The basic Helix-Loop-Helix (bHLH) proteins are transcription factors that play important roles during the development of various metazoans including fly, nematode, and vertebrates. They are also involved in human diseases, particularly in cancerogenesis. We made an extensive search for bHLH sequences in the completely sequenced genomes of Caenorhabditis elegans and of Drosophila melanogaster. We found 35 and 56 different genes, respectively, which may represent the complete set of bHLH of these organisms. A phylogenetic analysis of these genes, together with a large number (>350) of bHLH from other sources, led us to define 44 orthologous families among which 36 include bHLH from animals only, and two have representatives in both yeasts and animals. In addition, we identified two bHLH motifs present only in yeast, and four that are present only in plants; however, the latter number is certainly an underestimate. Most animal families (35/38) comprise fly, nematode, and vertebrate genes, suggesting that their common ancestor, which lived in pre-Cambrian times (600 million years ago) already owned as many as 35 different bHLH genes.

碱性/螺旋-环-螺旋（bHLH）转录因子是植物中第二大转录因子家族，该家族广泛存在于各种植物的基因组中，并在植物生长发育、次生代谢、非生物逆境胁迫响应等方面发挥着重要的调控作用。本文全面综述了植物bHLH基因家族成员的结构特征、分类规则及其生物学功能的研究进展，重点梳理总结了bHLH在植物生长发育和非生物胁迫（干旱、低温、盐、重金属）中的应答和调控，以及在次生代谢产物生物合成及动态积累过程中的重要作用，可为深入研究bHLH在生长发育、植物抗逆及品质形成等方面的分子调控机制及种质资源的开发提供指导。同时，因bHLH广泛参与调控植物次生代谢产物的合成和积累，已成为分子生药学和中药生态农业研究的热点。为此，本文进一步总结了近年来研究较为透彻的两种药用植物（丹参Salvia Miltiorrhiza、黄花蒿Artemisia annua）中bHLH基因家族及其成员的研究进展，以期为药用植物bHLH基因家族的深入研究提供参考，并为药用植物的分子育种、拟境栽培等工作的开展以及中药生态农业的发展提供新思路。

bHLH是真核生物中重要的一类转录因子，其主要由碱性氨基酸区和螺旋-环-螺旋区组成。本文利用生物信息学的方法鉴定到122个绿豆bHLH转录因子，并对其理化性质、保守结构域、基因结构、在染色体上的分布、系统进化以及部分典型基因的组织表达差异等进行分析。结果表明，bHLH转录因子理化性质差异较大；含有2个保守结构域，分别位于N端的碱性氨基酸区和C端的螺旋-环-螺旋区，碱性氨基酸区含有His5-Glu9-Arg13保守序列，与靶基因结合有关，HLH区含有Arg23和Arg55，与形成二聚体有关，同时含有5种保守元件；bHLH基因在11条染色体上分布不均匀，5号、7号和8号染色体上分布较多，1号、4号和10号染色体上分布较少，大部分基因含有1-9个不等的内含子，在染色体上成簇状分布；122个bHLH转录因子可分为11个亚家族。多数bHLH基因在绿豆根、茎、叶、花和种子等组织中都有表达，但具有组织表达特异性，且不同基因表达量差异较大。本研究为进一步研究绿豆bHLH转录因子家族的生物学功能奠定基础。

The basic/helix-loop-helix (bHLH) proteins are a superfamily of transcription factors that bind as dimers to specific DNA target sites and that have been well characterized in nonplant eukaryotes as important regulatory components in diverse biological processes. Based on evidence that the bHLH protein PIF3 is a direct phytochrome reaction partner in the photoreceptor's signaling network, we have undertaken a comprehensive computational analysis of the Arabidopsis genome sequence databases to define the scope and features of the bHLH family. Using a set of criteria derived from a previously defined consensus motif, we identified 147 bHLH protein-encoding genes, making this one of the largest transcription factor families in Arabidopsis. Phylogenetic analysis of the bHLH domain sequences permits classification of these genes into 21 subfamilies. The evolutionary and potential functional relationships implied by this analysis are supported by other criteria, including the chromosomal distribution of these genes relative to duplicated genome segments, the conservation of variant exon/intron structural patterns, and the predicted DNA binding activities within subfamilies. Considerable diversity in DNA binding site specificity among family members is predicted, and marked divergence in protein sequence outside of the conserved bHLH domain is observed. Together with the established propensity of bHLH factors to engage in varying degrees of homodimerization and heterodimerization, these observations suggest that the Arabidopsis bHLH proteins have the potential to participate in an extensive set of combinatorial interactions, endowing them with the capacity to be involved in the regulation of a multiplicity of transcriptional programs. We provide evidence from yeast two-hybrid and in vitro binding assays that two related phytochrome-interacting members in the Arabidopsis family, PIF3 and PIF4, can form both homodimers and heterodimers and that all three dimeric configurations can bind specifically to the G-box DNA sequence motif CACGTG. These data are consistent, in principle, with the operation of this combinatorial mechanism in Arabidopsis.

The bHLH (basic helix-loop-helix) transcription factor family is the second largest in plants. It occurs in all three eukaryotic kingdoms, and plays important roles in regulating growth and development. However, family members have not previously been studied in apple. Here, we identified 188 MdbHLH proteins in apple "Golden Delicious" (Malus x domestica Borkh.), which could be classified into 18 groups. We also investigated the gene structures and 12 conserved motifs in these MdbHLHs. Coupled with expression analysis and protein interaction network prediction, we identified several genes thatmight be responsible for abiotic stress responses. This study provides insight and rich resources for subsequent investigations of such proteins in apple.

The basic helix–loop–helix (bHLH) superfamily is considered the second largest transcription factor (TF) family. It plays regulatory roles in the developmental processes of plants and in their defense responses. In recent years, many bHLH superfamily genes have been identified and characterized in herbaceous and woody plants. However, the comprehensive genomic and functional analyses of these genes in eggplant (Solanum melongenaL.) have not been reported. In this study, 121 bHLH TFs were identified in the recently released eggplant genome. The phylogeny, gene structure and conserved motifs of theSmbHLHgene were comprehensively studied. Subsequently, the phylogenetic relationship between the bHLH of eggplant and the bHLH of other species was analyzed, and the proteins were classified into 17 subfamilies. Among these protein sequences, 16 subgroups were clustered into the functional clades ofArabidopsis. Two candidate genes (SmbHLH1,SmbHLH117) that may be involved in anthocyanin biosynthesis were screened. The tissue specificity or differential expression of thebHLHgenes in different tissues and under various light and temperature conditions suggested the differential regulation of tissue development and metabolism. This study not only provides a solid foundation for the functional dissection of the eggplantbHLHgene family but may also be useful for the future synthesis of anthocyanins in eggplant.

bHLH转录因子是植物第二大转录因子家族, 在调节植物生长发育、信号转导和逆境胁迫响应等方面发挥着重要的作用。为了研究玉米bHLH家族基因在逆境胁迫响应中的功能, 本研究从玉米根组织中克隆了ZmbHLH161 (AC: NC_AQK75074)基因。生物信息学分析表明: 该基因包含3个外显子, cDNA全长1460 bp, 编码序列全长1059 bp, 编码352个氨基酸; 在玉米基因组中以单拷贝形式存在, 功能未知; ZmbHLH161蛋白分子量为37.1 kD, 理论等电点为6.10, 具有bHLH转录因子家族特有的保守结构域, 但不具跨膜结构, 无信号肽, 为亲水性蛋白, 蛋白二级结构无规则卷曲所占比例最大, 为42.05%。玉米原生质体瞬时表达试验表明, ZmbHLH161定位在细胞核内。实时荧光定量PCR (qPCR)分析表明, 正常生长条件下, ZmbHLH161主要在根系和幼胚中表达; 在脱水和干旱处理下, ZmbHLH161在玉米苗期叶片中上调表达。转基因异源表达ZmbHLH161拟南芥株系经不同浓度NaCl处理后, 其根长与野生型差异不显著, 而不同浓度甘露醇处理后其根长优于野生型。由此推测ZmbHLH161基因可能参与玉米对渗透胁迫应答。

Integration of diverse environmental and endogenous signals to coordinately regulate growth, development, and defense is essential for plants to survive in their natural habitat. The hormonal signals gibberellin (GA) and jasmonate (JA) antagonistically and synergistically regulate diverse aspects of plant growth, development, and defense. GA and JA synergistically induce initiation of trichomes, which assist seed dispersal and act as barriers to protect plants against insect attack, pathogen infection, excessive water loss, and UV irradiation. However, the molecular mechanism underlying such synergism between GA and JA signaling remains unclear. In this study, we revealed a mechanism for GA and JA signaling synergy and identified a signaling complex of the GA pathway in regulation of trichome initiation. Molecular, biochemical, and genetic evidence showed that the WD-repeat/bHLH/MYB complex acts as a direct target of DELLAs in the GA pathway and that both DELLAs and JAZs interacted with the WD-repeat/bHLH/MYB complex to mediate synergism between GA and JA signaling in regulating trichome development. GA and JA induce degradation of DELLAs and JASMONATE ZIM-domain proteins to coordinately activate the WD-repeat/bHLH/MYB complex and synergistically and mutually dependently induce trichome initiation. This study provides deep insights into the molecular mechanisms for integration of different hormonal signals to synergistically regulate plant development.

Jasmonates (JAs) are plant hormones that regulate the balance between plant growth and responses to biotic and abiotic stresses. Although recent studies have uncovered the mechanisms for JA-induced responses in Arabidopsis thaliana, the mechanisms by which plants attenuate the JA-induced responses remain elusive. Here, we report that a basic helix-loop-helix–type transcription factor, ABA-INDUCIBLE BHLH-TYPE TRANSCRIPTION FACTOR/JA-ASSOCIATED MYC2-LIKE1 (JAM1), acts as a transcriptional repressor and negatively regulates JA signaling. Gain-of-function transgenic plants expressing the chimeric repressor for JAM1 exhibited substantial reduction of JA responses, including JA-induced inhibition of root growth, accumulation of anthocyanin, and male fertility. These plants were also compromised in resistance to attack by the insect herbivore Spodoptera exigua. Conversely, jam1 loss-of-function mutants showed enhanced JA responsiveness, including increased resistance to insect attack. JAM1 and MYC2 competitively bind to the target sequence of MYC2, which likely provides the mechanism for negative regulation of JA signaling and suppression of MYC2 functions by JAM1. These results indicate that JAM1 negatively regulates JA signaling, thereby playing a pivotal role in fine-tuning of JA-mediated stress responses and plant growth.

【目的】研究单盐(NaCl)和复盐(Na₂SO₄、NaHCO₃)胁迫下对胡麻萌发期和苗期生长以及耐盐阈值的影响,为胡麻耐盐材料的筛选提供基础参考。【方法】选用耐盐性不同的2个胡麻品种,设置不同浓度的盐胁迫处理,分析不同指标的变化,采用相关性和回归分析,萌发期单盐胁迫选取总鲜重、复盐胁迫选取发芽率,苗期单盐胁迫选取根干重、复盐胁迫选取总鲜重等指标与盐胁迫浓度建立了二次回归方程:Y=42.949x²-263.158x+224.564、Y=5.194x²-111.128x+109.467、Y=81.772x²-302.544x+382.076、Y=68.568x²-278.918x+326.567,以选定指标较对照降低50%为标准,分别计算萌发期和苗期的耐盐阈值。【结果】萌发期单盐(NaCl)胁迫阈值为103.722 25 mmol/L;萌发期复盐(Na₂SO₄、NaHCO₃)胁迫阈值为55.201 5 mmol/L;;苗期单盐(NaCl)胁迫阈值为251.247 mmol/L;;苗期复盐(Na₂SO₄、NaHCO₃)胁迫阈值为204.25 mmol/L。【结论】胡麻对中性盐比碱性盐的耐受力要强;胡麻不同组织对盐胁迫的耐受力存在较大差别;胡麻萌发期和苗期的耐盐性不具有一致性。

The two most commonly used methods to analyze data from real-time, quantitative PCR experiments are absolute quantification and relative quantification. Absolute quantification determines the input copy number, usually by relating the PCR signal to a standard curve. Relative quantification relates the PCR signal of the target transcript in a treatment group to that of another sample such as an untreated control. The 2(-Delta Delta C(T)) method is a convenient way to analyze the relative changes in gene expression from real-time quantitative PCR experiments. The purpose of this report is to present the derivation, assumptions, and applications of the 2(-Delta Delta C(T)) method. In addition, we present the derivation and applications of two variations of the 2(-Delta Delta C(T)) method that may be useful in the analysis of real-time, quantitative PCR data.Copyright 2001 Elsevier Science (USA).

Flax (Linum usitatissimum) bast fibres are located in the stem cortex where they play an important role in mechanical support. They contain high amounts of cellulose and so are used for linen textiles and in the composite industry. In this study, we screened the annotated flax genome and identified 14 distinct cellulose synthase (CESA) genes using orthologous sequences previously identified. Transcriptomics of 'primary cell wall' and 'secondary cell wall' flax CESA genes showed that some were preferentially expressed in different organs and stem tissues providing clues as to their biological role(s) in planta. The development for the first time in flax of a virus-induced gene silencing (VIGS) approach was used to functionally evaluate the biological role of different CESA genes in stem tissues. Quantification of transcript accumulation showed that in many cases, silencing not only affected targeted CESA clades, but also had an impact on other CESA genes. Whatever the targeted clade, inactivation by VIGS affected plant growth. In contrast, only clade 1- and clade 6-targeted plants showed modifications in outer-stem tissue organization and secondary cell wall formation. In these plants, bast fibre number and structure were severely impacted, suggesting that the targeted genes may play an important role in the establishment of the fibre cell wall. Our results provide new fundamental information about cellulose biosynthesis in flax that should facilitate future plant improvement/engineering.© 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Lignin and cellulose represent the two main components of plant secondary walls and the most abundant polymers on Earth. Quantitatively one of the principal products of the phenylpropanoid pathway, lignin confers high mechanical strength and hydrophobicity to plant walls, thus enabling erect growth and high-pressure water transport in the vessels. Lignin is characterized by a high natural heterogeneity in its composition and abundance in plant secondary cell walls, even in the different tissues of the same plant. A typical example is the stem of fibre crops, which shows a lignified core enveloped by a cellulosic, lignin-poor cortex. Despite the great value of fibre crops for humanity, however, still little is known on the mechanisms controlling their cell wall biogenesis, and particularly, what regulates their spatially-defined lignification pattern. Given the chemical complexity and the heterogeneous composition of fibre crops' secondary walls, only the use of multidisciplinary approaches can convey an integrated picture and provide exhaustive information covering different levels of biological complexity. The present review highlights the importance of combining high throughput -omics approaches to get a complete understanding of the factors regulating the lignification heterogeneity typical of fibre crops.

Background: Basic helix-loop-helix/helix-loop-helix (bHLH/HLH) transcription factors play important roles in plant development. Many reports have suggested that bHLH/HLH proteins participate in brassinosteroid (BR) hormone signaling pathways to promote cell elongation. Cotton fibers are single-cells and derived from seed surface. To explore the roles of bHLH/HLH proteins in cotton fiber development progress by modulating BR signaling pathway, we performed a systematic analysis of the bHLH/HLH gene family in upland cotton (Gossypium hirsutum) genome.Results: In this study, we identified 437 bHLH/HLH genes in upland cotton (G. hirsutum) genome. Phylogenetic analysis revealed that GhbHLH/HLH proteins were split into twenty six clades in the tree. These GhbHLH/HLH genes are distributed unevenly in different chromosomes of cotton genome. Segmental duplication is the predominant gene duplication event and the major contributor for amplification of GhbHLH/HLH gene family. The GhbHLH/HLHs within the same group have conserved exon/intron pattern and their encoding proteins show conserved motif composition. Based on transcriptome data, we identified 77 GhbHLH/HLH candidates that are expressed at relatively high levels in cotton fibers. As adding exogenous BR (brassinolide, BL) or brassinazole (Brz, a BR biosynthesis inhibitor), expressions of these GhbHLH/HLH genes were up-regulated or down-regulated in cotton fibers. Furthermore, overexpression of GhbHLH282 (one of the BR-response genes) in Arabidopsis not only promoted the plant growth, but also changed plant response to BR signaling.Conclusion: Collectively, these data suggested that these GhbHLH/HLH genes may participate in BR signaling transduction during cotton fiber development. Thus, our results may provide a valuable reference data as the basis for further studying the roles of these bHLH/HLH genes in cotton fiber development.