bHLH Family Research Articles

Genomic Characterization and Expression Analysis of Basic Helix-Loop-Helix (bHLH) Family Genes in Traditional Chinese Herb Dendrobium officinale.

Dendrobium officinale Kimura et Migo is of great importance as a traditional Chinese herb due to its abundant metabolites. The family of basic helix-loop-helix (bHLH) transcription factors widely exists in plants and plays an essential role in plant growth and development, secondary metabolism as well as responses to environmental changes. However, there is limited information on bHLH genes in D. officinale. In the present study, a total of 98 putative DobHLH genes were identified at the genomic level, which could be classified into 18 clades. Gene structures and conserved motifs in DobHLH genes showed high conservation during their evolution. The conserved amino acids and DNA bindings of DobHLH proteins were predicted, both of which are pivotal for their function. Furthermore, gene expression from eight tissues showed that some DobHLH genes were ubiquitously expressed while other DobHLH genes were expressed in the specific tissues. Expressional changes of DobHLH genes under MeJA and ABA treatments were detected by qRT-PCR. The protein–protein interactions between DobHLHs were predicted and several interactions were confirmed by yeast two hybrid. Therefore, our results here contribute to the understanding of bHLH genes in D. officinale and lay a foundation for the further functional study of its biological processes.

Genome-wide identification and characterization of bHLH family genes from Ginkgo biloba

Basic helix–loop–helix (bHLH) proteins, one of the most important and largest transcription factor family in plants, play important roles in regulating growth and development, stress response. In recent years, many bHLH family genes have been identified and characterized in woody plants. However, a systematic analysis of the bHLH gene family has not been reported in Ginkgo biloba, the oldest relic plant species. In this study, we identifed a total of 85 GbbHLH genes from the genomic and transcriptomic databases of G. biloba, which were classified into 17 subfamilies based on the phylogenetic analysis. Gene structures analysis indicated that the number of exon–intron range in GbbHLHs from 0 to 12. The MEME analysis showed that two conserved motifs, motif 1 and motif 2, distributed in most GbbHLH protein. Subcellular localization analysis exhibited that most GbbHLHs located in nucleus and a few GbbHLHs were distributed in chloroplast, plasma membrane and peroxisome. Promoter cis-element analysis revealed that most of the GbbHLH genes contained abundant cis-elements that involved in plant growth and development, secondary metabolism biosynthesis, various abiotic stresses response. In addition, correlation analysis between gene expression and flavonoid content screened seven candidate GbbHLH genes involved in flavonoid biosynthesis, providing the targeted gene encoding transcript factor for increase the flavonoid production through genetic engineering in G. biloba.

Genome-Wide Analysis Reveals Transcription Factors Regulated by Spider-Mite Feeding in Cucumber (Cucumis sativus).

To gain insight into the regulatory networks that underlie the induced defense in cucumber against spider mites, genes encoding transcription factors (TFs) were identified in the cucumber (Cucumis sativus) genome and their regulation by two-spotted spider mite (Tetranychus urticae) herbivory was analyzed using RNA-seq. Of the total 1212 annotated TF genes in the cucumber genome, 119 were differentially regulated upon spider-mite herbivory during a period of 3 days. These TF genes belong to different categories but the MYB, bHLH, AP2/ERF and WRKY families had the highest relative numbers of differentially expressed genes. Correlation analysis of the expression of TF genes with defense-associated genes during herbivory and pathogen infestation, and in different organs resulted in the putative identification of regulators of herbivore-induced terpenoid and green-leaf-volatile biosynthesis. Analysis of the cis-acting regulatory elements (CAREs) present in the promoter regions of the genes responsive to spider-mite feeding revealed potential TF regulators. This study describes the TF genes in cucumber that are potentially involved in the regulation of induced defense against herbivory by spider mites.

RETRACTED ARTICLE: Contribution of anthocyanin pathways to fruit flesh coloration in pitayas

BackgroundColor formation in Hylocereus spp. (pitayas) has been ascribed to the accumulation of betalains. However, several studies have reported the presence of anthocyanins in pitaya fruit and their potential role in color formation has not yet been explored. In this study, we profiled metabolome and transcriptome in fruit of three cultivars with contrasting flesh colors (red, pink and white) to investigate their nutritional quality and the mechanism of color formation involving anthocyanins.ResultsResults revealed that pitaya fruit is enriched in amino acid, lipid, carbohydrate, polyphenols, vitamin and other bioactive components with significant variation among the three cultivars. Anthocyanins were detected in the fruit flesh and accumulation levels of Cyanidin 3-glucoside, Cyanidin 3-rutinoside, Delphinidin 3-O-(6-O-malonyl)-beta-glucoside-3-O-beta-glucoside and Delphinidin 3-O-beta-D-glucoside 5-O-(6-coumaroyl-beta-D-glucoside) positively correlated with the reddish coloration. Transcriptome data showed that the white cultivar tends to repress the anthocyanin biosynthetic pathway and divert substrates to other competing pathways. This perfectly contrasted with observations in the red cultivar. The pink cultivar however seems to keep a balance between the anthocyanin biosynthetic pathway and the competing pathways. We identified several active transcription factors of the MYB and bHLH families which can be further investigated as potential regulators of the anthocyanin biosynthetic genes.ConclusionsCollectively, our results suggest that anthocyanins partly contribute to color formation in pitaya fruit. Future studies aiming at manipulating the biosynthetic pathways of anthocyanins and betalains will better clarify the exact contribution of each pathway in color formation in pitayas. This will facilitate efforts to improve pitaya fruit quality and appeal.

The bHLH protein OsIRO3 is critical for plant survival and iron (Fe) homeostasis in rice (Oryza sativa L.) under Fe-deficient conditions

ABSTRACT Iron (Fe) is an essential element for most organisms since it is involved in various biological reactions. Fe deficiency is a major constraint for plant growth and leads to low crop yields and poor nutritional quality. Fe uptake and translocation in plants require the coordinated expression of multiple genes, which is regulated by various transcription factors (TFs), especially members of the basic helix-loop-helix (bHLH) family. OsIRO3 was first reported as a repressor related to Fe homeostasis gene regulation in roots based on evidence in overexpressing rice. However, its functions in regulating rice growth and Fe homeostasis between roots and shoots during Fe deficiency is poorly understood. Here, we developed osiro3 knockout mutants by the CRISPR/Cas9 system. The leaves of osiro3 knockout mutants showed abnormal growth and development after Fe-deficiency treatment. Strong lesions occurred on the young leaves of osiro3 knockout mutants, which even led to plant death during severe Fe deficiency. Furthermore, accumulation of reactive oxygen species (ROS), Fe and manganese concentrations were all elevated in the shoots of osiro3 knockout mutants under the Fe deficiency stress. Quantitative RT-PCR showed that OsNAC4 for ROS induced plant hypersensitive cell death and OsFerritins for Fe storage and overload were all significantly activated in osiro3 knockout shoots after Fe deficiency treatment. In osiro3 knockout mutants, Fe-deficiency response genes were induced in roots, but suppressed in shoots, and furthermore Fe sufficiency/overload genes were induced in shoots under Fe-deficient conditions according to DNA microarray. Yeast two-hybrid analysis revealed a positive interaction between OsIRO3 and the homologous protein OsbHLH062, which was also transcriptionally increased in Fe-deficient roots. OsIRO3 is essential for maintaining plant survival in rice under Fe-deficient conditions and plays an important role in signal transmission from shoots to roots which is critical for plants to prevent Fe overload.

Genome-wide analysis of basic helix-loop-helix transcription factors in papaya (Carica papaya L.).

The basic helix-loop-helix (bHLH) transcription factors (TFs) have been identified and functionally characterized in many plants. However, no comprehensive analysis of the bHLH family in papaya (Carica papaya L.) has been reported previously. Here, a total of 73 CpbHLHs were identified in papaya, and these genes were classified into 18 subfamilies based on phylogenetic analysis. Almost all of the CpbHLHs in the same subfamily shared similar gene structures and protein motifs according to analysis of exon/intron organizations and motif compositions. The number of exons in CpbHLHs varied from one to 10 with an average of five. The amino acid sequences of the bHLH domains were quite conservative, especially Leu-27 and Leu-63. Promoter cis-element analysis revealed that most of the CpbHLHs contained cis-elements that can respond to various biotic/abiotic stress-related events. Gene ontology (GO) analysis revealed that CpbHLHs mainly functions in protein dimerization activity and DNA-binding, and most CpbHLHs were predicted to localize in the nucleus. Abiotic stress treatment and quantitative real-time PCR (qRT-PCR) revealed some important candidate CpbHLHs that might be responsible for abiotic stress responses in papaya. These findings would lay a foundation for further investigate of the molecular functions of CpbHLHs.

SmbHLH53 is relevant to jasmonate signaling and plays dual roles in regulating the genes for enzymes in the pathway for salvianolic acid B biosynthesis in Salvia miltiorrhiza.

Basic helix-loop-helix (bHLH) transcription factors play essential roles in myriad regulatory processes, including secondary metabolism. In this study with Salvia miltiorrhiza, we isolated and characterized SmbHLH53, which encodes a bHLH family member. Expression of this gene was significantly induced by wounding and multiple hormones, including methyl jasmonic acid; transcript levels were highest in the leaves and roots. Phylogenetic analysis indicated that SmbHLH53 clusters withAtbHLH17 and AtbHLH13, two negative regulators of jasmonate (JA) responses, and is localized in the nucleus and cell membrane. Yeast two-hybrid and bimolecular fluorescent complementation assays indicated that SmbHLH53 forms a homodimer as well as a heterodimer with SmbHLH37. It also interacts with both SmJAZs1/3/8 and SmMYC2, the core members of the JA signal pathway. Unexpectedly, we noted that overexpression of SmbHLH53 did not significantly influence the concentrations of rosmarinic acid and salvianolic acid B in transgenic plants. Results from yeast one-hybrid assays showed that SmbHLH53 binds to the promoters of SmTAT1, SmPAL1, and Sm4CL9, the key genes for enzymes in the pathway for phenolic acid synthesis. Assays of transient transcriptional activity demonstrated that SmbHLH53 represses the promoter of SmTAT1 while activating the promoter of Sm4CL9. Thus, the present work revealed that SmbHLH53 may play dual roles in regulating the genes for enzymes in the pathway for Sal B biosynthesis.

Ldb1 is required for Lmo2 oncogene–induced thymocyte self-renewal and T-cell acute lymphoblastic leukemia

AbstractProlonged or enhanced expression of the proto-oncogene Lmo2 is associated with a severe form of T-cell acute lymphoblastic leukemia (T-ALL), designated early T-cell precursor ALL, which is characterized by the aberrant self-renewal and subsequent oncogenic transformation of immature thymocytes. It has been suggested that Lmo2 exerts these effects by functioning as component of a multi-subunit transcription complex that includes the ubiquitous adapter Ldb1 along with b-HLH and/or GATA family transcription factors; however, direct experimental evidence for this mechanism is lacking. In this study, we investigated the importance of Ldb1 for Lmo2-induced T-ALL by conditional deletion of Ldb1 in thymocytes in an Lmo2 transgenic mouse model of T-ALL. Our results identify a critical requirement for Ldb1 in Lmo2-induced thymocyte self-renewal and thymocyte radiation resistance and for the transition of preleukemic thymocytes to overt T-ALL. Moreover, Ldb1 was also required for acquisition of the aberrant preleukemic ETP gene expression signature in immature Lmo2 transgenic thymocytes. Co-binding of Ldb1 and Lmo2 was detected at the promoters of key upregulated T-ALL driver genes (Hhex, Lyl1, and Nfe2) in preleukemic Lmo2 transgenic thymocytes, and binding of both Ldb1 and Lmo2 at these sites was reduced following Cre-mediated deletion of Ldb1. Together, these results identify a key role for Ldb1, a nonproto-oncogene, in T-ALL and support a model in which Lmo2-induced T-ALL results from failure to downregulate Ldb1/Lmo2-nucleated transcription complexes which normally function to enforce self-renewal in bone marrow hematopoietic progenitors.

Characterization and Analysis of the Full-Length Transcriptomes of Multiple Organs in Pseudotaxus chienii (W.C.Cheng) W.C.Cheng.

Pseudotaxus chienii, a rare tertiary relict species with economic and ecological value, is a representative of the monotypic genus Pseudotaxus that is endemic to China. P. chienii can adapt well to habitat isolation and ecological heterogeneity under a variety of climate and soil conditions, and is able to survive in harsh environments. However, little is known about the molecular and genetic resources of this long-lived conifer. Herein, we sequenced the transcriptomes of four organs of P. chienii using the PacBio Isoform Sequencing and Illumina RNA Sequencing platforms. Based on the PacBio Iso-Seq data, we obtained 44,896, 58,082, 50,485, and 67,638 full-length unigenes from the root, stem, leaf, and strobilus, respectively, with a mean length of 2692 bp, and a mean N50 length of 3010.75 bp. We then comprehensively annotated these unigenes. The number of organ-specific expressed unigenes ranged from 4393 in leaf to 9124 in strobilus, suggesting their special roles in physiological processes, organ development, and adaptability in the different four organs. A total of 16,562 differentially expressed genes (DEGs) were identified among the four organs and clustered into six subclusters. The gene families related to biotic/abiotic factors, including the TPS, CYP450, and HSP families, were characterized. The expression levels of most DEGs in the phenylpropanoid biosynthesis pathway and plant–pathogen interactions were higher in the root than in the three other organs, suggesting that root constitutes the main organ of defensive compound synthesis and accumulation and has a stronger ability to respond to stress. The sequences were analyzed to predict transcription factors, long non-coding RNAs, and alternative splicing events. The expression levels of most DEGs of C2H2, C3H, bHLH, and bZIP families in the root and stem were higher than those in the leaf and strobilus, indicating that these TFs may play a crucial role in the survival of the root and stem. These results comprise the first comprehensive gene expression profiles obtained for different organs of P. chienii. Our findings will facilitate further studies on the functional genomics, adaptive evolution, and phylogeny of P. chienii, and lay the foundation for the development of conservation strategies for this endangered conifer.

PsbHLH1, a novel transcription factor involved in regulating anthocyanin biosynthesis in tree peony (Paeonia suffruticosa)

Flower color is one of the most important features of ornamental plants. Anthocyanin composition and concentration are usually closely related to flower color formation. The biosynthesis of anthocyanin is regulated by a series of structural genes and regulatory genes. The basic helix-loop-helix proteins (bHLHs) are considered as one of the key transcription factors known as the regulators of anthocyanin biosynthesis. However, the bHLH transcription factor family of tree peony (Paeonia suffruticosa) has not been systematically studied in previous studies, especially for the regulation of petal pigmentation. The aim of this study was to identify bHLH genes and unravel their underlying molecular mechanism involved in the regulation of anthocyanin biosynthesis in tree peony. Based on transcriptome profiling analysis, we identified three bHLHs candidate anthocyanin regulators, PsbHLH1, PsbHLH2, and PsbHLH3. PsbHLH1-3 were phylogenetically clustered in the IIIf bHLH subgroup, which is involved in anthocyanin biosynthesis in other plant species. In addition, three bHLH proteins were localized in the nucleus and displayed transcriptional activation activity in a yeast hybrid system. Through a series of functional experiments, we further demonstrated that PsbHLH1 could transcriptionally activate the expression of PsDFR and PsANS via directly binding to their promoters. These results laid a solid foundation to better understand the regulatory mechanisms of anthocyanin biosynthesis in P. suffruticosa and to benefit molecular breeding of tree peony cultivars with novel color.

Genome-wide identification and characterization of cucumber bHLH family genes and the functional characterization of CsbHLH041 in NaCl and ABA tolerance in Arabidopsis and cucumber

BackgroundThe basic/helix-loop-helix (bHLH) transcription factor family exists in all three eukaryotic kingdoms as important participants in biological growth and development. To date, the comprehensive genomic and functional analyses of bHLH genes has not been reported in cucumber (Cucumis sativus L.).ResultsHere, a total of 142 bHLH genes were identified and classified into 32 subfamilies according to the conserved motifs, phylogenetic analysis and gene structures in cucumber. The sequences of CsbHLH proteins were highly conserved based on the results of multiple sequence alignment analyses. The chromosomal distribution, synteny analysis, and gene duplications of these 142 CsbHLHs were further analysed. Many elements related to stress responsiveness and plant hormones were present in the promoter regions of CsbHLH genes based on a cis-element analysis. By comparing the phylogeny of cucumber and Arabidopsis bHLH proteins, we found that cucumber bHLH proteins were clustered into different functional clades of Arabidopsis bHLH proteins. The expression analysis of selected CsbHLHs under abiotic stresses (NaCl, ABA and low-temperature treatments) identified five CsbHLH genes that could simultaneously respond to the three abiotic stresses. Tissue-specific expression profiles of these five genes were also analysed. In addition, 35S:CsbHLH041 enhanced the tolerance to salt and ABA in transgenic Arabidopsis and in cucumber seedlings, suggesting CsbHLH041 is an important regulator in response to abiotic stresses. Lastly, the functional interoperability network among the CsbHLH proteins was analysed.ConclusionThis study provided a good foundation for further research into the functions and regulatory mechanisms of CsbHLH proteins and identified candidate genes for stress resistance in cucumber.

Association mapping for yield traits in Paeonia rockii based on SSR markers within transcription factors of comparative transcriptome

BackgroundAllelic variation underlying the quantitative traits in plants is caused by the extremely complex regulation process. Tree peony originated in China is a peculiar ornamental, medicinal and oil woody plant. Paeonia rockii, one of tree peony species, is a precious emerging woody oil crop. However, in this valuable plant, the study of functional loci associated with yield traits has rarely been identified. Therefore, to explore the genetic architecture of 24 yield quantitative traits, the association mapping was first reported in 420 unrelated cultivated P. rockii individuals based on the next-generation sequencing (NGS) and single-molecule long-read sequencing (SMLRS).ResultsThe developed 58 pairs of polymorphic expressed sequence tag-simple sequence repeat (EST-SSR) markers from 959 candidate transcription factors (TFs) associated with yield were used for genotyping the 420 P. rockii accessions. We observed a high level of genetic diversity (polymorphic information content, PIC = 0.514) and low linkage disequilibrium (LD) between EST-SSRs. Moreover, four subpopulations in the association population were revealed by STRUCTURE analyses. Further, single-marker association analysis identified 141 significant associations, involving 17 quantitative traits and 41 EST-SSRs. These loci were mainly from AP2, TCP, MYB, HSF, bHLH, GATA, and B3 gene families and showed a small proportion of the phenotypic variance (3.79 to 37.45%).ConclusionsOur results summarize a valuable collection of functional loci associated with yield traits in P. rockii, and provide a precious resource that reveals allelic variation underlying quantitative traits in Paeonia and other woody oil crops.

Comparative transcriptome analysis reveals the responses of winter rye to cold stress

Rye is an important economic crop with remarkable resistance to biotic and abiotic stresses and high nutrient utilization. Identification of genes related to cold stress response in winter rye to improve understanding of the molecular mechanism of low temperature response. We performed RNA-seq on winter rye transcriptome under four different cold stress conditions: CK (room temperature), 24 h (4 °C), 48 h (4 °C) and 72 h (4 °C). A total of 128,744 Unigenes were identified, of which 29,874 genes were differentially expressed, including genes related to photosynthesis, plasma membrane stability, glucose and energy metabolism, as well as cold-response transcription factors. GO and KEGG results showed that winter rye protects against low temperature damage by synthesizing extracellular components such as Cutin, suberine and wax. In addition, Oligosaccharides and chitin play a key role in cold resistance of winter rye and MNS1 and MNS3 were the candidate genes. The transcription factors (TFs) results showed that MYB and bHLH families showed a close relationship with cold resistance. In conclusion, the cold tolerance of winter rye is achieved through a variety of biological pathways, including “chitin binding”, “glycolytic process”, and “tricarboxylic acid cycle”. Winter rye can also respond to cold stress by regulating cutin, suberine and wax synthesis. The expression of common cold stress genes such as HSPs, MYBs, bHLHs, NAC, bZIPs, C2H2 and CBF changed after cold stimulation. This study can provide a valuable resource for studies related to cold tolerance, helping to explore genes associated with cold stress and for genetic improvement of crops.

Identification of the bHLH gene family in Dracaena cambodiana reveals candidate genes involved in flavonoid biosynthesis

The basic helix-loop-helix (bHLH) transcription factors (TFs) regulate multiple biological processes by altering the expression of target genes. However, the bHLH superfamily genes in the dragon tree (Dracaena cambodiana Pierre ex Gagnep) have not been characterized, and their role in flavonoid biosynthesis remain unclear. Here, 62 bHLH genes in D. cambodiana were identified, and these genes were classified into 19 subfamilies based on a phylogenetic comparison with proteins from Arabidopsis. Transcriptomic analysis revealed that 8 DcbHLHs were up-regulated when exposed to the inducer of dragon’s blood and ultraviolet radiation b (UVB). Interaction network analysis revealed that 3 bHLHs (DcbHLH2, DcbHLH5 and DcbHLH56) are likely involved in flavonoid biosynthesis. In addition, DcbHLH5 and flavonoids biosynthetic genes were co-expressed in response to the inducer and UVB exposure. Furthermore, DcbHLH5 could bind to the promoters of flavonoid biosynthesis genes and activate their expression. This study suggests that DcbHLH proteins maybe involved in flavonoid biosynthesis.

The bHLH gene family and its response to saline stress in Jilin ginseng, Panax ginseng C.A. Meyer.

Basic helix-loop-helix (bHLH) gene family is a gene family of transcription factors that plays essential roles in plantgrowth and development, secondary metabolism and response to biotic and abiotic stresses. Therefore, a comprehensive knowledge of the bHLH gene family is paramount to understand the molecular mechanisms underlying these processes and develop advanced technologies to manipulate the processes efficiently. Ginseng, Panax ginseng C.A. Meyer, is a well-known medicinal herb; however, little is known about the bHLH genes (PgbHLH) in the species. Here, we identified 137 PgbHLH genes from Jilin ginseng cultivar, Damaya, widely cultivated in Jilin, China, of which 50 are newly identified by pan-genome analysis. These 137 PgbHLH genes were phylogenetically classified into 26 subfamilies, suggesting their sequence diversification. They are alternatively spliced into 366 transcripts in a 4-year-old plant and involved in 11 functional subcategories of the gene ontology, indicating their functional differentiation in ginseng. The expressions of the PgbHLH genes dramatically vary spatio-temporally and across 42 genotypes, but they are still somehow functionally correlated. Moreover, the PgbHLH gene family, at least some of its genes, is shown to have roles in plant response to the abiotic stress of saline. These results provide a new insight into the evolution and functional differentiation of the bHLH gene family in plants, new bHLH genes to the PgbHLH gene family, and saline stress-responsive genes for genetic improvement in ginseng and other plant species.

Genome-wide transcriptome analysis of roots in two rice varieties in response to alternate wetting and drying irrigation

Alternate wetting and drying (AWD) irrigation has been widely used as an efficient rice production method to obtain better yield without continuous flooding (CF) of the paddy field. However, how this practice affects gene expression to regulate rice physiology and morphology is largely unknown. In this study, we used two rice varieties, Nipponbare, a lowland rice cultivar, and Gaoshan 1, an upland cultivar, and found that root dry weight (RDW) and root oxidation activity (ROA) in both cultivars substantially increased in response to AWD. We then analyzed the differences in transcriptome profiles of their roots irrigated in AWD vs. CF conditions. AWD responsive genes are mainly involved in lignin biosynthetic pathway and phytohormone signal transduction pathway and belong mainly to bHLH, bZIP, NAC, WRKY, and HSF transcription factor families. We discussed how these differentially expressed genes may contribute to the morphological adaptations observed in roots exposed to AWD. This analysis also provides useful information to explain the similarities and differences in adaptation to AWD irrigation between the two rice ecotypes.

Transcriptome Analysis Reveals Clues into leaf-like flower mutant in Chinese orchid Cymbidium ensifolium

The floral morphology of Cymbidium ensifolium, a well-known orchid in China, has increasingly attracted horticultural and commercial attention. However, the molecular mechanisms that regulate flower development defects in C. ensifolium mutants are poorly understood. In this work, we examined a domesticated variety of C. ensifolium named ‘CuiYuMuDan’, or leaf-like flower mutant, which lacks typical characteristics of orchid floral organs but continues to produce sepal-to leaf-like structures along the inflorescence. We used comparative transcriptome analysis to identify 6234 genes that are differentially expressed between mutant and wild-type flowers. The majority of these differentially expressed genes are involved in membrane-building, anabolism regulation, and plant hormone signal transduction, implying that in the leaf-like mutant these processes play roles in the development of flower defects. In addition, we identified 152 differentially expressed transcription factors, including the bHLH, MYB, MIKC, and WRKY gene families. Moreover, we found 20 differentially expressed genes that are commonly involved in flower development, including MADS-box genes, CLAVATA3 (CLV3), WUSCHEL (WUS), and PERIANTHIA (PAN). Among them, floral homeotic genes were further investigated by phylogenetic analysis and expression validation, which displayed distinctive spatial expression patterns and significant changes between the wild type and the mutant. This is the first report on the C. ensifolium leaf-like flower mutant transcriptome. Our results shed light on the molecular regulation of orchid flower development, and may improve our understanding of floral patterning regulation and advance molecular breeding of Chinese orchids.

Physiological and transcriptomic analyses reveal the mechanisms underlying the salt tolerance of Zoysia japonica Steud

BackgroundAreas with saline soils are sparsely populated and have fragile ecosystems, which severely restricts the sustainable development of local economies. Zoysia grasses are recognized as excellent warm-season turfgrasses worldwide, with high salt tolerance and superior growth in saline-alkali soils. However, the mechanism underlying the salt tolerance of Zoysia species remains unknown.ResultsThe phenotypic and physiological responses of two contrasting materials, Zoysia japonica Steud. Z004 (salt sensitive) and Z011 (salt tolerant) in response to salt stress were studied. The results show that Z011 was more salt tolerant than was Z004, with the former presenting greater K+/Na+ ratios in both its leaves and roots. To study the molecular mechanisms underlying salt tolerance further, we compared the transcriptomes of the two materials at different time points (0 h, 1 h, 24 h, and 72 h) and from different tissues (leaves and roots) under salt treatment. The 24-h time point and the roots might make significant contributions to the salt tolerance. Moreover, GO and KEGG analyses of different comparisons revealed that the key DEGs participating in the salt-stress response belonged to the hormone pathway, various TF families and the DUF family.ConclusionsZoysia salt treatment transcriptome shows the 24-h and roots may make significant contributions to the salt tolerance. The auxin signal transduction family, ABA signal transduction family, WRKY TF family and bHLH TF family may be the most important families in Zoysia salt-stress regulation.

Genome-wide survey of the bHLH super gene family in Brassica napus

BackgroundThe basic helix-loop-helix (bHLH) gene family is one of the largest transcription factor families in plants and is functionally characterized in diverse species. However, less is known about its functions in the economically important allopolyploid oil crop, Brassica napus.ResultsWe identified 602 potential bHLHs in the B. napus genome (BnabHLHs) and categorized them into 35 subfamilies, including seven newly separated subfamilies, based on phylogeny, protein structure, and exon-intron organization analysis. The intron insertion patterns of this gene family were analyzed and a total of eight types were identified in the bHLH regions of BnabHLHs. Chromosome distribution and synteny analyses revealed that hybridization between Brassica rapa and Brassica oleracea was the main expansion mechanism for BnabHLHs. Expression analyses showed that BnabHLHs were widely in different plant tissues and formed seven main patterns, suggesting they may participate in various aspects of B. napus development. Furthermore, when roots were treated with five different hormones (IAA, auxin; GA3, gibberellin; 6-BA, cytokinin; ABA, abscisic acid and ACC, ethylene), the expression profiles of BnabHLHs changed significantly, with many showing increased expression. The induction of five candidate BnabHLHs was confirmed following the five hormone treatments via qRT-PCR. Up to 246 BnabHLHs from nine subfamilies were predicted to have potential roles relating to root development through the joint analysis of their expression profiles and homolog function.ConclusionThe 602 BnabHLHs identified from B. napus were classified into 35 subfamilies, and those members from the same subfamily generally had similar sequence motifs. Overall, we found that BnabHLHs may be widely involved in root development in B. napus. Moreover, this study provides important insights into the potential functions of the BnabHLHs super gene family and thus will be useful in future gene function research.

Comparative transcriptomic analysis reveals novel roles of transcription factors and hormones during the flowering induction and floral bud differentiation in sweet cherry trees (Prunus avium L. cv. Bing).

In sweet cherry trees, flowering is commercially important because the flowers, after fertilization, will generate the fruits. In P. avium, the flowering induction and flower organogensis are the first developmental steps towards flower formation and they occur within specialized organs known as floral buds during the summer, nine months before blooming. During this period the number of floral buds per tree and the bud fruitfulness (number of flowers per bud) are stablished affecting the potential yield of orchards and the plant architecture. The floral bud development is sensitive to any type of stress and the hotter and drier summers will interfere with this process and are calling for new adapted cultivars. A better understanding of the underlying molecular and hormonal mechanisms would be of help, but unlike the model plant Arabidopsis, very little is known about floral induction in sweet cherry. To explore the molecular mechanism of floral bud differentiation, high-throughput RNA sequencing was used to detect differences in the gene expression of P. avium floral buds at five differentiation stages. We found 2,982 differentially expressed genes during floral bud development. We identified genes associated with floral initiation or floral organ identity that appear to be useful biomarkers of floral development and several transcription factor families (ERF, MYB, bHLH, MADS-box and NAC gene family) with novel potential roles during floral transition in this species. We analyzed in deep the MADS-box gene family and we shed light about their key role during floral bud and organs development in P. avium. Furthermore, the hormonal-related signatures in the gene regulatory networks and the dynamic changes of absicic acid, zeatin and indolacetic acid contents in buds suggest an important role for these hormones during floral bud differentiation in sweet cherry. These data provide a rich source of novel informacion for functional and evolutionary studies about floral bud development in sweet cherry and new tools for biotechnology and breeding.