPP process. The recognition pathway of ubiquitinated UPP substrate is different in different species, which indicates that the mechanism and function of RPN10 are different in different species. However, the homologous ZmRPN10 in maize has not been studied. In this study, the changing of leaf angle and gene expression in leaves in maize wild-type B73 and mutant rpn10 under exogenous brassinosteroids (BRs) were investigated. The regulation effect of BR on the leaf angle of rpn10 was significantly stronger than that of B73. Transcriptome analysis showed that among the differentially expressed genes, CRE1, A-ARR and SnRK2 were significantly up-regulated, and PP2C, BRI1 AUX/IAA, JAZ and MYC2 were significantly down-regulated. This study revealed the regulation mechanism of ZmRPN10 on maize leaf angle and provided a promising gene resource for maize breeding.

Network Analysis of Different Exogenous Hormones on the Regulation of Deep Sowing Tolerance in Maize Seedlings.

The planting method of deep sowing can make the seeds make full use of water in deep soil, which is considered to be an effective way to respond to drought stress. However, deep sowing will affect the growth and development of maize (Zea mays L.) at seedling stage. To better understand the response of maize to deep sowing stress and the mechanism of exogenous hormones [Gibberellin (GA3), Brassinolide (BR), Strigolactone (SL)] alleviates the damaging effects of deep-sowing stress, the physiological and transcriptome expression profiles of seedlings of deep sowing sensitive inbred line Zi330 and the deep-tolerant inbred line Qi319 were compared under deep sowing stress and the conditions of exogenous hormones alleviates stress. The results showed that mesocotyl elongated significantly after both deep sowing stress and application of exogenous hormones, and its elongation was mainly through elongation and expansion of cell volume. Hormone assays revealed no significant changes in zeatin (ZT) content of the mesocotyl after deep sowing and exogenous hormone application. The endogenous GA3 and auxin (IAA) contents in the mesocotyl of the two inbred lines increased significantly after the addition of exogenous GA3, BR, and SL under deep sowing stress compared to deep sowing stress, while BR and SL decreased significantly. Transcriptome analysis showed that the deep seeding stress was alleviated by GA3, BR, and SLs, the differentially expressed genes (DEGs) mainly included cellulose synthase, expansin and glucanase, oxidase, lignin biosynthesis genes and so on. We also found that protein phosphatase 2C and GA receptor GID1 enhanced the ability of resist deep seeding stress in maize by participating in the abscisic acid (ABA) and the GA signaling pathway, respectively. In addition, we identified two gene modules that were significantly related to mesocotyl elongation, and identified some hub genes that were significantly related to mesocotyl elongation by WGCNA analysis. These genes were mainly involved in transcription regulation, hydrolase activity, protein binding and plasma membrane. Our results from this study may provide theoretical basis for determining the maize deep seeding tolerance and the mechanism by which exogenous hormones regulates deep seeding tolerance.

Transcriptional analysis reveals sodium nitroprusside affects alfalfa in response to PEG-induced osmotic stress at germination stage.

Drought is one of the most common environmental factors that affect alfalfa germination and development. Nitric oxide (NO) could mediate stress tolerance in plants. The goal of this study was to determine exogenous NO donor-mediated drought adaption molecular mechanisms during the alfalfa germination stage. In this study, physiological and transcriptome analyses were performed on 7 days of the growth period seedlings by sodium nitroprusside (SNP) and polyethylene glycol (PEG) treatment. The results showed that SNP supplementation alleviated malondialdehyde accumulation, increased levels of proline and soluble sugars, and enhanced antioxidant enzyme activity under osmotic stress conditions. RNA-Seq experiments identified 5828 genes exhibiting differential expression in seedlings treated with PEG, SNP, or SNP+PEG relative to seedlings treated with distilled water. Of these DEGs, 3235 were upregulated, and 2593 were downregulated relative to the controls. Fifteen DEGs were amplified by qRT-PCR to verify the changes in expression determined by RNA-Seq, revealing that PIF3, glnA, PLCG1, and RP-S11e exhibited enhanced expression under the SNP+PEG treatment. SNP was found to modulate redox homeostasis-related genes such as GSTs, SOD2, GPX, and RBOH, and triggered calcium signaling transduction. It also induced some key genes relating to the abscisic acid, ethylene, and auxin signaling transduction in response to PEG stress. Conversely, genes associated with secondary metabolite biosynthesis and the metabolism of starch and sucrose during osmotic stress were downregulated by SNP. These results provide new insights into SNP-mediated drought adaption mechanisms at transcriptome-wide in alfalfa and reveal key drought tolerance pathways in this species.

Endogenous NO-mediated transcripts involved in photosynthesis and carbohydrate metabolism in alfalfa (Medicago sativa L.) seedlings under drought stress.

Alfalfa (Medicago sativa L.) is an important perennial legume and used as a forage crop worldwide, and has extensive resistance to various abiotic stresses. Nitric oxide (NO) plays a critical role in response to external and internal cues to regulate plant growth and development. However, endogenous NO-mediated molecular mechanisms of drought tolerance in alfalfa is poorly understood. To get a deeper insight into the regulate pathway of NO, RNA-Seq was used to profile transcriptome changes of alfalfa seedlings, which were treated with NO scavenger under normal and drought conditions. A total of 1,025 and 3,461 differently-expressed genes (FDR < 0.0001; fold change ≥ 2) were observed while NO absence under normal and drought conditions, respectively. Based on GO enrich and KEGG pathway analysis, we found NO absence induced photosynthesis, carbon fixation in photosynthetic organisms and primary metabolism were significantly up-enriched. Most oxidoreductase, dehydrogenase, reductase and transferase genes were down-regulated in the above processes. Moreover, NO absence restrained chlorophyll biosynthesis and decreased different sugar content. Therefore, this work provides insights into the mechanism that NO-mediated enhanced photosynthesis and carbohydrate metabolism in alfalfa under drought stress.

Transgenic potato plants expressing cry3A gene confer resistance to Colorado potato beetle.

The Colorado potato beetle (Leptinotarsa decemlineata Say, CPB) is a fatal pest, which is a quarantine pest in China. The CPB has now invaded the Xinjiang Uygur Autonomous Region and is constantly spreading eastward in China. In this study, we developed transgenic potato plants expressing cry3A gene. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis indicated that the cry3A gene expressed in leaves, stems and roots of the transgenic plants under the control of CaMV 35S promoter, while they expressed only in leaves and stems under the control of potato leaf and stem-specific promoter ST-LS1. The mortality of the larvae was higher (28% and 36%) on the transgenic plant line 35S1 on the 3rd and 4th days, and on ST3 (48%) on the 5th day after inoculation with instar larvae. Insect biomass accumulation on the foliage of the transgenic plant lines 35S1, 35S2 and ST3 was significantly lower (0.42%, 0.43% and 0.42%). Foliage consumption was lowest on transgenic lines 35S8 and ST2 among all plant foliage (7.47 mg/larvae/day and 12.46 mg/larvae/day). The different transgenic plant foliages had varied inhibition to larval growth. The survivors on the transgenic lines obviously were smaller than their original size and extremely weak. The transgenic potato plants with CPB resistance could be used to develop germplasms or varieties for controlling CPB damage and halting its spread in China.