TCOD: an integrated resource for tropical crops.

Tropical crops are vital for tropical agriculture, with resource scarcity, functional diversity and extensive market demand, providing considerable economic benefits for the world's tropical agriculture-producing countries. The rapid development of sequencing technology has promoted a milestone in tropical crop research, resulting in the generation of massive amount of data, which urgently needs an effective platform for data integration and sharing. However, the existing databases cannot fully satisfy researchers' requirements due to the relatively limited integration level and untimely update. Here, we present the Tropical Crop Omics Database (TCOD, https://ngdc.cncb.ac.cn/tcod), a comprehensive multi-omics data platform for tropical crops. TCOD integrates diverse omics data from 15 species, encompassing 34 chromosome-level de novo assemblies, 1 255 004 genes with functional annotations, 282 436 992 unique variants from 2048 WGS samples, 88 transcriptomic profiles from 1997 RNA-Seq samples and 13 381 germplasm items. Additionally, TCOD not only employs genes as a bridge to interconnect multi-omics data, enabling cross-species comparisons based on homology relationships, but also offers user-friendly online tools for efficient data mining and visualization. In short, TCOD integrates multi-species, multi-omics data and online tools, which will facilitate the research on genomic selective breeding and trait biology of tropical crops.

Jasmonate signalling in the regulation of rubber biosynthesis in laticifer cells of rubber tree, Hevea brasiliensis.

Rubber trees are the world's major source of natural rubber. Rubber-containing latex is obtained from the laticifer cells of the rubber tree (Hevea brasiliensis) via regular tapping. Rubber biosynthesis is a typical isoprenoid metabolic process in the laticifer cells; however, little is known about the positive feedback regulation caused by the loss of latex that occurs through tapping. In this study, we demonstrate the crucial role of jasmonate signalling in this feedback regulation. The endogenous levels of jasmonate, the expression levels of rubber biosynthesis-related genes, and the efficiency of in vitro rubber biosynthesis were found to be significantly higher in laticifer cells of regularly tapped trees than those of virgin (i.e. untapped) trees. Application of methyl jasmonate had similar effects to latex harvesting in up-regulating the rubber biosynthesis-related genes and enhancing rubber biosynthesis. The specific jasmonate signalling module in laticifer cells was identified as COI1-JAZ3-MYC2. Its activation was associated with enhanced rubber biosynthesis via up-regulation of the expression of a farnesyl pyrophosphate synthase gene and a small rubber particle protein gene. The increase in the corresponding proteins, especially that of farnesyl pyrophosphate synthase, probably contributes to the increased efficiency of rubber biosynthesis. To our knowledge, this is the first study to reveal a jasmonate signalling pathway in the regulation of rubber biosynthesis in laticifer cells. The identification of the specific jasmonate signalling module in the laticifer cells of the rubber tree may provide a basis for genetic improvement of rubber yield potential.

Comparative transcriptome analysis of latex from rubber tree clone CATAS8-79 and PR107 reveals new cues for the regulation of latex regeneration and duration of latex flow.

BACKGROUND: Rubber tree (Hevea brasiliensis Muell. Arg.) is the primarily commercial source of natural rubber in the world. Latex regeneration and duration of latex flow after tapping are the two factors that determine rubber yield of rubber tree, and exhibit a huge variation between rubber tree clones CATAS8-79 and PR107. RESULTS: To dissect the molecular mechanism for the regulation of latex regeneration and duration of latex flow, we sequenced and comparatively analyzed latex of rubber tree clone CATAS8-79 and PR107 at transriptome level. More than 26 million clean reads were generated in each pool and 51,829 all-unigenes were totally assembled. A total of 6,726 unigenes with differential expression patterns were detected between CATAS8-79 and PR107. Functional analysis showed that genes related to mass of categories were differentially enriched between the two clones. Expression pattern of genes which were involved in latex regeneration and duration of latex flow upon successive tapping was analyzed by quantitative PCR. Several genes related to rubber biosynthesis, cellulose and lignin biosynthesis and rubber particle aggregation were differentially expressed between CATAS8-79 and PR107. CONCLUSIONS: This is the first report about probing latex regeneration and duration of latex flow by comparative transcriptome analysis. Among all the suggested factors, it is more important that the level of endogenous jasmonates, carbohydrate metabolism, hydroxymethylglutaryl-CoA reductase (HMGR) and Hevea rubber transferase (HRT) in mevalonate (MVA) parthway for latex regeneration while the level of endogenous ethylene (ETH), lignin content of laticifer cell wall, antioxidants and glucanases for the duration of latex flow. These data will provide new cues for understanding the molecular mechanism for the regulation of latex regeneration and duration of latex flow in rubber tree.

Genomic insight into domestication of rubber tree.

Understanding the genetic basis of rubber tree (Hevea brasiliensis) domestication is crucial for further improving natural rubber production to meet its increasing demand worldwide. Here we provide a high-quality H. brasiliensis genome assembly (1.58 Gb, contig N50 of 11.21 megabases), present a map of genome variations by resequencing 335 accessions and reveal domestication-related molecular signals and a major domestication trait, the higher number of laticifer rings. We further show that HbPSK5, encoding the small-peptide hormone phytosulfokine (PSK), is a key domestication gene and closely correlated with the major domestication trait. The transcriptional activation of HbPSK5 by myelocytomatosis (MYC) members links PSK signaling to jasmonates in regulating the laticifer differentiation in rubber tree. Heterologous overexpression of HbPSK5 in Russian dandelion (Taraxacum kok-saghyz) can increase rubber content by promoting laticifer formation. Our results provide an insight into target genes for improving rubber tree and accelerating the domestication of other rubber-producing plants.

Genome-wide identification of the rubber tree superoxide dismutase (SOD) gene family and analysis of its expression under abiotic stress.

Background: The rubber tree (Hevea brasiliensis) is the only species capable of producing high-quality natural rubber for commercial use, and is often subjected to various abiotic stresses in non-traditional rubber plantation areas. Superoxide dismutase (SOD) is a vital metalloenzyme translated by a SOD gene family member and acts as a first-line of protection in plant cells by catalysing the disproportionation of reactive oxygen species (ROS) to produce H2O2 and O2. However, the SOD gene family is not reported in rubber trees. Methods: Here, we used hidden markov model (HMM) and BLASTP methods to identify SOD genes in the H. brasiliensis genome. Phylogenetic tree, conserved motifs, gene structures, cis elements, and gene ontology annotation (GO) analyses were performed using MEGA 6.0, MEME, TBtools, PlantCARE, and eggNOG database, respectively. HbSOD gene expression profiles were analysed using quantitative reverse transcription polymerase chain reaction (qRT-PCR). Results: We identified nine HbSOD genes in the rubber tree genome, including five HbCSDs, two HbFSDs, and two HbMSDs. Phylogenetic relationship analysis classified the SOD proteins from the rubber tree and other related species into three subfamilies. The results of gene structure and conserved motif analysis illustrated that most HbSOD genes have similar exon-intron numbers and conserved motifs in the same evolutionary branch. Five hormone-related, four stress-related, and light-responsive elements were detected in the HbSODs' promoters. HbSODs were expressed in different tissues, gradually increased with leaf development, and were abundantly expressed in mature leaves. HbCSD2 and HbCSD4 was significantly upregulated under low and high temperatures, and salt stress, except for HbCSD2, by heat. Furthermore, most HbSOD genes were significantly upregulated by drought, except HbMSD2. These findings imply that these genes may play vital roles in rubber tree stress resistance. Our results provide a basis for further studies on the functions of HbSOD genes in rubber trees and stress response mechanisms.

Genome-wide identification and expression analysis of the phosphatase 2A family in rubber tree (Hevea brasiliensis).

The protein phosphatase 2As (PP2As) play a key role in manipulating protein phosphorylation. Although a number of proteins in the latex of laticifers are phosphorylated during latex regeneration in rubber tree, information about the PP2A family is limited. In the present study, 36 members of the HbPP2A family were genome-wide identified. They were clustered into five subgroups: the subgroup HbPP2AA (4), HbPP2AB' (14), HbPP2AB'' (6), HbPP2AB55 (4), and HbPP2AC (8). The members within the same subgroup shared highly conserved gene structures and protein motifs. Most of HbPP2As possessed ethylene- and wounding-responsive cis-acting elements. The transcripts of 29 genes could be detected in latex by using published high-throughput sequencing data. Of the 29 genes, seventeen genes were significantly down-regulated while HbPP2AA1-1 and HbPP2AB55α/Bα-1were up-regulated by tapping. Of the 17 genes, 14 genes were further significantly down-regulated by ethrel application. The down-regulated expression of a large number of HbPP2As may attribute to the enhanced phosphorylation of the proteins in latex from the tapped trees and the trees treated with ethrel application.

Genome-Wide Identification and Characterization of the JAZ Gene Family in Rubber Tree (Hevea brasiliensis).

Jasmonate signaling plays a vital role in the regulation of secondary laticifer differentiation and natural rubber biosynthesis in Hevea brasiliensis. Jasmonate ZIM-domain (JAZ) proteins are the master regulators of jasmonate signaling. Although several JAZs have been reported in the laticifer cells of H. brasiliensis, the genome-wide screening of HbJAZ members has not yet been explored. In the present study, 18 HbJAZs were identified based on the recent H. brasiliensis genome. Phylogenetic construction revealed that the HbJAZs were clustered into five subgroups and that members within the same subgroup shared highly conserved gene structures and protein motifs. Cis-element analysis of HbJAZ promoters suggested the presence of hormone, stress and development-related cis-elements. HbJAZ1.0, HbJAZ2.0, and HbJAZ5.0 interacted with CORONATINE INSENSITIVE1 (COI1) in the presence of coronatine (COR, a JA mimic). HbJAZ1.0, HbJAZ2.0, HbJAZ5.0, and HbJAZ12.0 could also interact with each other. Of the 18 HbJAZs, transcripts of 15 HbJAZs were present in the vascular cambium region except for that of HbJAZ7.0, HbJAZ8.0d, and HbJAZ13.0. Fourteen of the 15 HbJAZs were significantly up-regulated upon COR treatment. The transcripts of three genes that were absent from vascular cambium region were also absent from the latex. Among the 15 HbJAZs in the latex, the expression patterns of 13 HbJAZs were different between the tapping and ethrel treatments. Eight of the 14 COR-up-regulated HbJAZs in the vascular cambium region were also activated by tapping in latex. Of the eight tapping-activated HbJAZs, 5 HbJAZs were repressed by ethrel application. Based on the computational analyses and gene expression patterns described in this study, the HbJAZ5.0 and HbJAZ10.0b may be associated with laticifer differentiation while the HbJAZ8.0b is a negative regulator for natural rubber biosynthesis in H. brasiliensis.

Comparative transcriptome analysis reveals phytohormone signalings, heat shock module and ROS scavenger mediate the cold-tolerance of rubber tree.

Two contrasting cold response rubber tree clones, the cold-resistant '93-114' and cold-sensitive 'Reken501', were subject to a global transcriptome response assessing via high-throughput RNA-seq technique and comprehensive bioinformatics analysis using the referenced rubber tree genome with the purpose of exploring the potential molecular cues underlying the tolerance of rubber trees to cold stress. As a result, a total of 1919 genes had significantly higher expression, while 2929 genes had significantly lower expression in '93-114' than in 'Reken501' without cold stress. Upon cold stress, the numbers of genes with significantly higher expression decreased to 1501 at 1 h treatment and to 1285 at 24 h treatment in '93-114' than that of 'Reken501', conversely, the numbers of genes with significantly lower expression increased to 7567 at 1 h treatment and to 5482 at 24 h treatment. Functional annotation of the differentially expressed genes between '93-114' and 'Reken501' suggests that down-regulation of auxin and ethylene signaling and activation of heat shock module and ROS scavengers is a primary strategy for H. brasiliensis to cope with cold stress. Our identified vital differentially expressed genes may be beneficial for elucidation of the molecular mechanisms underlying cold tolerance and for genetic improvement of H. brasiliensis clones.

Transcript Profiling of Hevea brasiliensis during Latex Flow.

Latex exploitation enhances latex regeneration in rubber trees. The latex exploitation-caused latex flow lasts from 10 min to a few hours, which is convenient for exploring the transcript profiling of latex metabolism-related genes at the different stages of latex flow. In the present study, the expression pattern of 62 latex metabolism-related genes involved in water transportation, carbohydrate metabolism, natural rubber biosynthesis, hormone signaling, ROS generation and scavenging, and latex coagulum across three stages of latex flow between rubber tree clones CATAS7-33-97 and CATAS8-79 were comparatively analyzed by quantitative real-time PCR. The two clones show differences in latex regeneration and have a different duration of latex flow. The results showed that the expression levels of 38 genes were significantly higher in CATAS8-79 latex than in CATAS7-33-97 during latex regeneration, while 45 genes had a notably higher expression level in CATAS8-79 latex during latex flow. Together with the activation of the MEP pathway and jasmonate pathway in CATAS8-79 latex, HbPIP1;3, HbPIP1;4, HbSUT3, HbSus3, HbHMGS1-2, HbMK should contribute to the high latex regeneration ability. The up-regulation of ethylene signaling and Hb44KD and the down-regulation of latex coagulation-related genes in CATAS8-79 latex might contribute to its longer latex flow duration. This study provides some cues for revealing the regulation of latex metabolism in rubber trees.

Evaluation of Reference Genes for Quantitative Real-Time PCR Analysis of the Gene Expression in Laticifers on the Basis of Latex Flow in Rubber Tree (Hevea brasiliensis Muell. Arg.).

Latex exploitation-caused latex flow is effective in enhancing latex regeneration in laticifer cells of rubber tree. It should be suitable for screening appropriate reference gene for analysis of the expression of latex regeneration-related genes by quantitative real-time PCR (qRT-PCR). In the present study, the expression stability of 23 candidate reference genes was evaluated on the basis of latex flow by using geNorm and NormFinder algorithms. Ubiquitin-protein ligase 2a (UBC2a) and ubiquitin-protein ligase 2b (UBC2b) were the two most stable genes among the selected candidate references in rubber tree clones with differential duration of latex flow. The two genes were also high-ranked in previous reference gene screening across different tissues and experimental conditions. By contrast, the transcripts of latex regeneration-related genes fluctuated significantly during latex flow. The results suggest that screening reference gene during latex flow should be an efficient and effective clue for selection of reference genes in qRT-PCR.

Transcriptome Analysis of the Signalling Networks in Coronatine-Induced Secondary Laticifer Differentiation from Vascular Cambia in Rubber Trees.

The secondary laticifer in rubber tree (Hevea brasiliensis Muell. Arg.) is a specific tissue within the secondary phloem. This tissue differentiates from the vascular cambia, and its function is natural rubber biosynthesis and storage. Given that jasmonates play a pivotal role in secondary laticifer differentiation, we established an experimental system with jasmonate (JA) mimic coronatine (COR) for studying the secondary laticifer differentiation: in this system, differentiation occurs within five days of the treatment of epicormic shoots with COR. In the present study, the experimental system was used to perform transcriptome sequencing and gene expression analysis. A total of 67,873 unigenes were assembled, and 50,548 unigenes were mapped at least in one public database. Of these being annotated unigenes, 15,780 unigenes were differentially expressed early after COR treatment, and 19,824 unigenes were differentially expressed late after COR treatment. At the early stage, 8,646 unigenes were up-regulated, while 7,134 unigenes were down-regulated. At the late stage, the numbers of up- and down-regulated unigenes were 7,711 and 12,113, respectively. The annotation data and gene expression analysis of the differentially expressed unigenes suggest that JA-mediated signalling, Ca2+ signal transduction and the CLAVATA-MAPK-WOX signalling pathway may be involved in regulating secondary laticifer differentiation in rubber trees.

Gene expression analysis and SNP/InDel discovery to investigate yield heterosis of two rubber tree F1 hybrids.

As an important industrial material, natural rubber is mainly harvested from the rubber tree. Rubber tree breeding is inefficient, expensive and time-consuming, whereas marker-assisted selection is a feasible method for early selection of high-yield hybrids. We thus sequenced and analyzed the transcriptomes of two parent rubber trees (RRIM 600 and PR 107) and their most productive hybrids (RY 7-33-97 and RY 7-20-59) to understand their gene expression patterns and genetic variations including single nucleotide polymorphisms (SNPs) and small insertions/deletions (InDels). We discovered >31,000 genetic variations in 112,702 assembled unigenes. Our results showed that the higher yield in F1 hybrids was positively associated with their higher genome heterozygosity, which was further confirmed by genotyping 10 SNPs in 20 other varieties. We also showed that RY 7-33-97 and RY 7-20-59 were genetically closer to RRIM 600 and PR 107, respectively, in agreement with both their phenotypic similarities and gene expression profiles. After identifying ethylene- and jasmonic acid-responsive genes at the transcription level, we compared and analyzed the genetic variations underlying rubber biosynthesis and the jasmonic acid and ethylene pathways in detail. Our results suggest that genome-wide genetic variations play a substantive role in maintaining rubber tree heterosis.

Cloning, heterologous expression and characterization of ascorbate peroxidase (APX) gene in laticifer cells of rubber tree (Hevea brasiliensis Muell. Arg.).

Ascorbate peroxidases (APXs) are a kind of crucial enzymes for removing reactive oxygen species (ROS) in plant cell. In the present study, a full-length cDNA encoding an APX, designated HbAPX, was isolated from Hevea brasiliensis by the rapid amplification of cDNA ends (RACE) method. HbAPX was 1174-bp in length and contained a 912-bp open reading frame (ORF) encoding a putative protein of 304 amino acids. The predicted molecular mass of HbAPX was 27.6 kDa (kDa) with an isoelectric point (pI) of 6.73. The phylogenetic analysis showed that HbAPX belonged to the cytosolic subgroup and was more relative to PtAPX and MdAPX2. By using PlantCare online analysis, such cis-acting elements as W-box and MRE were detected in the promoter region of HbAPX. Overproduction of recombinant HbAPX protein either in Escherichia coli or yeast enhanced their tolerance to such abiotic stresses as Cu(2+), Zn(2+), Na(2+) and hydrogen peroxide (H2O2). Ethrel application significantly down-regulated the expression of HbAPX and inhibited the activity of HbAPX in vivo. The ethrel-caused down-regulation of HbAPX may disturb the redox homeostasis in laticifer cells of rubber tree.

Cytological and transcriptional dynamics analysis of host plant revealed stage-specific biological processes related to compatible rice-Ustilaginoidea virens interaction.

Rice false smut, a fungal disease caused by Ustilaginoidea virens is becoming a severe detriment to rice production worldwide. However, little is known about the molecular response of rice to attacks by the smut pathogen. In this article, we define the initial infection process as having three stages: initial colonization on the pistil (stage 1, S1), amplification on the anther (stage 2, S2) and sporulation in the anther chambers (stage 3, S3). Based on the transcriptome of rice hosts in response to U. virens in two separate years, we identified 126, 204, and 580 specific regulated genes in their respective stages S1, S2, and S3, respectively, by excluding common expression patterns in other openly biotic/abiotic databases using bioinformatics. As the disease progresses, several stage-specific biological processes (BP) terms were distinctively enriched: "Phosphorylation" in stage S1, "PCD" in S2, and "Cell wall biogenesis" in S3, implying a concise signal cascade indicative of the tactics that smut pathogens use to control host rice cells during infection. 113 regulated genes were coexpressed among the three stages. They shared highly conserved promoter cis-element in the promoters in response to the regulation of WRKY and Myb for up-regulation, and ABA and Ca2+ for down regulation, indicating their potentially critical roles in signal transduction during rice-U. virens interaction. We further analyzed seven highly regulated unique genes; four were specific to pollen development, implying that pollen-related genes play critical roles in the establishment of rice susceptibility to U. virens. To my knowledge, this is the first report about probing of molecular response of rice to smut pathogen infection, which will greatly expand our understanding of the molecular events surrounding infection by rice false smut.