Exogenous strigolactone enhanced the drought tolerance of pepper (Capsicum chinense) by mitigating oxidative damage and altering the antioxidant mechanism.

KEY MESSAGE: Exogenous SL positively regulates pepper DS by altering the root morphology, photosynthetic character, antioxidant enzyme activity, stomatal behavior, and SL-related gene expression. Drought stress (DS) has always been a problem for the growth and development of crops, causing significant negative impacts on crop productivity. Strigolactone (SL) is a newly discovered class of plant hormones that are involved in plants' growth and development and environmental stresses. However, the role of SL in response to DS in pepper remains unknown. DS considerably hindered photosynthetic pigments content, damaged root architecture system, and altered antioxidant machinery. In contrast, SL application significantly restored pigment concentration modified root architecture system, and increased relative chlorophyll content (SPAD). Additionally, SL treatment reduced oxidative damage by reducing hydrogen peroxide (H2O2) (24-57%) and malondialdehyde (MDA) (79-89%) accumulation in pepper seedlings. SL-pretreated pepper seedlings showed significant improvement in antioxidant enzyme activity, proline accumulation, and soluble sugar content. Furthermore, SL-related genes (CcSMAX2, CcSMXL6, and CcSMXL3) were down-regulated under DS. These findings suggest that the foliar application of SL can alleviate the adverse effects of drought tolerance by up-regulating chlorophyll content and activating antioxidant defense mechanisms.

A comprehensive analysis of transcriptomic data for comparison of cold tolerance in two Brassica napus genotypes.

Brassica napus is an important oil crop and cold stress severely limits its productivity. To date, several studies have reported the regulatory genes and pathways involved in cold-stress responses in B. napus. However, transcriptome-scale identification of the regulatory genes is still lacking. In this study, we performed comparative transcriptome analysis of cold-tolerant C18 (CT - C18) and cold-sensitive C6 (CS - C6) Brassica napus genotypes under cold stress for 7 days, with the primary purpose of identifying cold-responsive transcription in B. napus. A total of 6061 TFs belonging to 58 families were annotated in the B. napus genome, of which 3870 were expressed under cold stress in both genotypes. Among these, 451 TFs were differentially expressed (DE), with 21 TF genes expressed in both genotypes. Most TF members of the MYB (26), bHLH (23), and NAC (17) families were significantly expressed in the CT - C18 genotype compared with the CS - C6 B. napus genotype. GO classification showed a significant role in transcription regulation, DNA-binding transcription factor activity, response to chitin, and the ethylene-activated signaling pathway. KEGG pathway annotation revealed these TFs are involved in regulating more pathways, resulting in more tolerance. In conclusion, the results provide insights into the molecular regulation mechanisms of B. napus in response to freezing treatment, expanding our understanding of the complex molecular mechanisms in plants' response to freezing stress.

Evolution and Dynamic Transcriptome of Key Genes of Photoperiodic Flowering Pathway in Water Spinach (Ipomoea aquatica).

The photoperiod is a major environmental factor in flowering control. Water spinach flowering under the inductive short-day condition decreases the yield of vegetative tissues and the eating quality. To obtain an insight into the molecular mechanism of the photoperiod-dependent regulation of the flowering time in water spinach, we performed transcriptome sequencing on water spinach under long- and short-day conditions with eight time points. Our results indicated that there were 6615 circadian-rhythm-related genes under the long-day condition and 8691 under the short-day condition. The three key circadian-rhythm genes, IaCCA1, IaLHY, and IaTOC1, still maintained single copies and similar IaCCA1, IaLHY, and IaTOC1 feedback expression patterns, indicating the conservation of reverse feedback. In the photoperiod pathway, highly conserved GI genes were amplified into two copies (IaGI1 and IaGI2) in water spinach. The significant difference in the expression of the two genes indicates functional diversity. Although the photoperiod core gene FT was duplicated to three copies in water spinach, only IaFT1 was highly expressed and strongly responsive to the photoperiod and circadian rhythms, and the almost complete inhibition of IaFT1 in water spinach may be the reason why water spinach does not bloom, no matter how long it lasts under the long-day condition. Differing from other species (I. nil, I. triloba, I. trifida) of the Ipomoea genus that have three CO members, water spinach lacks one of them, and the other two CO genes (IaCO1 and IaCO2) encode only one CCT domain. In addition, through weighted correlation network analysis (WGCNA), some transcription factors closely related to the photoperiod pathway were obtained. This work provides valuable data for further in-depth analyses of the molecular regulation of the flowering time in water spinach and the Ipomoea genus.

Physiological and Transcriptome Analysis of the Effects of Exogenous Strigolactones on Drought Responses of Pepper Seedlings.

Drought stress significantly restricts the growth, yield, and quality of peppers. Strigolactone (SL), a relatively new plant hormone, has shown promise in alleviating drought-related symptoms in pepper plants. However, there is limited knowledge on how SL affects the gene expression in peppers when exposed to drought stress (DS) after the foliar application of SL. To explore this, we conducted a thorough physiological and transcriptome analysis investigation to uncover the mechanisms through which SL mitigates the effects of DS on pepper seedlings. DS inhibited the growth of pepper seedlings, altered antioxidant enzyme activity, reduced relative water content (RWC), and caused oxidative damage. On the contrary, the application of SL significantly enhanced RWC, promoted root morphology, and increased leaf pigment content. SL also protected pepper seedlings from drought-induced oxidative damage by reducing MDA and H2O2 levels and maintaining POD, CAT, and SOD activity. Moreover, transcriptomic analysis revealed that differentially expressed genes were enriched in ribosomes, ABC transporters, phenylpropanoid biosynthesis, and Auxin/MAPK signaling pathways in DS and DS + SL treatment. Furthermore, the results of qRT-PCR showed the up-regulation of AGR7, ABI5, BRI1, and PDR4 and down-regulation of SAPK6, NTF4, PYL6, and GPX4 in SL treatment compared with drought-only treatment. In particular, the key gene for SL signal transduction, SMXL6, was down-regulated under drought. These results elucidate the molecular aspects underlying SL-mediated plant DS tolerance, and provide pivotal strategies for effectively achieving pepper drought resilience.

Genome-Wide Identification of WRKY Gene Family and Functional Characterization of CcWRKY25 in Capsicum chinense.

Pepper is renowned worldwide for its distinctive spicy flavor. While the gene expression characteristics of the capsaicinoid biosynthesis pathway have been extensively studied, there are already a few reports regarding transcriptional regulation in capsaicin biosynthesis. In this study, 73 WRKYs were identified in the genome of Capsicum chinense, and their physicochemical traits, DNA, and protein sequence characteristics were found to be complex. Combining RNA-seq and qRT-PCR data, the WRKY transcription factor CA06g13580, which was associated with the accumulation tendency of capsaicinoid, was screened and named CcWRKY25. CcWRKY25 was highly expressed in the placenta of spicy peppers. The heterologous expression of CcWRKY25 in Arabidopsis promoted the expression of genes PAL, 4CL1, 4CL2, 4CL3, CCR, and CCoAOMT and led to the accumulation of lignin and flavonoids. Furthermore, the expression of the capsaicinoid biosynthesis pathway genes (CBGs) pAMT, AT3, and KAS was significantly reduced in CcWRKY25-silenced pepper plants, resulting in a decrease in the amount of capsaicin. However, there was no noticeable difference in lignin accumulation. The findings suggested that CcWRKY25 could be involved in regulating capsaicinoid synthesis by promoting the expression of genes upstream of the phenylpropanoid pathway and inhibiting CBGs' expression. Moreover, the results highlighted the role of CcWRKY25 in controlling the pungency of pepper and suggested that the competitive relationship between lignin and capsaicin could also regulate the spiciness of the pepper.

Melatonin enhanced the heavy metal-stress tolerance of pepper by mitigating the oxidative damage and reducing the heavy metal accumulation.

Heavy metals (HMs), like vanadium (V), chromium (Cr), cadmium (Cd), and nickel (Ni) toxicity due to anthropogenic, impair plant growth and yield, which is a challenging issue for agricultural production. Melatonin (ME) is a stress mitigating molecule, which alleviates HM-induced phytotoxicity, but the possible underlying mechanism of ME functions under HMs' phytotoxicity is still unclear. Current study uncovered key mechanisms for ME-mediated HMs-stress tolerance in pepper. HMs toxicity greatly reduced growth by impeding leaf photosynthesis, root architecture system, and nutrient uptake. Conversely, ME supplementation markedly enhanced growth attributes, mineral nutrient uptake, photosynthetic efficiency, as measured by chlorophyll content, gas exchange elements, chlorophyll photosynthesis genes' upregulation, and reduced HMs accumulation. ME treatment showed a significant decline in the leaf/root V, Cr, Ni, and Cd concentration which was about 38.1/33.2%, 38.5/25.9%, 34.8/24.9%, and 26.6/25.1%, respectively, when compared with respective HM treatment. Furthermore, ME remarkably reduced the ROS (reactive oxygen species) accumulation, and reinstated the integrity of cellular membrane via activating antioxidant enzymes (SOD, superoxide dismutase; CAT, catalase; APX, ascorbate peroxidase; GR, glutathione reductase; POD, peroxidase; GST, glutathione S-transferase; DHAR, dehydroascorbate reductase; MDHAR, monodehydroascorbate reductase) and as well as regulating ascorbate-glutathione (AsA-GSH) cycle. Importantly, oxidative damage showed efficient alleviations through upregulating the genes related to key defense such as SOD, CAT, POD, GR, GST, APX, GPX, DHAR, and MDHAR; along with the genes related to ME biosynthesis. ME supplementation also enhanced the level of proline and secondary metabolites, and their encoding genes expression, which may control excessive H2O2 (hydrogen peroxide) production. Finally, ME supplementation enhanced the HM stress tolerance of pepper seedlings.

The creation of autotetraploid provides insights into critical features of DNA methylome changes after genome doubling in water spinach (Ipomoea aquatica Forsk).

Water spinach (Ipomoea aquatica Forsk) is an essential green leafy vegetable in Asia. In this study, we induced autotetraploid water spinach by colchicine. Furthermore, DNA methylation and transcriptome of tetraploid and diploid were compared using Whole Genome Bisulfite Sequencing (WGBS) and RNA-sequencing techniques. Autotetraploid water spinach was created for the first time. Compared with the diploid parent, autotetraploid water spinach had wider leaves, thicker petioles and stems, thicker and shorter adventitious roots, longer stomas, and larger parenchyma cells. The whole genome methylation level of the autotetraploid was slightly higher than that of the diploid. Compared with the diploid, 12281 Differentially Methylated Regions (DMRs)were found in the autotetraploid, including 2356 hypermethylated and 1310 hypomethylated genes, mainly enriched in 'Arginine and Proline metabolism', 'beta - Alanine metabolism', 'Plant homone signal translation', 'Ribome', and 'Plant - pathgen interaction' pathways. Correlation analysis of transcriptome and DNA methylation data showed that 121 differentially expressed genes undergone differential methylation, related to four pathways 'Other types of O-glycan biosynthesis', 'Terpenoid backbone biosynthesis', 'Biosynthesis of secondary metabolites', and 'Metabolic paths'. This work obtained important autotetraploid resources of water spinach and revealed the genomic DNA methylation changes after genome doubling, being helpful for further studying the molecular mechanism of variations caused by polyploids of the Ipomoea genus.

Analysis of Volatile Aroma Components and Regulatory Genes in Different Kinds and Development Stages of Pepper Fruits Based on Non-Targeted Metabolome Combined with Transcriptome.

Aroma is a crucial attribute affecting the quality of pepper and its processed products, which has significant commercial value. However, little is known about the composition of volatile aroma compounds (VACs) in pepper fruits and their potential molecular regulatory mechanisms. In this study, HS-SPME-GC-MS combined with transcriptome sequencing is used to analyze the composition and formation mechanism of VACs in different kinds and development stages of pepper fruits. The results showed that 149 VACs, such as esters, alcohols, aldehydes, and terpenoids, were identified from 4 varieties and 3 development stages, and there were significant quantitative differences among different samples. Volatile esters were the most important aroma components in pepper fruits. PCA analysis showed that pepper fruits of different developmental stages had significantly different marker aroma compounds, which may be an important provider of pepper's characteristic aroma. Transcriptome analysis showed that many differential genes (DEGs) were enriched in the metabolic pathways related to the synthesis of VACs, such as fatty acids, amino acids, MVA, and MEP in pepper fruits. In addition, we identified a large number of differential transcription factors (TFs) that may regulate the synthesis of VACs. Combined analysis of differential aroma metabolites and DEGs identified two co-expression network modules highly correlated with the relative content of VACs in pepper fruit. This study confirmed the basic information on the changes of VACs in the fruits of several Chinese spicy peppers at different stages of development, screened out the characteristic aroma components of different varieties, and revealed the molecular mechanism of aroma formation, providing a valuable reference for the quality breeding of pepper.

MYB24 Negatively Regulates the Biosynthesis of Lignin and Capsaicin by Affecting the Expression of Key Genes in the Phenylpropanoid Metabolism Pathway in Capsicum chinense.

The wide application of pepper is mostly related to the content of capsaicin, and phenylpropanoid metabolism and its branch pathways may play an important role in the biosynthesis of capsaicin. The expression level of MYB24, a transcription factor screened from the transcriptome data of the pepper fruit development stage, was closely related to the spicy taste. In this experiment, CcMYB24 was cloned from Hainan Huangdenglong pepper, a hot aromatic pepper variety popular in the world for processing, and its function was confirmed by tissue expression characteristics, heterologous transformation in Arabidopsis thaliana, and VIGS technology. The results showed that the relative expression level of CcMYB24 was stable in the early stage of pepper fruit development, and increased significantly from 30 to 50 days after flowering. Heterologous expression led to a significant increase in the expression of CcMYB24 and decrease in lignin content in transgenic Arabidopsis thaliana plants. CcMYB24 silencing led to a significant increase in the expression of phenylpropanoid metabolism pathway genes PAL, 4CL, and pAMT; lignin branch CCR1 and CAD; and capsaicin pathway CS, AT3, and COMT genes in the placenta of pepper, with capsaicin content increased by more than 31.72% and lignin content increased by 20.78%. However, the expression of PAL, pAMT, AT3, COMT, etc., in the corresponding pericarps did not change significantly. Although CS, CCR1, and CAD increased significantly, the relative expression amount was smaller than that in placental tissue, and the lignin content did not change significantly. As indicated above, CcMYB24 may negatively regulate the formation of capsaicin and lignin by regulating the expression of genes from phenylpropanoid metabolism and its branch pathways.

Analysis of the molecular and biochemical mechanisms involved in the symbiotic relationship between Arbuscular mycorrhiza fungi and Manihot esculenta Crantz.

Introduction: Plants and arbuscular mycorrhizal fungi (AMF) mutualistic interactions are essential for sustainable agriculture production. Although it is shown that AMF inoculation improves cassava physiological performances and yield traits, the molecular mechanisms involved in AM symbiosis remain largely unknown. Herein, we integrated metabolomics and transcriptomics analyses of symbiotic (Ri) and asymbiotic (CK) cassava roots and explored AM-induced biochemical and transcriptional changes. Results: Three weeks (3w) after AMF inoculations, proliferating fungal hyphae were observable, and plant height and root length were significantly increased. In total, we identified 1,016 metabolites, of which 25 were differentially accumulated (DAMs) at 3w. The most highly induced metabolites were 5-aminolevulinic acid, L-glutamic acid, and lysoPC 18:2. Transcriptome analysis identified 693 and 6,481 differentially expressed genes (DEGs) in the comparison between CK (3w) against Ri at 3w and 6w, respectively. Functional enrichment analyses of DAMs and DEGs unveiled transport, amino acids and sugar metabolisms, biosynthesis of secondary metabolites, plant hormone signal transduction, phenylpropanoid biosynthesis, and plant-pathogen interactions as the most differentially regulated pathways. Potential candidate genes, including nitrogen and phosphate transporters, transcription factors, phytohormone, sugar metabolism-related, and SYM (symbiosis) signaling pathway-related, were identified for future functional studies. Discussion: Our results provide molecular insights into AM symbiosis and valuable resources for improving cassava production.

Glucosinolate Biosynthetic Genes of Cabbage: Genome-Wide Identification, Evolution, and Expression Analysis.

Cabbage (Brassica oleracea var. capitata) is a vegetable rich in glucosinolates (GSLs) that have proven health benefits. To gain insights into the synthesis of GSLs in cabbage, we systematically analyzed GSLs biosynthetic genes (GBGs) in the entire cabbage genome. In total, 193 cabbage GBGs were identified, which were homologous to 106 GBGs in Arabidopsis thaliana. Most GBGs in cabbage have undergone negative selection. Many homologous GBGs in cabbage and Chinese cabbage differed in expression patterns indicating the unique functions of these homologous GBGs. Spraying five exogenous hormones significantly altered expression levels of GBGs in cabbage. For example, MeJA significantly upregulated side chain extension genes BoIPMILSU1-1 and BoBCAT-3-1, and the expression of core structure construction genes BoCYP83A1 and BoST5C-1, while ETH significantly repressed the expression of side chain extension genes such as BoIPMILSU1-1, BoCYP79B2-1, and BoMAMI-1, and some transcription factors, namely BoMYB28-1, BoMYB34-1, BoMYB76-1, BoCYP79B2-1, and BoMAMI-1. Phylogenetically, the CYP83 family and CYP79B and CYP79F subfamilies may only be involved in GSL synthesis in cruciferous plants. Our unprecedented identification and analysis of GBGs in cabbage at the genome-wide level lays a foundation for the regulation of GSLs synthesis through gene editing and overexpression.

Establishment of in vitro regeneration system and molecular analysis of early development of somatic callus in Capsicum chinense and Capsicum baccatum.

Regeneration is extremely important to pepper genetic development; however, the molecular mechanisms of how the callus reactivates cell proliferation and promotes cell reprogramming remain elusive in pepper. In the present study, C. baccatum (HNUCB81 and HNUCB226) and C. chinense (HNUCC22 and HNUCC16) were analyzed to reveal callus initiation by in vitro regeneration, histology, and transcriptome. We successfully established an efficient in vitro regeneration system of two cultivars to monitor the callus induction of differential genotypes, and the regenerated plants were obtained. Compared to C. chinense, there was a higher callus induction rate in C. baccatum. The phenotype of C. baccatum changed significantly and formed vascular tissue faster than C. chinense. The KEGG enrichment analysis found that plant hormone transduction and starch and sucrose metabolism pathways were significantly enriched. In addition, we identified that the WOX7 gene was significantly up-regulated in HNUCB81 and HNUCB226 than that in HNUCC22 and HNUCC16, which may be a potential function in callus formation. These results provided a promising strategy to improve the regeneration and transformation of pepper plants.

Insight into the effect of low temperature treatment on trichome density and related differentially expressed genes in Chinese cabbage.

Trichome is important for help plant resist adversity and external damage. However, it often affects the appearance and taste of vegetables. In the present study, the trichome density of leaves from two Chinese cabbage cultivars with and without trichomes treated at low temperature are analyzed by biological microscope, and the differentially expressed genes related to trichomes formation were screened through transcriptome sequencing. The results showed that the number of leaves trichomes was reduced by 34.7% at low temperature compared with room temperature. A total of 661 differentially expression genes effecting trichomes formation were identified at the CT vs C, LCT vs LC, CT vs LCT. Several differentially expression genes from every comparison group were enriched in plant hormone signal transduction and amino acid biosynthesis pathway. Combined with the central genes obtained by WGCNA analysis, five candidate genes Bra029778, Bra026393, Bra030270, Bra037264 and Bra009655 were screened. qRT-PCR analysis verified that the gene expression differences were in line with the trend of transcriptome data. This study not only found possible new key genes and laid a foundation for revealing the molecular mechanism regulating the formation of trichome in Chinese cabbage, but also provided a new way to study plant surface trichomes.

Response of Anthocyanin Accumulation in Pepper (Capsicum annuum) Fruit to Light Days.

Light is the key factor affecting the synthesis of anthocyanins in pepper. In this study, pepper fruit under different light days was used as experimental material to explore the synthesis of anthocyanins in purple pepper. A total of 38 flavonoid metabolites were identified in the purple pepper germplasm HNUCA21 by liquid chromatography-tandem mass spectrometry (LC-MS/MS), of which 30 belong to anthocyanins. The detected anthocyanin with the highest content was Delphinidin-3-O-glucoside (17.13 µg/g), which reached the maximum after 168 h of light treatment. Through weighted gene co-expression network analysis (WGCNA), the brown module was identified to be related to the early synthesis of anthocyanins. This module contains many structural genes related to flavonoid synthesis, including chalcone synthase (CHS 107871256, 107864266), chalcone isomerase (CHI 107871144, 107852750), dihydroflavonol 4-reductase (DFR 107860031), flavonoid 3' 5'-hydroxylase (F3'5'H 107848667), flavonoid 3'-monooxygenase (F3M 107862334), leucoanthocyanidin dioxygenase (LDOX 107866341), and trans-cinnamate 4-monooxygenase (TCM 107875406, 107875407). The module also contained some genes related to anthocyanin transport function, such as glutathione S-transferase (GST 107861273), anthocyanidin 3-O-glucosyltransferase (UDPGT 107861697, 107843659), and MATE (107863234, 107844661), as well as some transcription factors, such as EGL1 (107865400), basic helix-loop-helix 104 (bHLH104 107864591), and WRKY44 (107843538, 107843524). The co-expression regulatory network indicated the involvement of CHS, DFR, CHI, and EGL1, as well as two MATE and two WRKY44 genes in anthocyanin synthesis. The identified genes involved in early, middle, and late light response provided a reference for the further analysis of the regulatory mechanism of anthocyanin biosynthesis in pepper.

Integrated Transcriptomic and Metabolomic Analyses of Cold-Tolerant and Cold-Sensitive Pepper Species Reveal Key Genes and Essential Metabolic Pathways Involved in Response to Cold Stress.

Cold stress, triggered by particularly low temperatures, is one of the most severe forms of abiotic stress in pepper plants and a major constraint to the global pepper industry, threatening crop production and food security. To acclimatize to extreme conditions, the plant undergoes numerous modifications, including genetic and metabolic modulations. A thorough study of both the genetic and metabolic alterations of plants in response to cold stress is vital to understanding and developing the cold stress resistance mechanism. This study implemented transcriptome and metabolome analyses to evaluate the cold stress response in cold-tolerant and cold-sensitive pepper species. The weighted gene co-expression network revealed three significant modules related to cold stress tolerance in Capsicum pubescens. We identified 17 commonly enriched genes among both species at different time points in 10 different comparisons, including the AP2 transcription factor, LRR receptor-like serine, hypersensitivity-related 4-like protein, and uncharacterized novel.295 and novel.6172 genes. A pathway enrichment analysis indicated that these DEGs were mainly associated with the MAPK signaling pathway, hormone signaling pathway, and primary and secondary metabolism. Additionally, 21 significantly differentially accumulated metabolites (DAMs) were identified in both species after 6 h of cold stress. A transcriptome and metabolome integrated analysis revealed that 54 genes correlated with metabolites enriched in five different pathways. Most genes and metabolites involved in carbohydrate metabolism, the TCA cycle, and flavonoid biosynthesis pathways were upregulated in cold-tolerant plants under cold stress. Together, the results of this study provide a comprehensive gene regulatory and metabolic network in response to cold stress and identified some key genes and metabolic pathways involved in pepper cold tolerance. This study lays a foundation for the functional characterization and development of pepper cultivars with improved cold tolerance.

Response of anthocyanin biosynthesis to light by strand-specific transcriptome and miRNA analysis in Capsicum annuum.

BACKGROUND: Anthocyanins have distinct biological functions in plant coloring, plant defense against strong light, UV irradiation, and pathogen infection. Aromatic hydroxyl groups and ortho-dihydroxyl groups in anthocyanins are able to inhibit free-radical chain reactions and hydroxyl radicals. Thus, anthocyanins play an antioxidative role by removing various types of ROS. Pepper is one of the solanaceous vegetables with the largest cultivation area in China. The purple-fruited pepper is rich in anthocyanins, which not only increases the ornamental nature of the pepper fruit but also benefits the human body. In this experiment, light-induced regulatory pathways and related specific regulators of anthocyanin biosynthesis were examined through integrative transcriptomic and metabolomic analysis. RESULTS: Results revealed that delphinium 3-O-glucoside significantly accumulated in light exposed surface of pepper fruit after 48 h as compared to shaded surface. Furthermore, through strand-specific sequencing technology, 1341 differentially expressed genes, 172 differentially expressed lncRNAs, 8 differentially expressed circRNAs, and 28 differentially expressed miRNAs were identified significantly different among both surfaces. The flavonoid synthesis pathway was significantly enriched by KEGG analysis including SHT (XM_016684802.1), AT-like (XM_016704776.1), CCoAOMT (XM_016698340.1, XM_016698341.1), CHI (XM_016697794.1, XM_016697793.1), CHS2 (XM_016718139.1), CHS1B (XM_016710598.1), CYP98A2-like (XM_016688489.1), DFR (XM_016705224.1), F3'5'H (XM_016693437.1), F3H (XM_016705025.1), F3'M (XM_016707872.1), LDOX (XM_016712446.1), TCM (XM_016722116.1) and TCM-like (XM_016722117.1). Most of these significantly enriched flavonoid synthesis pathway genes may be also regulated by lncRNA. Some differentially expressed genes encoding transcription factors were also identified including MYB4-like (XM_016725242.1), MYB113-like (XM_016689220.1), MYB308-like (XM_016696983.1, XM_016702244.1), and EGL1 (XM_016711673.1). Three 'lncRNA-miRNA-mRNA' regulatory networks with sly-miR5303, stu-miR5303g, stu-miR7997a, and stu-miR7997c were constructed, including 28 differentially expressed mRNAs and 6 differentially expressed lncRNAs. CONCLUSION: Possible light regulated anthocyanin biosynthesis and transport genes were identified by transcriptome analysis, and confirmed by qRT-PCR. These results provide important data for further understanding of the anthocyanin metabolism in response to light in pepper.

Integrated Analysis of mRNA and Non-coding RNA Transcriptome in Pepper (Capsicum chinense) Hybrid at Seedling and Flowering Stages.

Pepper is an important vegetable in the world. In this work, mRNA and ncRNA transcriptome profiles were applied to understand the heterosis effect on the alteration in the gene expression at the seedling and flowering stages between the hybrid and its parents in Capsicum chinense. Our phenotypic data indicated that the hybrid has dominance in leaf area, plant scope, plant height, and fruit-related traits. Kyoto Encyclopedia of Genes and Genomes analysis showed that nine members of the plant hormone signal transduction pathway were upregulated in the seedling and flowering stages of the hybrid, which was supported by weighted gene coexpression network analysis and that BC332_23046 (auxin response factor 8), BC332_18317 (auxin-responsive protein IAA20), BC332_13398 (ethylene-responsive transcription factor), and BC332_27606 (ethylene-responsive transcription factor WIN1) were candidate hub genes, suggesting the important potential role of the plant hormone signal transduction in pepper heterosis. Furthermore, some transcription factor families, including bHLH, MYB, and HSF were greatly over-dominant. We also identified 2,525 long ncRNAs (lncRNAs), 47 micro RNAs (miRNAs), and 71 circle RNAs (circRNAs) in the hybrid. In particular, downregulation of miR156, miR169, and miR369 in the hybrid suggested their relationship with pepper growth vigor. Moreover, we constructed some lncRNA-miRNA-mRNA regulatory networks that showed a multi-dimension to understand the ncRNA relationship with heterosis. These results will provide guidance for a better understanding of the molecular mechanism involved in pepper heterosis.

Early Response of Radish to Heat Stress by Strand-Specific Transcriptome and miRNA Analysis.

Radish is a crucial vegetable crop of the Brassicaceae family with many varieties and large cultivated area in China. Radish is a cool season crop, and there are only a few heat tolerant radish varieties in practical production with little information concerning the related genes in response to heat stress. In this work, some physiological parameter changes of young leaves under short-term heat stress were detected. Furthermore, we acquired 1802 differentially expressed mRNAs (including encoding some heat shock proteins, heat shock factor and heat shock-related transcription factors), 169 differentially expressed lncRNAs and three differentially expressed circRNAs (novel_circ_0000265, novel_circ_0000325 and novel_circ_0000315) through strand-specific RNA sequencing technology. We also found 10 differentially expressed miRNAs (ath-miR159b-3p, athmiR159c, ath-miR398a-3p, athmiR398b-3p, ath-miR165a-5p, ath-miR169g-3p, novel_86, novel_107, novel_21 and ath-miR171b-3p) by small RNA sequencing technology. Through function prediction and enrichment analysis, our results suggested that the significantly possible pathways/complexes related to heat stress in radish leaves were circadian rhythm-plant, photosynthesis-antenna proteins, photosynthesis, carbon fixation in photosynthetic organisms, arginine and proline metabolism, oxidative phosphorylation, peroxisome and plant hormone signal transduction. Besides, we identified one lncRNA-miRNA-mRNAs combination responsive to heat stress. These results will be helpful for further illustration of molecular regulation networks of how radish responds to heat stress.

Discovery of putative capsaicin biosynthetic genes by RNA-Seq and digital gene expression analysis of pepper.

The Indian pepper 'Guijiangwang' (Capsicum frutescens L.), one of the world's hottest chili peppers, is rich in capsaicinoids. The accumulation of the alkaloid capsaicin and its analogs in the epidermal cells of the placenta contribute to the pungency of Capsicum fruits. To identify putative genes involved in capsaicin biosynthesis, RNA-Seq was used to analyze the pepper's expression profiles over five developmental stages. Five cDNA libraries were constructed from the total RNA of placental tissue and sequenced using an Illumina HiSeq 2000. More than 19 million clean reads were obtained from each library, and greater than 50% of the reads were assignable to reference genes. Digital gene expression (DGE) profile analysis using Solexa sequencing was performed at five fruit developmental stages and resulted in the identification of 135 genes of known function; their expression patterns were compared to the capsaicin accumulation pattern. Ten genes of known function were identified as most likely to be involved in regulating capsaicin synthesis. Additionally, 20 new candidate genes were identified related to capsaicin synthesis. We use a combination of RNA-Seq and DGE analyses to contribute to the understanding of the biosynthetic regulatory mechanism(s) of secondary metabolites in a nonmodel plant and to identify candidate enzyme-encoding genes.