Screening and Identification of the Biomarkers Applied for the Evaluation of Acute and Chronic Thermal Tolerance Ability in Largemouth Bass (Micropterus salmoides).

Affected by the continuously rising temperature, thermal stress leads to a delinked growth rate and resistance to stress in cultured largemouth bass (Micropterus salmoides, LMB) in China. Identification of LMB with better thermal resistance will benefit the breeding of new varieties. However, there has been limited reporting on the evaluation to identify LMB with better thermal resistance. LMB consists of the northern LMB (Micropterus salmoides salmoides, NLMB) and the Florida LMB (Micropterus salmoides floridanus, FLMB). Due to their different geographical distributions, it has been suggested that FLMB exhibit better thermal resistance compared to NLMB. In this study, NLMB and FLMB were subjected to thermal stress for 3 h (acute) and 60 d (chronic) at 33 °C, respectively. Subsequently, the variations of 12 candidate biomarkers between NLMB and FLMB were analyzed. Exposure to acute thermal stress significantly increased plasma cortisol, blood glucose, and lactate levels; activities of superoxide dismutase (SOD), glutathione peroxidase (GPX), catalase (CAT), glucose kinase (GK), pyruvate kinase (PK), lactate dehydrogenase (LDH), and glucose 6 phosphatase (G6Pase); and the expressions of hsp70 and hsp90 in both NLMB and FLMB (p < 0.05). Compared to NLMB, FLMB exhibited a lower plasma cortisol level and a higher expression of hsp90 under acute thermal stress (p < 0.05). Exposure to chronic thermal stress significantly increased plasma cortisol and blood glucose levels, as well as activities of GK, PK, LDH, and G6Pase, as well as expressions of hsp70 and hsp90 in both NLMB and FLMB (p < 0.05). Additionally, FLMB showed a lower expression of hsp70 compared to NLMB (p < 0.05). In conclusion, our results showed that LMB with lower plasma cortisol level and higher expression of hsp90 under acute thermal stress, as well as lower expression of hsp70 under chronic thermal stress were suggested to have better thermal resistance. Our study provides valuable information for identifying and breeding LMB varieties with better thermal resistance in the future.

Comprehensive investigation of BZR gene family in four dicots and the function of PtBZR9 and PtBZR12 under drought stress.

Brassinazole-resistant (BZR) transcription factor plays an important role in plant growth and stress resistance through brassinosteroid (BR) signal transduction. However, systematic analysis of the BZR family in dicots remains limited. In this study, we conducted a genome-wide study of four typical dicots: Arabidopsis thaliana, Carica papaya, Vitis vinifera and Populus trichocarpa. Thirty-four BZR gene family members were identified and classified them into three subfamilies. Analysis of promoter and expression patterns revealed crucial role of a pair of homologous BZR genes, PtBZR9 and PtBZR12, in poplar may play a critical role under abiotic stress. PtBZR9 and PtBZR12 were localised in the nucleus and exhibited mutual interactions. Moreover, transient overexpression (OE) of PtBZR9 and PtBZR12 in poplar enhanced tolerance to drought stress. The phenotypic and physiological characteristics of PtBZR9 and PtBZR12 OE in Arabidopsis mirrored those of transient OE in the poplar. Additionally, PtBZR9 and PtBZR12 can bind to the E-box element. Under exogenous BR treatment, transgenic lines displayed a greater decrease in root length than the wild type. Thus, these findings provide a solid foundation for future research on the complex regulatory mechanisms of BZR genes.

Geranyl diphosphate synthase large subunits OfLSU1/2 interact with small subunit OfSSUII and are involved in aromatic monoterpenes production in Osmanthus fragrans.

Osmanthus fragrans is a famous ornamental tree species for its pleasing floral fragrance. Monoterpenoids are the core floral volatiles of O. fragrans flowers, which have tremendous commercial value. Geranyl diphosphate synthase (GPPS) is a key enzyme that catalyzes the formation of GPP, the precursor of monoterpenoids. However, there are no reports of GPPSs in O. fragrans. Here, we performed RNA sequencing on the O. fragrans flowers and identified three GPPSs. Phylogenetic tree analysis showed that OfLSU1/2 belonged to the GPPS.LSU branch, while the OfSSUII belonged to the GPPS.SSU branch. OfLSU1, OfLSU2 and OfSSUII were all localized in chloroplasts. Y2H and pull-down assays showed that OfLSU1 or OfLSU2 interacted with OfSSUII to form heteromeric GPPSs. Site mutation experiments revealed that the conserved CXXXC motifs of OfLSU1/2 and OfSSUII were essential for the interaction between OfLSU1/2 and OfSSUII. Transient expression experiments showed that OfLSU1, OfLSU2 and OfSSUII co-expressed with monoterpene synthase genes OfTPS1 or OfTPS2 improved the biosynthesis of monoterpenoids (E)-ß-ocimene and linalool. The heteromeric GPPSs formed by OfLSU1/2 interacting with OfSSUII further improves the biosynthesis of monoterpenoids. Overall, these preliminary results suggested that the GPPSs play a key role in regulating the production of aromatic monoterpenes in O. fragrans.

The R2R3-MYB transcription factor OfMYB21 positively regulates linalool biosynthesis in Osmanthus fragrans flowers.

Osmanthus fragrans is a well-known landscape ornamental tree species for its pleasing floral fragrance and abundance of flowers. Linalool, the core floral volatiles of O. fragrans, has tremendous economic value in the pharmaceuticals, cleaning products and cosmetics industries. However, the transcriptional regulatory network for the biosynthesis of linalool in O. fragrans remains unclear. Here, OfMYB21, a potential transcription factor regulating the linalool synthetase OfTPS2, was identified using RNA-seq data and qRT-PCR analysis. Yeast one-hybrid, dual-luciferase and EMSA showed that OfMYB21 directly binds to the promoter of OfTPS2 and activates its expression. Overexpression of OfMYB21 in the petals of O. fragrans led to up-regulation of OfTPS2 and increased accumulation of linalool, while silencing of OfMYB21 led to down-regulation of OfTPS2 and decreased biosynthesis of linalool. Subsequently, yeast two-hybrid, pull-down and BiFC experiments showed that OfMYB21 interacts with JA signaling factors OfJAZ2/3 and OfMYC2. Interestingly, the interaction between OfMYC2 and OfMYB21 further enhanced the transcription of OfTPS2, whereas OfJAZ3 attenuated this effect. Overall, our studies provided novel finding on the regulatory mechanisms responsible for the biosynthesis of the volatile monoterpenoid linalool in O. fragrans.

Genome-Wide Identification, Evolution and Expression Profile Analysis of NAC Transcription Factor in Simmondsia chinensis.

NAC transcription factors (TFs) are one of the largest plant-specific gene families and play important roles in plant growth, development, and the biotic and abiotic stress response. Although the sequencing of Jojoba (Simmondsia chinensis) has been completed, the genome-wide identification and analysis of its NAC TFs has not been reported. In this study, a total of 57 genes were identified in Jojoba, which were divided into eight groups based on phylogenetic analysis. The genes clustered in the same groups have a similar gene structure and motif distribution. Based on the analysis of cis-elements in NAC TFs, nine cis-acting elements were identified in the promoter region that involved in light response, hormonal response, and stress response. Synteny analysis showed a greater collinearity between Jojoba and V. vinifera than Arabidopsis thaliana. The 24 genes in the Jojoba NAC TFs are derived from fragment replication, which may be the main source of NAC amplification. Gene expression analysis identified seven genes that were highly expressed in seeds. The differential expression analysis of NAC TFs in cotyledon and embryonic axis tissues showed that the expression of 10 genes was up-regulated and 1 gene was down-regulated. This study provides more information on the classification, gene structure, conserved motif, and evolution of NAC TFs in Jojoba, facilitating further exploration of their specific functional analysis in Jojoba seed development.

Whole-genome identification and multiple abiotic stresses expression pattern profiling analysis of PLATZ transcription factor family members in Pecan (Carya illinoensis).

Plant AT-rich sequence and zinc-binding (PLATZ), as a plant-specific transcription factor, have been identified and studied in a variety of plants. However, there are no reports about PLATZ proteins in Carya illinoensis (pecan). Here, 24 C. illinoensis CiPLATZs have been identified and divided into 4 groups. Gene structure, motif composition, conserved domain and cis-acting elements analysis indicated that the PLATZ gene family was highly conserved. Transcriptome data combined with qRT-PCR analysis revealed that CiPLATZ6, CiPLATZ12, CiPLATZ13, CiPLATZ14 and CiPLATZ23 were highly expressed in multiple tissues of C. illinoensis and strongly responded to drought, salt and heat stress. Among them, CiPLATZ6, CiPLATZ12 and CiPLATZ23 were all located in the nucleus and had no transcriptional autoactivation ability in yeast cells, and acted as transcriptional suppressors in plants. In addition, the CiPLATZ23-overexpressing transgenic Arabidopsis thaliana showed enhanced tolerance to drought. Measurements of physiological indicators and analysis of stress-related genes expression levels in transgenic A. thaliana were used to support this conclusion. The results of this study are helpful to understand the structural feature and function of CiPLATZs, and provide candidate genes for molecular breeding of drought tolerance of C. illinoensis.

Comprehensive analysis of MAPK gene family in Populus trichocarpa and physiological characterization of PtMAPK3-1 in response to MeJA induction.

Mitogen-activated protein kinases (MAPKs) play important roles in plant growth and development, as well as hormone and stress responses by signaling to eukaryotic cells, through MAPK cascade, the presence of various cues; thereby, regulating various responses. The MAPK cascade consists mainly of three gene families, MAPK, MAPKK, and MAPKKK, which activate downstream signaling pathways through sequential phosphorylation. Although the MAPK cascade gene family has been reported in several species, there is a lack of comprehensive analysis in poplar. We identified 21 MAPK genes, 11 MAPKK genes, and 104 MAPKKK genes in Populus trichocarpa. The phylogenetic classification was supported by conservative motif, gene structure and motif analysis. Whole genome duplication has an important role in the expansion of MAPK cascade genes. Analysis of promoter cis-elements and expression profiles indicates that MAPK cascade genes have important roles in plant growth and development, abiotic and biotic stresses, and phytohormone response. Expression profiling revealed a significant upregulation of PtMAPK3-1 expression in response to drought, salt and disease stresses. Poplar transiently overexpressing PtMAPK3-1 and treated with methyl jasmonic acid (MeJA) had higher catalase and peroxidase levels than non-overexpressing poplar. This work represents the first complete inventory of the MAPK cascade in P. trichocarpa, which reveals that PtMAPK3-1 is induced by the MeJA hormone and participates in the MeJA-induced enhancement of the antioxidant enzyme system.

Correction: Wu et al. Homologous Drought-Induced 19 Proteins, PtDi19-2 and PtDi19-7, Enhance Drought Tolerance in Transgenic Plants. Int. J. Mol. Sci. 2022, 23, 3371.

The authors wish to make the following corrections to this paper [...].

SAUR8, a small auxin-up RNA gene in poplar, confers drought tolerance to transgenic Arabidopsis plants.

SAUR (small auxin-up RNA) is an early auxin-responsive gene. In this study, a novel SAUR gene PtSAUR8 was cloned from poplar (Populus trichocarpa), and subcellular location analysis showed that it is targeted to the nuclear membrane. In addition, PtSAUR8 overexpression in Arabidopsis improved the plant resistance to drought stress. Meanwhile, the loss of function mutant saur53 showed more drought sensitivity compared to the WT. PtSAUR8 conferred drought tolerance in transgenic Arabidopsis, as determined through phenotypic and stress-associated physiological indicator analyses, namely, root length, germination rate, relative water content, proline content, CAT content, POD content, malondialdehyde content, hydrogen peroxide content, and relative conductivity. In addition, after the 1 µM abscisic acid (ABA) treatment, the PtSAUR8-OE lines promoted stomata closure. Quantitative fluorescence analysis of related genes induced by drought mutant stress further confirmed that overexpression of PtSAUR8 can improve drought resistance in transgenic Arabidopsis lines. Therefore, PtSAUR8 may play a role in plant drought resistance through ABA-mediated pathways; thus, providing new research materials for molecular breeding of poplar resistance.

Homologous Drought-Induced 19 Proteins, PtDi19-2 and PtDi19-7, Enhance Drought Tolerance in Transgenic Plants.

Drought-induced 19 (Di19) proteins play important roles in abiotic stress responses. Thus far, there are no reports about Di19 family in woody plants. Here, eight Di19 genes were identified in poplar. We analyzed phylogenetic tree, conserved protein domain, and gene structure of Di19 gene members in seven species. The results showed the Di19 gene family was very conservative in both dicotyledonous and monocotyledonous forms. On the basis of transcriptome data, the expression patterns of Di19s in poplar under abiotic stress and ABA treatment were further studied. Subsequently, homologous genes PtDi19-2 and PtDi19-7 with strong response to drought stress were identified. PtDi19-2 functions as a nuclear transcriptional activator with a transactivation domain at the C-terminus. PtDi19-7 is a nuclear and membrane localization protein. Additionally, PtDi19-2 and PtDi19-7 were able to interact with each other in yeast two-hybrid system. Overexpression of PtDi19-2 and PtDi19-7 in Arabidopsis was found. Phenotype identification and physiological parameter analysis showed that transgenic Arabidopsis increased ABA sensitivity and drought tolerance. PtDi19-7 was overexpressed in hybrid poplar 84K (Populus alba × Populus glandulosa). Under drought treatment, the phenotype and physiological parameters of transgenic poplar were consistent with those of transgenic Arabidopsis. In addition, exogenous ABA treatment induced lateral bud dormancy of transgenic poplar and stomatal closure of transgenic Arabidopsis. The expression of ABA/drought-related marker genes was upregulated under drought treatment. These results indicated that PtDi19-2 and PtDi19-7 might play a similar role in improving the drought tolerance of transgenic plants through ABA-dependent signaling pathways.

The poplar VQ1 gene confers salt tolerance and pathogen resistance in transgenic Arabidopsis plants via changes in hormonal signaling.

The VQ protein family is plant-specific, and is involved in growth, development, and biotic and abiotic stress responses. In this study, we found that the gene expression of poplar VQ1(Potri.001G029700) from Populus trichocarpa varied remarkably under salt stress and hormones associated with disease. A subcellular localization experiment showed that VQ1 was localized in the nucleus and cytomembrane in tobacco. The overexpression of VQ1 in Arabidopsis thaliana enhanced its resistance to salt stress and disease, and was also responsive to it through abscisic acid. Compared with wild-type, transgenic Arabidopsis lines had significantly increased levels of abscisic acid and salicylic acid. The expression of some stress-related genes, such as MPK6, NPR1, and PDF1.2, was significantly up-regulated by salt in transgenic plants, while WRKY70, ABI1, KUP6, and NCED2 were significantly down-regulated by Pseudomonas syringae pv. tomato DC3000 in transgenic plants. Together, these results demonstrate that VQ1 modulates hormonal signaling to confer multiple biotic and abiotic stress responses in transgenic Arabidopsis plants.

Systematic analysis of the Serine/Arginine-Rich Protein Splicing Factors (SRs) and focus on salt tolerance of PtSC27 in Populus trichocarpa.

Serine/Arginine-Rich Protein Splicing Factors (SRs) are indispensable splicing factors, which play significant roles in spliceosome assembly, splicing regulation and regulation of plant stress. However, a comprehensive analysis and function research of SRs in the woody plant is still lacking. In this report, we conducted the identification and comprehensive analysis of the 71 SRs in poplar and three other dicots, including basic characterization, phylogenetic, conserved motifs, gene duplication, promoter and splice isoform of these genes. Based on the publicly available transcriptome data, expression pattern of SRs in poplar under low temperature, high temperature, drought and salt stress were further analyzed. Subsequently, a key candidate gene PtSC27 that responded to salt stress was screened. More importantly, overexpression of PtSC27 increased plant survival rate under salt stress, and enhanced salt tolerance by regulating malondialdehyde (MDA) content, peroxidase (POD) and catalase (CAT) enzyme activities in transgenic plants. Meanwhile, overexpression of PtSC27 made transgenic plants insensitive to exogenous ABA and improved the expression of some ABA signal-related genes under salt stress. Overall, our studies lay a foundation for understanding the structure and function of SRs in the poplar and provide useful gene resources for breeding through genetic engineering.

GEPSdb: The Gene Expression Database of Poplar under Stress.

As a model tree species, poplar (Populus L.) has important economic and ecological value. Here, we constructed the GEPSdb (Gene Expression Database of Poplar under Stress; http://gepsdb.ahau-edu.cn/), which is an integrated database of poplar gene expression profiles derived from RNA-seq and microarray library data. This database provides a comprehensive collection of gene expression data from poplar exposed to 14 types of environmental stress from 11 high-quality RNA-seq experiments and 51 microarray libraries. The GEPSdb includes 56 genes from previous literature that have been examined in poplar and functionally verified. By incorporating data from numerous expression analyses, GEPSdb provides a user-friendly web interface for querying, browsing, and visualizing the expression profiles of related genes. Consequently, GEPSdb can be used to link transcription data with phenotypes and can enhance our understanding of important biological processes and mechanisms underlying complex agronomic traits in poplar.

Comparative analysis of the stellacyanins (SCs) family and focus on drought resistance of PtSC18 in Populus trichocarpa.

Stellacyanin (SC) is a type I (blue) copper protein, which plays a crucial role in plant growth and stress response. However, the comprehensive analysis and functional research of SCs in the woody plant is still lacking. Here, a total of 74 SCs were collected and identified from Arabidopsis, papaya, grape, rice and poplar. Bioinformatics was used to analyze the gene structure, protein structure and evolutionary relationship of 74 genes, especially 19 SCs in Populus trichocarpa. Based on the RNA-seq data, expression pattern of SCs in poplar under cold, high temperature, drought and salt stress were further analyzed. Subsequently, a key candidate gene PtSC18 that strongly responded to drought stress was screened. Subcellular localization experiment exhibited that PtSC18 was localized in the nucleus and plasma membrane. Overexpression of PtSC18 enhanced drought tolerance of transgenic Arabidopsis by improving water retention and reducing oxidative damage. Measurements of physiological indicators, including chlorophyll, H2O2, malondialdehyde content, peroxidase and catalase enzyme activities and electrical conductivity, all supported this conclusion. More importantly, PtSC18 enhanced the expression of some stress-related genes in transgenic Arabidopsis. Overall, our results lay a foundation for understanding the structure and function of PtSCs and provide useful gene resources for breeding through genetic engineering.

Genome and transcriptome analysis to understand the role diversification of cytochrome P450 gene under excess nitrogen treatment.

BACKGROUND: Panax notoginseng (Burk.) F. H. Chen (P. notoginseng) is a medicinal plant. Cytochrome P450 (CYP450) monooxygenase superfamily is involved in the synthesis of a variety of plant hormones. Studies have shown that CYP450 is involved in the synthesis of saponins, which are the main medicinal component of P. notoginseng. To date, the P. notoginseng CYP450 family has not been systematically studied, and its gene functions remain unclear. RESULTS: In this study, a total of 188 PnCYP genes were identified, these genes were divided into 41 subfamilies and clustered into 9 clans. Moreover, we identified 40 paralogous pairs, of which only two had Ka/Ks ratio greater than 1, demonstrating that most PnCYPs underwent purification selection during evolution. In chromosome mapping and gene replication analysis, 8 tandem duplication and 11 segmental duplication events demonstrated that PnCYP genes were continuously replicating during their evolution. Gene ontology (GO) analysis annotated the functions of 188 PnCYPs into 21 functional subclasses, suggesting the functional diversity of these gene families. Functional divergence analyzed the members of the three primitive branches of CYP51, CYP74 and CYP97 at the amino acid level, and found some critical amino acid sites. The expression pattern of PnCYP450 related to nitrogen treatment was studied using transcriptome sequencing data, 10 genes were significantly up-regulated and 37 genes were significantly down-regulated. Combined with transcriptome sequencing analysis, five potential functional genes were screened. Quantitative real-time PCR (qRT-PCR) indicated that these five genes were responded to methyl jasmonate (MEJA) and abscisic acid (ABA) treatment. CONCLUSIONS: These results provide a valuable basis for comprehending the classification and biological functions of PnCYPs, and offer clues to study their biological functions in response to nitrogen treatment.

Genome-wide identification and expression analysis of the NF-Y transcription factor family in Populus.

In the past few years, many studies have reported that the transcription factor Nuclear Factor Y (NF-Y) gene family plays important roles in embryonic development, photosynthesis, flowering time regulation and stress response, in various plants. Although the NF-Y gene family has been systematically studied in many species, little is known about NF-Y genes in Populus. In this study, the NF-Y gene family in the Populus genome was identified and its structural characteristics were described. Fifty-two NF-Y genes were authenticated in the Populus trichocarpa genome and categorized into three subfamilies (NF-YA/B/C) by phylogenetic analysis. Chromosomal localization of these genes revealed that they were distributed randomly across 17 of the 19 chromosomes. Segmental duplication played a vital role in the amplification of Populus NF-Y gene family. Moreover, microsynteny analysis indicated that, among Populus trichocarpa, Arabidopsis thaliana, Vitis vinifera and Carica papaya, NF-Y duplicated regions were more conserved between Populus trichocarpa and Vitis vinifera. Redundant stress-related cis-elements were also found in the promoters of most 13 NF-YA genes and their expression levels varied widely following drought, salt, ABA and cold treatments. Subcellular localization experiments in tobacco showed that PtNF-YA3 was localized in nucleus and cytomembrane, while PtNF-YA4 was only in the nucleus in tobacco. According to the transcriptional activity experiments, neither of them had transcriptional activity in yeast. In summary, a comprehensive analysis of the Populus NF-Y gene family was performed to establish a theoretical basis for further functional studies on this family.

Identification of Orphan Genes in Unbalanced Datasets Based on Ensemble Learning.

Orphan genes are associated with regulatory patterns, but experimental methods for identifying orphan genes are both time-consuming and expensive. Designing an accurate and robust classification model to detect orphan and non-orphan genes in unbalanced distribution datasets poses a particularly huge challenge. Synthetic minority over-sampling algorithms (SMOTE) are selected in a preliminary step to deal with unbalanced gene datasets. To identify orphan genes in balanced and unbalanced Arabidopsis thaliana gene datasets, SMOTE algorithms were then combined with traditional and advanced ensemble classified algorithms respectively, using Support Vector Machine, Random Forest (RF), AdaBoost (adaptive boosting), GBDT (gradient boosting decision tree), and XGBoost (extreme gradient boosting). After comparing the performance of these ensemble models, SMOTE algorithms with XGBoost achieved an F1 score of 0.94 with the balanced A. thaliana gene datasets, but a lower score with the unbalanced datasets. The proposed ensemble method combines different balanced data algorithms including Borderline SMOTE (BSMOTE), Adaptive Synthetic Sampling (ADSYN), SMOTE-Tomek, and SMOTE-ENN with the XGBoost model separately. The performances of the SMOTE-ENN-XGBoost model, which combined over-sampling and under-sampling algorithms with XGBoost, achieved higher predictive accuracy than the other balanced algorithms with XGBoost models. Thus, SMOTE-ENN-XGBoost provides a theoretical basis for developing evaluation criteria for identifying orphan genes in unbalanced and biological datasets.

A Moso bamboo gene VQ28 confers salt tolerance to transgenic Arabidopsis plants.

MAIN CONCLUSION: Overexpression ofPeVQ28in Arabidopsis regulated the expression of salt/ABA-responsive genes and indicated thatPeVQ28may affect the ABA synthesis induced by stress in plants by regulating salt tolerance. Plant-specific VQ proteins, which contain a conserved short FxxhVQxhTG amino acid sequence motif, play an important role in abiotic stress responses, but their functions have not been previously studied in Moso bamboo (Phyllostachys edulis). In this study, real-time quantitative PCR analysis indicated that expression of PeVQ28 was induced by salt and abscisic acid stresses. A subcellular localization experiment showed that PeVQ28 was localized in the nuclei of tobacco leaf cells. Yeast two-hybrid and bimolecular fluorescence complementation analyses indicated that PeVQ28 and WRKY83 interactions occurred in the nucleus. The PeVQ28-overexpressing Arabidopsis lines showed increased resistance to salt stress and enhanced sensitivity to ABA. Compared with wild-type plants under salt stress, PeVQ28-transgenic plants had lower malondialdehyde and higher proline contents, which might enhance stress tolerance. Overexpression of PeVQ28 in Arabidopsis enhanced expression of salt- and ABA-responsive genes. These results suggest that PeVQ28 functions in the positive regulation of salt tolerance mediated by an ABA-dependent signaling pathway.

Genome-Wide Identification and Expression Analyses of the bZIP Transcription Factor Genes in moso bamboo (Phyllostachys edulis).

The basic leucine zipper (bZIP) transcription factor (TF) family is one of the largest gene families, and play crucial roles in many processes, including stress responses, hormone effects. The TF family also participates in plant growth and development. However, limited information is available for these genes in moso bamboo (Phyllostachys edulis), one of the most important non-timber forest products in the world. In the present study, 154 putative PhebZIP genes were identified in the moso bamboo genome. The phylogenetic analyses indicate that the PhebZIP gene proteins classify into 9 subfamilies and the gene structures and conserved motifs that analyses identified among all PhebZIP proteins suggested a high group-specificity. Microsynteny and evolutionary patterns analyses of the non-synonymous (Ka) and synonymous (Ks) substitution rates and their ratios indicated that paralogous pairs of PhebZIP genes in moso bamboo underwent a large-scale genome duplication event that occurred 7-15 million years ago (MYA). According to promoter sequence analysis, we further selected 18 genes which contain the higher number of cis-regulatory elements for expression analysis. The result showed that these genes are extensively involved in GA-, ABA- and MeJA-responses, with possibly different mechanisms. The tissue-specific expression profiles of PhebZIP genes in five plant tissues/organs/developmental stages suggested that these genes are involved in moso bamboo organ development, especially seed development. Subcellular localization and transactivation activity analysis showed that PhebZIP47 and PhebZIP126 were localized in the nucleus and PhebZIP47 with no transcriptional activation in yeast. Our research provides a comprehensive understanding of PhebZIP genes and may aid in the selection of appropriate candidate genes for further cloning and functional analysis in moso bamboo growth and development, and improve their resistance to stress during their life.

The trihelix family of transcription factors: functional and evolutionary analysis in Moso bamboo (Phyllostachys edulis).

BACKGROUND: Trihelix transcription factors (TTFs) are photoresponsive proteins that have a representative three-helix structure (helix-loop-helix-loop-helix). Members of this gene family have been reported to play roles in many plant processes. RESULTS: In this study, we performed a functional and evolutionary analysis of the TTFs in Moso bamboo (Phyllostachys edulis). A total of 35 genes were identified and grouped into five subfamilies (GT-1, GT-γ, GT-2, SIP1 and SH4) according to their structural properties. Gene structure analysis showed that most genes in the PeTTF family had fewer introns. A unique motif (Motif 16) to the GT-γ subfamily was identified by conserved motif analysis. Promoter analysis revealed various cis-acting elements related to plant growth and development, abiotic and biotic stresses, and phytohormone responses. Data for the 35 Moso bamboo TTF genes were used to generate heat maps, which indicated that these genes were expressed in different tissues or developmental stages. Most of the TTF genes identified here had high expression in leaves and panicles according to the expression profile analysis. The expression levels of the TTF members in young leaves were studied using quantitative real-time PCR to determine their tissue specificity and stress-related expression patterns to help functionally characterize individual members. CONCLUSIONS: The results indicated that members of the TTF gene family may be involved in plant responses to stress conditions. Additionally, PeTTF29 was shown to be located in the nucleus by subcellular localization analysis and to have transcriptional activity in a transcriptional activity assay. Our research provides a comprehensive summary of the PeTTF gene family, including functional and evolutionary perspectives, and provides a basis for functionally characterizing these genes.