Plant Hormone Biosynthesis Research Articles

Combined BSA-Seq Based Mapping and RNA-Seq Profiling Reveal Candidate Genes Associated with Plant Architecture in Brassica napus.

Plant architecture involves important agronomic traits affecting crop yield, resistance to lodging, and fitness for mechanical harvesting in Brassica napus. Breeding high-yield varieties with plant architecture suitable for mechanical harvesting is the main goal of rapeseed breeders. Here, we report an accession of B. napus (4942C-5), which has a dwarf and compact plant architecture in contrast to cultivated varieties. A BC8 population was constructed by crossing a normal plant architecture line, 8008, with the recurrent parent 4942C-5. To investigate the molecular mechanisms underlying plant architecture, we performed phytohormone profiling, bulk segregant analysis sequencing (BSA-Seq), and RNA sequencing (RNA-Seq) in BC8 plants with contrasting plant architecture. Genetic analysis indicated the plant architecture traits of 4942C-5 were recessive traits controlled by multiple genes. The content of auxin (IAA), gibberellin (GA), and abscisic acid (ABA) differed significantly between plants with contrasting plant architecture in the BC8 population. Based on BSA-Seq analysis, we identified five candidate intervals on chromosome A01, namely those of 0 to 6.33 Mb, 6.45 to 6.48 Mb, 6.51 to 6.53 Mb, 6.77 to 6.79 Mb, and 7 to 7.01 Mb regions. The RNA-Seq analysis revealed a total of 4378 differentially expressed genes (DEGs), of which 2801 were up-regulated and 1577 were down-regulated. There, further analysis showed that genes involved in plant hormone biosynthesis and signal transduction, cell structure, and the phenylpropanoid pathway might play a pivotal role in the morphogenesis of plant architecture. Association analysis of BSA-Seq and RNA-Seq suggested that seven DEGs involved in plant hormone signal transduction and a WUSCHEL-related homeobox (WOX) gene (BnaA01g01910D) might be candidate genes responsible for the dwarf and compact phenotype in 4942C-5. These findings provide a foundation for elucidating the mechanisms underlying rapeseed plant architecture and should contribute to breed new varieties suitable for mechanization.

Transcriptomics-Based Identification of Genes Related to Tapetum Degradation and Microspore Development in Lily.

Lily is a popular and economically ornamental crop around the world. However, its high production of pollen grains causes serious problems to consumers, including allergies and staining of clothes. During anther development, the tapetum is a crucial step for pollen formation and microspore release. Therefore, it is important to understand the mechanism of tapetum degradation and microspore development in lily where free pollen contamination occurs. Here, we used the cut lily cultivar ‘Siberia’ to characterize the process of tapetum degradation through the use of cytology and transcriptomic methods. The cytological observation indicated that, as the lily buds developed from 4 cm (Lo 4 cm) to 8 cm (Lo 8 cm), the tapetum completed the degradation process and the microspores matured. Furthermore, by comparing the transcriptome profiling among three developmental stages (Lo 4 cm, Lo 6 cm and Lo 8 cm), we identified 27 differentially expressed genes. These 27 genes were classed into 4 groups by function, namely, cell division and expansion, cell-wall morphogenesis, transcription factors, LRR-RLK (leucine-rich repeat receptor-like kinases), plant hormone biosynthesis and transduction. Quantitative real-time PCR was performed as validation of the transcriptome data. These selected genes are candidate genes for the tapetum degradation and microspore development of lily and our work provides a theoretical basis for breeding new lily cultivars without pollen.

NAC Transcription Factor Family Regulation of Fruit Ripening and Quality: A Review.

The NAC transcription factor (TF) family is one of the largest plant-specific TF families and its members are involved in the regulation of many vital biological processes during plant growth and development. Recent studies have found that NAC TFs play important roles during the ripening of fleshy fruits and the development of quality attributes. This review focuses on the advances in our understanding of the function of NAC TFs in different fruits and their involvement in the biosynthesis and signal transduction of plant hormones, fruit textural changes, color transformation, accumulation of flavor compounds, seed development and fruit senescence. We discuss the theoretical basis and potential regulatory models for NAC TFs action and provide a comprehensive view of their multiple roles in modulating different aspects of fruit ripening and quality.

New insights into the role of cyanide in the promotion of seed germination in tomato

BackgroundCyanide is a natural metabolite that exists widely in plants, and it is speculated to be involved in the regulation of various growth and development processes of plants in addition to being regarded as toxic waste. Previous studies have shown that exogenous cyanide treatment helps to improve seed germination, but the mechanism is still unclear. In this study, tomato (Solanum lycopersicum cv. Alisa Craig) was used as the material, and the effects of cyanide pretreatment at different concentrations on tomato seed germination were investigated.ResultsThe results showed that exogenous application of a lower concentration of cyanide (10 μmol/L KCN) for 12 h strongly increased the tomato seed germination rate. RNA-Seq showed that compared with the control, a total of 15,418 differentially expressed genes (P<0.05) were obtained after pretreatment with KCN for 12 h, and in the next 12 h, a total of 13,425 differentially expressed genes (P<0.05) were regulated. GO and KEGG analyses demonstrated that exogenous KCN pretreatment was involved in regulating the expression (mainly downregulation) of seed storage proteins, thereby accelerating the degradation of stored proteins for seed germination. In addition, KCN pretreatment was also involved in stimulating glycolysis, the TCA cycle and oxidative phosphorylation. Notably, it is shown that KCN acted on the regulation of plant hormone biosynthesis and perception, i.e., down-regulated the gene expression of ABA biosynthesis and signal transduction, but up-regulated the expression of genes related to GA biosynthesis and response. Consistent with this, plant hormone measurements confirmed that the levels of ABA were reduced, but GA levels were induced after pretreatment with KCN.ConclusionThese findings provide new insights into the regulation of seed germination by cyanide, that is cyanide-mediated seed germination occurs in a time- and dose-dependent manner, and is related to the mobilization of energy metabolism and the regulation of some plant hormone signals.

Integrated microbiology and metabolomics analysis reveal responses of soil microorganisms and metabolic functions to phosphorus fertilizer on semiarid farm

Localized fertilization of phosphorus has potential benefits in achieving higher crop productivity and nutrient use efficiency, but the underlying biological mechanisms of interactions between soil microorganisms and related metabolic cycle remain largely to be recognized. Here, we combined microbiology with non-target metabolomics to explore how P fertilizer levels and fertilization patterns affect wheat soil microbial communities and metabolic functions based on high-throughput sequencing and UPLC-MS/MS platforms. The results showed P fertilizer decreased the diversity of bacterial 16S rRNA genes and fungal ITS genes, and it did significantly change both soil bacterial and fungal overall community structures and compositions. The P levels and patterns also interfered with complexity of soil bacterial and fungal symbiosis networks. Moreover, metabolomics analysis showed that P fertilizer significantly changed soil metabolite spectrum, and the differential metabolites were significantly enriched to 7 main metabolic pathways, such as arginine and proline metabolism, biosynthesis of plant hormones, amino acids, plant secondary metabolites, and alkaloids derived from ornithine. Additionally, microbes also were closely related to the accumulation of metabolites through correlation analysis. Our results indicated that localized appropriate phosphorus fertilizer plays an important role in regulating soil microbial metabolism, and their interactions in soil providing valuable information for understanding how the changed phosphorus management practices affect the complex biological processes and the adaption capacity of plants to environments.

Diverse reactions catalyzed by cytochrome P450 and biosynthesis of steroid hormone.

Steroid hormones modulate numerous physiological processes in various higher organisms. Research on the physiology, biosynthesis, and metabolic degradation of steroid hormones is crucial for developing drugs, agrochemicals, and anthelmintics. Most steroid hormone biosynthetic pathways, excluding those in insects, have been elucidated, and the roles of several cytochrome P450s (CYPs, P450s), heme (iron protoporphyrin IX)-containing monooxygenases, have been identified. Specifically, P450s of the animal steroid hormone biosynthetic pathways and their three dimensional structures and reaction mechanisms have been extensively studied; however, the mechanisms of several uncommon P450 reactions involved in animal steroid hormone biosynthesis and structures and reaction mechanisms of various P450s involved in plant and insect steroid hormone biosynthesis remain unclear. Recently, we determined the crystal structure of P450 responsible for the first and rate-determining step in brassinosteroids biosynthesis and clarified the regio- and stereo-selectivity in the hydroxylation reaction mechanism. In this review, we have outlined the general catalytic cycle, reaction mechanism, and structure of P450s. Additionally, we have described the recent advances in research on the reaction mechanisms of steroid hormone biosynthesis-related P450s, some of which catalyze unusual P450 reactions including C–C bond cleavage reactions by utilizing either a heme–peroxo anion species or compound I as an active oxidizing species. This review article is an extended version of the Japanese article, Structure and mechanism of cytochrome P450s involved in steroid hormone biosynthesis, published in SEIBUTSU BUTSURI Vol. 61, p. 189–191 (2021).

Insight Into the Multiple Branches Traits of a Mutant in Larix olgensis by Morphological, Cytological, and Transcriptional Analyses.

Larix olgensis is a tall deciduous tree species that has many applications in the wood fiber industry. Bud mutations are somatic mutations in plants and are considered an ideal material to identify and describe the molecular mechanism of plant mutation. However, the molecular regulatory mechanisms of bud mutations in L. olgensis remain unknown. In this study, dwarfed (or stunted), short-leaved, and multi-branched mutants of L. olgensis were found and utilized to identify crucial genes and regulatory networks controlling the multiple branch structure of L. olgensis. The physiological data showed that the branch number, bud number, fresh and dry weight, tracheid length, tracheid length-width ratio, inner tracheid diameter, and epidermal cell area of mutant plants were higher than that of wild-type plants. Hormone concentration measurements found that auxin, gibberellin, and abscisic acid in the mutant leaves were higher than that in wild-type plants. Moreover, the transcriptome sequencing of all samples using the Illumina Hiseq sequencing platform. Transcriptome analysis identified, respectively, 632, 157, and 199 differentially expressed genes (DEGs) in buds, leaves, and stems between mutant plants and wild type. DEGs were found to be involved in cell division and differentiation, shoot apical meristem activity, plant hormone biosynthesis, and sugar metabolism. Furthermore, bZIP, WRKY, and AP2/ERF family transcription factors play a role in bud formation. This study provides new insights into the molecular mechanisms of L. olgensis bud and branch formation and establishes a fundamental understanding of the breeding of new varieties in L. olgensis.

Effects of gibberellin priming on seedling emergence and transcripts involved in mesocotyl elongation in rice under deep direct-seeding conditions.

Mesocotyl elongation is a key trait influencing seedling emergence and establishment in direct-seeding rice cultivation. The phytohormone gibberellin (GA) has positive effects on mesocotyl elongation in rice. However, the physiological and molecular basis underlying the regulation of mesocotyl elongation mediated by GA priming under deep-sowing conditions remains largely unclear. In the present study, we performed a physiological and comprehensive transcriptomic analysis of the function of GA priming in mesocotyl elongation and seedling emergence using a direct-seeding japonica rice cultivar ZH10 at a 5-cm sowing depth. Physiological experiments indicated that GA priming significantly improved rice seedling emergence by increasing the activity of starch-metabolizing enzymes and compatible solute content to supply the energy essential for subsequent development. Transcriptomic analysis revealed 7074 differentially expressed genes (false discovery rate of <0.05, |log2(fold change)| of ≥1) after GA priming. Furthermore, gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) enrichment analyses revealed that genes associated with transcriptional regulation, plant hormone biosynthesis or signaling, and starch and sucrose metabolism were critical for GA-mediated promotion of rice mesocotyl elongation. Further analyses showed that the expression of the transcription factor (TF) genes (v-myb avian myeloblastosis viral oncogene homolog (MYB) alternative splicing 1 (MYBAS1), phytochrome-interacting factors 1 (PIF1), Oryza sativa teosinte branched 1/cycloidea/proliferating cell factor 5 (OsTCP5), slender 1 (SLN1), and mini zinc finger 1 (MIF1)), plant hormone biosynthesis or signaling genes (brassinazole-resistant 1 (BZR1), ent-kaurenoic acid oxidase-like (KAO), GRETCHEN HAGEN 3.2 (GH3.2), and small auxin up RNA 36 (SAUR36)), and starch and sucrose metabolism genes (α-amylases (AMY2A and AMY1.4)) was highly correlated with the mesocotyl elongation and deep-sowing tolerance response. These results enhance our understanding of how nutrient metabolism-related substances and genes regulate rice mesocotyl elongation. This may facilitate future studies on related genes and the development of novel rice varieties tolerant to deep sowing.

Integrated Transcriptome Analysis and Single-Base Resolution Methylomes of Watermelon (Citrullus lanatus) Reveal Epigenome Modifications in Response to Osmotic Stress.

DNA methylation plays an important role against adverse environment by reshaping transcriptional profile in plants. To better understand the molecular mechanisms of watermelon response to osmotic stress, the suspension cultured watermelon cells were treated with 100mM mannitol, and then a methylated cytosines map was generated by whole genome bisulfite sequencing (WGBS). Combined with transcriptome sequencing, the effects of osmotic stress on differentially methylated expressed genes (DMEGs) were assessed. It was found that genes related to plant hormone synthesis, signal transduction, osmoregulatory substance-related and reactive oxygen species scavenging-related enzyme could rapidly respond to osmotic stress. The overall methylation level of watermelon decreased after osmotic stress treatment, and demethylation occurred in CG, CHG, and CHH contexts. Moreover, differentially methylated expressed genes (DMEGs) were significantly enriched in RNA transport, starch and sucrose metabolism, plant hormone signal transduction and biosynthesis of secondary metabolites, especially in biosynthesis of osmolytes synthase genes. Interestingly, demethylation of a key enzyme gene Cla014489 in biosynthesis of inositol upregulated its expression and promoted accumulation of inositol, which could alleviate the inhibition of cell growth caused by osmotic stress. Meanwhile, a recombinant plasmid pET28a-Cla014489 was constructed and transferred into Escherichia coli BL21 for prokaryotic expression and the expression of ClMIPS protein could improve the tolerance of E. coli to osmotic stress. The effect of methylation level on the expression properties of inositol and its related genes was further confirmed by application of DNA methylation inhibitor 5-azacytidine. These results provide a preliminary insight into the altered methylation levels of watermelon cells in response to osmotic stress and suggest a new mechanism that how watermelon cells adapt to osmotic stress.

Rambutan genome revealed gene networks for spine formation and aril development

Rambutan is a popular tropical fruit known for its exotic appearance, has long flexible spines on shells, extraordinary aril growth, desirable nutrition, and a favorable taste. The genome of an elite rambutan cultivar Baoyan 7 was assembled into 328 Mb in 16 pseudo-chromosomes. Comparative genomics analysis between rambutan and lychee revealed that rambutan chromosomes 8 and 12 are collinear with lychee chromosome 1, which resulted in a chromosome fission event in rambutan (n = 16) or a fusion event in lychee (n = 15) after their divergence from a common ancestor 15.7 million years ago. Root development genes played a crucial role in spine development, such as endoplasmic reticulum pathway genes, jasmonic acid response genes, vascular bundle development genes, and K+ transport genes. Aril development was regulated by D-class genes (STK and SHP1), plant hormone and phenylpropanoid biosynthesis genes, and sugar metabolism genes. The lower rate of male sterility of hermaphroditic flowers appears to be regulated by MYB24. Population genomic analyses revealed genes in selective sweeps during domestication that are related to fruit morphology and environment stress response. These findings enhance our understanding of spine and aril development and provide genomic resources for rambutan improvement.

Integrating transcriptome and physiological analyses to elucidate the molecular responses of buckwheat to graphene oxide

With the increasing application of nanomaterials, evaluation of the phytotoxicity of nanoparticles has attracted considerable interest. Buckwheat is an economically pseudocereal crop, which is a potential model for investigating the response of plants to hazardous materials. In this study, the response of buckwheat to graphene oxide (GO) was investigated by integrating physiological and transcriptome analysis. GO can penetrate into buckwheat root and stem, and high concentrations of GO inhibited seedlings growth. High concentration of GO improved ROS production and regulated the activities and gene expression of oxidative enzymes, which implying GO may affect plant growth via regulating ROS detoxification. Root and stem exhibit distinct transcriptomic responses to GO, and the GO-responsive genes in stem are more enriched in cell cycle and epigenetic regulation. GO inhibited plant hormone biosynthesis and signaling by analyzing the expression data. Additionally, 97 small secreted peptides (SSPs) encoding genes were found to be involved in GO response. The gene expression of 111 transcription factor (TFs) and 43 receptor-like protein kinases (RLKs) were regulated by GO, and their expression showed high correlation with SSPs. Finally, the TFs-SSPs-RLKs signaling networks in regulating GO response were proposed. This study provides insights into the molecular responses of plants to GO.

Molybdoenzyme Participation In Plant Biochemical Processes

Molybdenum is a key microelement in plant vital functioning. The microelement can be absorbed by the plants only in the form of molybdate-anion. The Molybdenum deficiency affects negatively to the most important agricultural growing. As molybdenum takes part in such vital mechanisms as nitrogen and sulfur metabolism, plant hormone biosynthesis, and purine banding catabolism. Molybdenum is included in enzyme content which is called molybdoenzyms. Having bonded with molybdopterin it creates molybdenum co-factor (Moco) and gets oxidation-reduction properties. Moco is included in active site of molybdoenzymes. They take part in sulfur and nitrogen metabolism, and detrimental compound detoxication. Molybdenum deficiency is characterized by the slow plant growth, low amount of chlorophyll ascorbic acid capacity.It was noticed that plants suffering from the molybdenum deficiency can be saved, sodium molibdate can be used, it can be put directly in the soil or plant leaves can sprayed with the solution. There are five plant molibdoenzymes which are currently known: sulfite oxidase (SO), xanthine dehydrogenase (XD), nitrate reductase (NR), aldehyde oxidase (AO) and mitochondrial amidoxim-regenerative component. Nitrate reductase catalyzes the first stage of nitrate assimilation, eucariotic organisms contain three isoforms of the molybdoezimes: A NADH, A NAD(P)H и NADPH. Xanthine dehydrogenase regulates purine metabolism. XD increases plant antioxidant ability and slows down leaves aging. Molybdoenzymess are involved in the process of the stress adaptation, defining of the mechanisms and their reaction to environmental stress conditions is important for plant stress resistance.

Transcriptome Reveals Allele Contribution to Heterosis in Maize.

Heterosis, which has greatly increased maize yields, is associated with gene expression patterns during key developmental stages that enhance hybrid phenotypes relative to parental phenotypes. Before heterosis can be more effectively used for crop improvement, hybrid maize developmental gene expression patterns must be better understood. Here, six maize hybrids, including the popular hybrid Zhengdan958 (ZC) from China, were studied. Maize hybrids created in-house were generated using an incomplete diallel cross (NCII)-based strategy from four elite inbred parental lines. Differential gene expression (DEG) profiles corresponding to three developmental stages revealed that hybrid partial expression patterns exhibited complementarity of expression of certain parental genes, with parental allelic expression patterns varying both qualitatively and quantitatively in hybrids. Single-parent expression (SPE) and parent-specific expression (PSE) types of qualitative variation were most prevalent, 43.73 and 41.07% of variation, respectively. Meanwhile, negative super-dominance (NSD) and positive super-dominance (PSD) types of quantitative variation were most prevalent, 31.06 and 24.30% of variation, respectively. During the early reproductive growth stage, the gene expression pattern differed markedly from other developmental stage patterns, with allelic expression patterns during seed development skewed toward low-value parental alleles in hybrid seeds exhibiting significant quantitative variation-associated superiority. Comparisons of qualitative gene expression variation rates between ZC and other hybrids revealed proportions of SPE-DEGs (41.36%) in ZC seed DEGs that significantly exceeded the average proportion of SPE-DEGs found in seeds of other hybrids (28.36%). Importantly, quantitative gene expression variation rate comparisons between ZC and hybrids, except for transgressive expression, revealed that the ZC rate exceeded the average rate for other hybrids, highlighting the importance of partial gene expression in heterosis. Moreover, enriched ZC DEGs exhibiting distinct tissue-specific expression patterns belonged to four biological pathways, including photosynthesis, plant hormone signal transduction, biology metabolism and biosynthesis. These results provide valuable technical insights for creating hybrids exhibiting strong heterosis.

Transcriptomic and Metabolomic Analyses Provide Insights Into an Aberrant Tissue of Tea Plant (Camellia sinensis).

Tea plant (Camellia sinensis (L.) O. Kuntze) is one of the most important economic crops with multiple mutants. Recently, we found a special tea germplasm that has an aberrant tissue on its branches. To figure out whether this aberrant tissue is associated with floral bud (FB) or dormant bud (DB), we performed tissue section, transcriptome sequencing, and metabolomic analysis of these tissues. Longitudinal sections indicated the aberrant tissue internal structure was more like a special bud (SB), but was similar to that of DB. Transcriptome data analysis showed that the number of heterozygous and homozygous SNPs was significantly different in the aberrant tissue compared with FB and DB. Further, by aligning the unmapped sequences of the aberrant tissue to the Non-Redundant Protein Sequences (NR) database, we observed that 36.13% of unmapped sequences were insect sequences, which suggested that the aberrant tissue might be a variation of dormant bud tissue influenced by the interaction of tea plants and insects or pathogens. Metabolomic analysis showed that the differentially expressed metabolites (DEMs) between the aberrant tissue and DB were significantly enriched in the metabolic pathways of biosynthesis of plant hormones and biosynthesis of phenylpropanoids. Subsequently, we analyzed the differentially expressed genes (DEGs) in the above mentioned two tissues, and the results indicated that photosynthetic capacity in the aberrant tissue was reduced, whereas the ethylene, salicylic acid and jasmonic acid signaling pathways were activated. We speculated that exogenous infection induced programmed cell death (PCD) and increased the lignin content in dormant buds of tea plants, leading to the formation of this aberrant tissue. This study advanced our understanding of the interaction between plants and insects or pathogens, providing important clues about biotic stress factors and key genes that lead to mutations and formation of the aberrant tissue.

Sulfuric Acid and Gibberellic Acid (GA3) Treatment Combined with Exposure to Cold Temperature Modulates Seed Proteins during Breaking of Dormancy to Germination in Tilia miqueliana.

Tilia miqueliana produces woody seeds that exhibit deep dormancy. In this study, we used cell biology methods, including Paraffin section determination and Coomassie brilliant blue staining, as well as proteomics-based methods, including two-dimensional electrophoresis with matrix-assisted laser desorption/ionisation tandem time-of-flight mass spectrometry (2DE-MALDI-TOF/TOF), to examine the effects of H2SO4-GA3 and cold stratification (3°C) treatment on proteins during dormancy release and germination in T. miqueliana seeds. The results revealed that during cold stratification, the area and density of proteins in the endosperm cells of H2SO4-GA3-treated seeds were significantly altered. Total protein content was continuously consumed and utilised. Storage proteins (albumin, globulin, prolamin, and glutelin) were degraded to varying degrees. Sixteen differential proteins were identified using mass spectrometry. Kyoto encyclopedia of genes (KEGG) pathway analysis revealed that the glycolysis/gluconeogenesis, secondary metabolite biosynthesis, glyoxylate and dicarboxylate metabolism, amino acid biosynthesis, and metabolic pathways were critical during dormancy release and germination. Gene ontology analysis and KEGG pathway annotation of differential proteins in the co-expression network indicated that the differential proteins are implicated in photosynthesis, glucose metabolism, biosynthesis of plant hormones, and glycolysis/gluconeogenesis. Synergistic interactions among these proteins accelerated dormancy release and germination. Therefore, H2SO4-GA3 cold stratification treatment is the best method for achieving rapid dormancy release and increasing the germination rate of T. miqueliana seeds.

Metabolite profiling and transcriptome analyses reveal novel regulatory mechanisms of melatonin biosynthesis in hickory

Studies have shown that melatonin regulates the expression of various elements in the biosynthesis and catabolism of plant hormones. In contrast, the effects of these different plant hormones on the biosynthesis and metabolism of melatonin and their underlying molecular mechanisms are still unclear. In this study, the melatonin biosynthesis pathway was proposed from constructed metabolomic and transcriptomic libraries from hickory (Carya cathayensis Sarg.) nuts. The candidate pathway genes were further identified by phylogenetic analysis, amino-acid sequence alignment, and subcellular localization. Notably, most of the transcription factor-related genes coexpressed with melatonin pathway genes were hormone-responsive genes. Furthermore, dual-luciferase and yeast one‐hybrid assays revealed that CcEIN3 (response to ethylene) and CcAZF2 (response to abscisic acid) could activate melatonin biosynthesis pathway genes, a tryptophan decarboxylase coding gene (CcTDC1) and an N-acetylserotonin methyltransferase coding gene (CcASMT1), by directly binding to their promoters, respectively. Our results provide a molecular basis for the characterization of novel melatonin biosynthesis regulatory mechanisms and demonstrate for the first time that abscisic acid and ethylene can regulate melatonin biosynthesis.

Embryogenic Calli Induction and Salt Stress Response Revealed by RNA-Seq in Diploid Wild Species Gossypium sturtianum and Gossypium raimondii.

Wild cotton species can contribute to a valuable gene pool for genetic improvement, such as genes related to salt tolerance. However, reproductive isolation of different species poses an obstacle to produce hybrids through conventional breeding. Protoplast fusion technology for somatic cell hybridization provides an opportunity for genetic manipulation and targeting of agronomic traits. Transcriptome sequencing analysis of callus under salt stress is conducive to study salt tolerance genes. In this study, calli were induced to provide materials for extracting protoplasts and also for screening salt tolerance genes. Calli were successfully induced from leaves of Gossypium sturtianum (C1 genome) and hypocotyls of G. raimondii (D5 genome), and embryogenic calli of G. sturtianum and G. raimondii were induced on a differentiation medium with different concentrations of 2, 4-D, KT, and IBA, respectively. In addition, embryogenic calli were also induced successfully from G. raimondii through suspension cultivation. Transcriptome sequencing analysis was performed on the calli of G. raimondii and G. sturtianum, which were treated with 200 mM NaCl at 0, 6, 12, 24, and 48 h, and a total of 12,524 genes were detected with different expression patterns under salt stress. Functional analysis showed that 3,482 genes, which were differentially expressed in calli of G. raimondii and G. sturtianum, were associated with biological processes of nucleic acid binding, plant hormone (such as ABA) biosynthesis, and signal transduction. We demonstrated that DEGs or TFs which related to ABA metabolism were involved in the response to salt stress, including xanthoxin dehydrogenase genes (ABA2), sucrose non-fermenting 1-related protein kinases (SnRK2), NAM, ATAT1/2, and CUC2 transcription factors (NAC), and WRKY class of zinc-finger proteins (WRKY). This research has successfully induced calli from two diploid cotton species and revealed new genes responding to salt stress in callus tissue, which will lay the foundation for protoplast fusion for further understanding of salt stress responses in cotton.

Transcriptome Analysis Revealed Plant Hormone Biosynthesis and Response Pathway Modification by Epichloëgansuensis in Achnatheruminebrians under Different Soil Moisture Availability.

The present study was designed to explore the effects of the endophyte Epichloë gansuensis on gene expression related to plant hormone biosynthesis and response pathways and the content of salicylic acid (SA) and jasmonic acid (JA) hormones of Achnatherum inebrians, under different moisture conditions. Through a pot experiment and transcriptome analysis, we found a total of 51 differentially expressed genes (DEGs) related to hormone biosynthesis and response pathways, including 12 auxin related genes, 8 cytokinin (CTK) related genes, 3 gibberellin (GA) related genes, 7 abscisic acid (ABA) related genes, 7 ethylene (ET) related genes, 12 JA related genes and 4 SA related genes. Furthermore, key genes of JA and SA biosynthesis and response pathways, such as LOX2S, AOS, OPR, ACX, JMT, JAZ, PAL, NPR1, TGA and PR-1, showed different degrees of upregulation or downregulation. Under 60% soil moisture content, the JA content of endophyte-free (EF) A. inebrians was significantly (p < 0.05) higher than that of endophyte-infected (EI) A. inebrians. Under 30% and 60% soil moisture content, the SA content of EF A. inebrians was significantly (p < 0.05) higher than that of EI A. inebrians. SA content of EI A. inebrians under 30% and 60% soil moisture content was significantly (p < 0.05) higher than that under 15% soil moisture content. With both EI and EF plants, the SA and JA levels, respectively, are very similar at 15% soil moisture content. This study has revealed that E. gansuensis differentially activated plant hormone synthesis and signal transduction pathways of A. inebrians plants under different soil moisture availability.

Metabolomic Analysis Provides New Insight Into Tolerance of Huanglongbing in Citrus.

There have been efforts to develop citrus cultivars that are tolerant of Huanglongbing (HLB), a catastrophic phloem-limited disease. Previous studies demonstrated that continuous plant growth with phloem regeneration is one of the major characteristics of HLB tolerance. In this study, the metabolic mechanisms of HLB tolerance in citrus were elucidated using a multiple pathway-targeted metabolomic approach. Comparative analysis of healthy and infected HLB-tolerant and HLB-sensitive mandarin cultivars (Citrus reticulata) revealed differentially expressed metabolic responses among different groups. Pathway enrichment analysis indicated aspartate and glutamate metabolism, purine metabolism, and biosynthesis of plant hormones were upregulated in the tolerant group, except salicylic acid signaling. Catabolic pathways linked to energy-yielding metabolism were also upregulated in the tolerant group. These metabolisms and pathways were interconnected with each other, unveiling a pivotal metabolic network associated with HLB tolerance. In the network, auxins and cytokinins, the plant hormones responsible for plant growth and phloem regeneration, were accumulated. In addition, purine metabolites serving as energy carriers and nitrogen sources of plants were increased. Only salicylic acid-related metabolites for plant defense responses were decreased in the tolerant group. Our findings may evidence the strategy of HLB-tolerant cultivars that sustain plant growth and phloem formation rather than displaying direct plant defense to overcome the disease.

Characterization of microRNAs involved in asymbiotic germination of Bletilla striata (Orchidaceae) seeds

Orchids are distributed worldwide, and some species have considerable economic value. Orchid seeds are minute in size, simple in structure, and deficient in nutrient reserves. Asymbiotic seed germination is an important propagation strategy for orchids. MicroRNAs (miRNAs) play an essential role in seed germination. However, few studies have examined miRNAs involved in seed germination in orchids. Here, we conducted comparative small RNA sequencing at five stages to characterize the miRNAs involved in asymbiotic seed germination in Bletilla striata. A total of 253 known and 125 novel miRNAs were identified. Of them, 71 known and 29 novel miRNAs showed distinct expression among the five stages. Quantitative PCR revealed negative correlations of expression between differentially expressed miRNAs (DE miRNAs) and their targets. Function annotation and enrichment analyses of the targets of DE miRNAs between adjacent stages indicate that miRNA-target regulations are involved in many important processes during germination, such as signaling, biosynthesis, and transport of plant hormones. Twenty-two miRNAs were inferred to participate in plant hormone-related processes. The contents of abscisic acid, gibberellin A3, indole-3-acetic acid, jasmonic acid, trans zeatin riboside, and N6-(Δ2-isopentenyl) adenine varied significantly among the five stages. Nine tested plant hormone-related miRNAs and their targets exhibited significant correlations with at least one plant hormone. 5′-RLM-RACE validated that a transcript encoding auxin response factor was cleaved by Bst-miR160e as predicted. For the first time, we characterized miRNAs associated with the asymbiotic seed germination of an orchid species, which will help understand the miRNA-mediated regulatory mechanism of seed germination in orchids.