High-throughput sequencing of highbush blueberry transcriptome and analysis of basic helix-loop-helix transcription factors

The highbush blueberry（Vaccinium corymbosum）,Duke,was used to construct a de novo transcriptome sequence library and to perform data statistical analysis.Mega 4,CLC Sequence Viewer 6 software,and quantitative PCR were employed for bioinformatics and expression analyses of the basic helix-loop-helix（BHLH）transcription factors of the sequencing library.The results showed that 28.38 gigabytes of valid data were obtained from transcriptome sequencing and were assembled into 108 033 unigenes.Functional annotation showed that 32 244 unigenes were annotated into Clusters of Orthologous Groups（COG）and Gene Ontology（GO）databases,whereas the rest of the 75 789 unigenes had no matching information.By using COG and GO classification tools,sequences with annotation information were divided into 25 and 52 categories,respectively,which involved transport and metabolism,transcriptional regulation,and signal transduction.Analysis of the transcriptome library identified a total of 59 BHLH genes.Sequence analysis revealed that 55 genes of that contained a complete BHLH domain.Furthermore,phylogenetic analysis showed that BHLH genes of blueberry（Duke）could be divided into 13 sub-groups.PCR results showed that 45 genes were expressed at various developmental stages of buds,stems,leaves,flowers,and fruits,suggesting that the function of BHLH was associated with the development of different tissues and organs of blueberry,Duke.The present study would provided a foundation for further investigations on the classification and functions of the blueberry BHLH family.

Function of pioneer neurons specified by the basic helix-loop-helix transcription factor atonal in neural development

Basic helix-loop-helix （bHLH） transcription factors regulate the differentiation of various tissues in a vast diversity of species. The bHLH protein Atonal was first identified as a proneural gene involved in the formation of mechanosensory cells and photoreceptor cells in Drosophila （larman et al., 1993, 1994）. Atonal is expressed in sensory organ precursors and is required and sufficient for the development of chordotonal organs （Jar- man et al., 1993）. Moreover, Atonal expression is observed in the developing eye and is essential for the differentiation of R8 photoreceptors, which are the first photoreceptors that appear during development. Atonal is not involved in the formation of other photoreceptors （R1-R7） directly. However, R8 photore- ceptors recruit other photoreceptors from the surrounding cells （Jarman et al., 1994）.

果蝇Tap蛋白结构与功能的生物信息学分析

目的利用生物信息学方法分析果蝇Tap蛋白的结构和功能。方法基于NCBI数据库中果蝇Tap蛋白的氨基酸序列,从蛋白质理化性质、跨膜区、信号肽、亚细胞定位、结构域、三维结构及物种间同源蛋白进化关系等方面进行分析。结果Tap蛋白为不稳定亲水性蛋白,无跨膜区和信号肽,在细胞核中发挥生物学效应,具有碱性螺旋-环-螺旋(bHLH)结构域。Tap蛋白与来源于NeuroD1的模板2ql2.1.B有61.02%的氨基酸序列一致,Tap蛋白与人类和啮齿动物的编码产物高度同源,在系统发生树中距离最近。结论果蝇Tap蛋白具有bHLH蛋白家族的典型结构,可能在果蝇胚胎发育早期的神经发生、分化等过程中发挥作用。

围绕中国新音乐发展九大论域的思潮之辩——“吕贺之争”的基本母题

"吕贺之争"在事关中国新音乐文化建设之"音乐本质论""音政关系论""中西关系论""古今关系论""雅俗关系论""形式技术论""批评标准论""批评方法论""统一战线论"等九大理论领域内,均发生过不同观念、主张和见解的思潮之辩,以吕骥、贺绿汀为代表的这两股音乐思潮,从正反两方面对中国新音乐文化建设产生了重大影响。

IQSEC1通过病毒蛋白PB1调控甲型流感病毒的增殖

目的探讨IQSEC1是否通过病毒蛋白PB1调控甲型流感病毒的增殖。方法首先克隆甲型流感病毒[A/Shanghai/02/2013(H7N9)]的8个基因;其次,通过免疫共沉淀检测IQ模体Sec7结构域蛋白1(IQSEC1)与聚合酶PB1(PB1)存在相互作用;此外,通过过表达或者敲低IQSEC1的方法检测IQSEC1对PB1核定位的影响;最后,过表达或者敲低IQSEC1后检测Influenza A virus[A/Shanghai/02/2013(H7N9)]。结果病毒感染条件下,外源IQSEC1和PB1存在相互作用。当过表达IQSEC1时,细胞中IQSEC1的表达量上升,相应的PB1在细胞核中的定位减少;当用敲低IQSEC1时,细胞中IQSEC1的表达量下降,相应的PB1在细胞核中的定位上升。过表达IQSEC1后,甲型流感病毒的增殖水平下降(P<0.05)。敲低IQSEC1后,甲型流感病毒的增殖水平上升(P<0.05)。结论IQSEC1通过减少甲型流感病毒蛋白PB1的核定位抑制甲型流感病毒的增殖。

我国基础研究管理及科研合作模式的多层次对比研究

根据我国现行旨在发展基础研究的科技计划,提出自由探索型和需求攻关型两种基础研究管理模式。基于CNKI数据库中国家自然科学基金与国家重点基础研究发展计划(“973计划”)发文数据构建科研合作网络,从宏观网络整体、中观网络成分和微观基元模体3个尺度,总结我国基础研究的4种科研合作模式,并对比分析两种管理模式的优缺点。结果表明,需求攻关型管理模式具有更多平衡型和流线型科研合作模式,自由探索型具有更多单点型、流线型和核心型科研合作模式,两种管理模式均需要大幅提升科研合作广度与深度;需求攻关型可以补短板但应对未来不确定性的能力不足,自由探索型可以应对不确定性但无法快速修补短板,两者分别作为长短期方案结合使用才能解决我国基础研究的“卡脖子”问题。

Expression of a novel bHLH-Zip gene in human testis

<abstract>Aim: To identify specifically expressed genes in the adult and fetal testes. Methods: A human testis cDNA microarray was established. Then the mRNA of adult and fetal testis was purified and probes were prepared by a reverse transcription reaction with the testis mRNA as template. The microarray was hybridized with probes of adult and fetal testes. The nucleic sequences of differentially expressed genes were determined and homologies were searched in the databases of the GenBank. Results: When hybridized with adult or fetal testis probes, the positive clones were 96.8 % and 95.4 %, respectively. Among these genes, one was a new testis-specific gene, which was named TSP1. TSP1 was highly expressed in human adult testis. The cDNA of TSP1 was 1,484 bp in length. The cDNA sequence of this clone was deposited in the Genbank (AF333098). TSP1 was also determined as Interim Gen Symbol (Unigene, No. Hs.98266). Protein analysis showed that TSP1 contained two functional domains: an N-terminal basic helix-loop-helix (bHLH) and a C-terminal leucine zipper (Zip). Homologous analysis showed that the 430 amino acid sequences deduced from the 1,293 bp open reading frame (ORF) had a homology with the human gene FLJ2509 (AK098575). TSP1 had also a sequence homology with Spz 1 protein of mouse. Expression profiles showed that TSP1 was specifically and strongly expressed in the testis. Conclusion: TSP1 is a gene highly expressed in adult testis. It may play an important role in spermatogenesis in the humans.

The bHLH transcription factor GhPAS1 mediates BR signaling to regulate plant development and architecture in cotton

Cotton (Gossypium spp.) is the most important natural textile fiber crop in the world.The ideal plant architecture of cotton is suitable for mechanical harvesting and productivity in modern agricultural production.However,cotton genes regulating plant development and architecture have not been fully identified.We identified a basic helix-loop-helix (b HLH) transcription factor,GhPAS1 (PAGODA1 SUPPRESSOR1) in G.hirsutum (Upland cotton).GhPAS1 was located in the nucleus and showed a strong transcription activation effect.Tissue-specific expression patterns showed that GhPAS1 was highly expressed in floral organs,followed by high expression in early stages of ovule development and rapid fiber elongation.GhPAS1 overexpression in Arabidopsis and BRZ (brassinazole,BR biosynthesis inhibitor) treatment indicated that GhPAS1 positively regulates and responds to the BR (brassinosteroid) signaling pathway and promotes cell elongation.GhPAS1 overexpression in Arabidopsis mediated plant development in addition to increasing plant biomass.Virus-induced gene silencing of GhPAS1 indicated that down-regulation of GhPAS1 inhibited cotton growth and development,as plant height,fruit branch length,and boll size of silenced plants were lower than in control plants.Fiber length and seed yield were also lower in silenced plants.We conclude that GhPAS1,a b HLH transcription factor,regulates plant development and architecture in cotton.These findings may help breeders and researchers develop cotton cultivars with desirable agronomic characteristics.

The contribution of oligodendrocytes and oligodendrocyte progenitor cells to central nervous system repair in multiple sclerosis： perspectives for remyelination therapeutic strategies

Oligodencrocytes（OLs） are the main glial cells of the central nervous system involved in myelination of axons. In multiple sclerosis（MS）, there is an imbalance between demyelination and remyelination processes, the last one performed by oligodendrocyte progenitor cells（OPCs） and OLs, resulting into a permanent demyelination, axonal damage and neuronal loss. In MS lesions, astrocytes and microglias play an important part in permeabilization of blood-brain barrier and initiation of OPCs proliferation. Migration and differentiation of OPCs are influenced by various factors and the process is finalized by insufficient acummulation of OLs into the MS lesion. In relation to all these processes, the author will discuss the potential targets for remyelination strategies.

Effects of histone acetylation and DNA methylation on p21^(WAF1)regulation

Cell cycle progression is regulated by interactions between cyclins and cyclin-dependent kinases (CDKs). p21(WAF1) is one of the CIP/KIP family which inhibits CDKs activity. Increased expression of p21(WAF1) may play an important role in the growth arrest induced in transformed cells. Although the stability of the p21( WAF1) mRNA could be altered by different signals, cell differentiation and numerous influencing factors. However, recent studies suggest that two known mechanisms of epigenesis, i.e.gene inactivation by methylation in promoter region and changes to an inactive chromatin by histone deacetylation, seem to be the best candidate mechanisms for inactivation of p21( WAF1). To date, almost no coding region p21(WAF1) mutations have been found in tumor cells, despite extensive screening of hundreds of various tumors. Hypermethylation of the p21(WAF1) promoter region may represent an alternative mechanism by which the p21(WAF1/CIP1) gene can be inactivated. The reduction of cellular DNMT protein levels also induces a corresponding rapid increase in the cell cycle regulator p21(WAF1) protein demonstrating a regulatory link between DNMT and p21(WAF1) which is independent of methylation of DNA. Both histone hyperacetylation and hypoacetylation appear to be important in the carcinoma process, and induction of the p21(WAF1) gene by histone hyperacetylation may be a mechanism by which dietary fiber prevents carcinogenesis. Here, we review the influence of histone acetylation and DNA methylation on p21(WAF1) transcription, and affection of pathways or factors associated such as p 53, E2A, Sp1 as well as several histone deacetylation inhibitors.

Genome-wide identification and characterization of basic helix-loop-helix transcription factors in Spodoptera litura upon pathogen infection

Basic helix-loop-helix(bHLH)transcription factors play an important role in a wide range of metabolic and developmental processes in eukaryotes,and bHLH proteins also participate in immune responses,especially in plants.However,their roles in insects upon entomopathogen infection are unknown.In this study,54 bHLH genes in 41 families were identified in a polyphagous pest,Spodoptera litura,including a new bHLH gene in group B,which is specifically present in Lepidoptera and was thus named Lep.The conserved amino acids in the bHLH domain,structural architecture,and chromosomal distribution of bHLH genes in S.litura were analyzed.The bHLH genes in Plutella xylostella and Apis mellifera were also updated,and genome-wide comparison and phylogenetic analysis of bHLH members in 5 holometabolous insects were performed.The expression profiles of S.litura bHLH(SlbHLH)genes in 3 tissues at different developmental stages and their responses to S.litura nucleopolyhedrovirus(SpltNPV),Nomuraea rileyi(Nr),and Bacillus thuringiensis(Bt)infection were investigated.More SlbHLHs in group B were expressed and differentially expressed during pathogen infections,and SlbHLHs tended to be downregulated in the midgut of S.litura larvae after B.thuringiensis treatment.Our study provides an overview of bHLH family members in S.litura and their responses to different pathogens used for pest biocontrol.These findings on bHLH members may contribute to uncovering the mechanism of host-pathogen interaction.

bHLH genes polymorphisms and their association with growth traits in the Pacifi c oyster Crassostrea gigas

The basic helix-loop-helix(bHLH)genes encode a large superfamily of transcription factors in the Pacific oyster(Crassostrea gigas),and play a very important role in regulation of growth and development.To investigate the oyster growth traits and the associations with bHLH genes variations,we analyzed the gene polymorphisms-traits association in a wild population,and confirmed the results in another independent wild population by targeted gene re-sequencing and SNPshot analysis.After screening the single nucleotide polymorphisms(SNPs)in three candidate genes of the bHLH family(88 bHLH genes in two wild oyster populations in total),we identified the allele CgLoblHLH4-T/G located in the exon of the CgLoblHLH4 gene.This allele is a non-synonymous mutation(Phe/Leu)with an extremely significant association with shell width(P<0.01)and allele G is beneficial to shell width.This SNP on the CgLoblHLH4 gene might have a potential value as a genetic marker of growth traits that could be used in breeding in C.gigas in the future.

中国民间义虎报恩型故事的形态分析

中国民间义虎报恩型故事,主要讲述的是人解救了受困的老虎以后,受到老虎以猎物或义举回报的故事。借鉴刘魁立的"民间故事生命树"的研究方法,从结构形态学的角度分析故事的类型变体,依据17 个义虎故事文本,可分析出7 个不同的故事类型变体。据此可绘制故事生命树的结构图,并可进一步找出义虎报恩型故事的情节基干即"虎落难→人救虎→虎报恩",与核心母题即"虎报恩"。

度量故事:情节类型、情节基干与核心序列

在完成划分情节类型和编制索引之后,如何推进形态研究,是当前民间故事领域的困局。本文重新思考情节类型这一概念的逻辑前提,设置几种基础性概念工具,兼及情节、功能两个层面,对故事之间的相似、重复与关联进行度量。在归纳民间故事情节类型的诸种方案之中,刘魁立“以情节基干为中心”的方法是具有规范操作性的尝试。这一方法是基于文本形态的平行比较,然而比较的前提则未被明确指出:一种情节类型的结构对应于一个核心序列。情节基干应是一个自足的故事,它不仅是所有文本在情节层的共有情节,而且在功能层的对应结构必须是一个核心序列或多个核心序列的衔接,呈现出完整的叙述逻辑。由若干故事文本叠加而成的生命树,其重合部分含以下三种情况:第一,重合部分只应用相同的情节链,它的结构不足以构成一个完整的核心序列,则所有文本并无共享的情节基干,自然也不属于同一情节类型;第二,重合部分不仅表现为相同的情节链,而且其在功能层的结构对应于一个完整的核心序列,多则文本在该情节基干上的段落可被视为同一情节类型;第三,重合部分在功能层对应于多个核心序列,则情节基干可能是同一情节类型的重复应用,或多个不同情节类型的衔接。用以区分情节类型的情节基干等参数取决于故事材料样本,一旦样本变化,须重新提取情节基干,已有的分类体系可能被推翻,情节类型会随着样本的更新不断移动叙事重心。以有限样本归纳的情节类型并非一成不变,而是开放、无本质的。

设计基础造型课程教学思考

设计基础造型课程对基础理论的研究,为设计基础教学提供了较为系统的认识论与方法论,成为设计专业教学内在主线,是造型设计的核心课程,是培养学生最基本设计观念和设计意识的最根本环节。在现有的教学过程中,设计基础造型教学存有明显的弊病,作为专业课重要基础的地位没有能够得以彰显。我们在教学过程中,在教学理念、模式、内容上需要认真研究、探索、改革以适应学习专业的需要,减少弯路和无用功。

Effect of endothelial PAS domain protein 1 and hypoxia inducible factor 1~ on vascular endothelial growth factor expression in human pancreatic carcinoma

Background Transcription factors hypoxia inducible factor 1α （HIF 1α） and endothelial PAS domain protein 1 （EPAS1） promote the transcription of vascular endothelial growth factor （VEGF）. VEGF enhances angiogenesis and vascular permeability of tumours, which promotes tumour growth and facilitates entry of cancer cells into blood circulation and metastasizing. This study examined whether HIF 1α and EPAS1 stimulated angiogenesis through activation of VEGF in human pancreatic carcinoma. Methods Specimens from pancreatic carcinoma and healthy parts of same pancreas were taken from 60 patients. Real time quantitative reverse transcription polymerase chain reaction estimated expression of HIF 1α, EPAS1, and VEGF mRNAs. Western blotting and immunohistochemical, streptavidin peroxidase method assessed expression of HIF 1α, EPAS1, and VEGF proteins. Microvessel density （MVD） was assessed. Results Highly significant increases in expression of EPAS1, VEGF, and MVD were found in pancreatic carcinoma tissue but not in normal pancreatic tissue： VEGF at mRNA and protein levels （t=17.32, P=-0.0001; t=98.41, P=0.0001）; EPAS1 protein level （t=22.51, P=0.0001）. Expression of HIF la was similar in pancreatic carcinoma and normal pancreatic tissues at both mRNA and protein levels. Significant correlations were observed between EPAS1 and VEGF （r=0.736, P=0.0041）, between VEGF and MVD （r=0.858, P=0.0001）, and between EPAS1 and MVD （r=0.641, P=0.0003）. No significant correlations were observed between HIF la and VEGF, or between HIF 1α and MVD. MVD and expression of EPAS1 and VEGF were significantly related with TNM staging, so was EPASI and VEGF with size of tumour. Conclusions EPAS1 and VEGF, but not HIFla, are overexpressed in pancreatic carcinoma. The expression of EPAS1 is correlated with that of VEGF and MVD. EPAS1 may be involved in the angiogenesis of pancreatic carcinoma by upregulating the expression of VEGE Targeting EPAS1 may be a new method of antiangiogenic tumour therapy for pancreatic carcinoma.

The jasmonate-induced bHLH gene SlJIG functions in terpene biosynthesis and resistance to insects and fungus

Jasmonic acid(JA)is a key regulator of plant defense responses.Although the transcription factor MYC2,the master regulator of the JA signaling pathway,orchestrates a hierarchical transcriptional cascade that regulates the JA responses,only a few transcriptional regulators involved in this cascade have been described.Here,we identified the basic helix-loop-helix(bHLH)transcription factor gene in tomato(Solanum lycopersicum),METHYL JASMONATE(MeJA)-INDUCED GENE(SlJIG),the expression of which was strongly induced by MeJA treatment.Genetic and molecular biology experiments revealed that SlJIG is a direct target of MYC2.SlJIG knockout plants generated by gene editing had lower terpene contents than the wild type from the lower expression of TERPENE SYNTHASE(TPS)genes,rendering them more appealing to cotton bollworm(Helicoverpa armigera).Moreover,SlJIG knockouts exhibited weaker JA-mediated induction of TPSs,suggesting that SlJIG may participate in JA-induced terpene biosynthesis.Knocking out SlJIG also resulted in attenuated expression of JA-responsive defense genes,which may contribute to the observed lower resistance to cotton bollworm and to the fungus Botrytis cinerea.We conclude that SlJIG is a direct target of MYC2,forms a MYC2-SlJIG module,and functions in terpene biosynthesis and resistance against cotton bollworm and B.cinerea.

Further Analysis of the Function of AtBHLH29 in Regulating the Iron Uptake Process in Arabidopsis thaliana

Using T-DNA insertion and chemical mutants, two recent studies have shown that AtBHLH29, encoding a putative basic helix-loop-helix （BHLH） protein, is involved in regulating the iron uptake process in Arabidopsis thaliana. Herein, we report that RNA interference （RNAi） mutants can be used to reveal more accurately the genetic function of AtBHLH29. We compared the iron deficiency responses of seven RNAi strains that contained decreasing amounts of AtBHLH29transcripts. Under high iron conditions （50 μmol/L iron）, only in the most severe RNAi strains （R101, R111, and R119） was plant growth significantly retarded. However, these mutants could still survive and produce seeds. This suggests that the function of AtBHLH29 is beneficial, but not absolutely essential, to plant growth when iron supply is not limiting. Under low iron conditions （less than 10 μmol/L iron）, the Rlll and R119 strains died prematurely, demonstrating that AtBHLH29 is absolutely necessary for plant survival when iron supply is restricted. The transcription of AtBHLH29 was essential for the expression of AtFRO2 （encoding the ferric chelate reductase）. In contrast, the expression of AtlRT1 （encoding the high-affinity iron transporter） was not so strongly dependent upon the transcription of AtBHLH29. By transient expression, we found that the AtBHLH29-GUS fusion protein was targeted specifically to the nucleus in plant cells. Interestingly, the nuclear localization of AtBHLH29-GUS was abolished when the four consecutive arginine residues located in the basic region of the putative AtBHLH29 protein were replaced by alanine residues by mutagenesis. The implications of our findings on further studies of the mechanism underlying the function of AtBHLH29 are discussed.

Cryptochrome 1 Inhibits Shoot Branching by Repressing the Self-Activated Transciption Loop of PIF4 in Arabidopsis

Cryptochrome 1(CRY1)is an important light receptor essential for de-etiolation of Arabidopsis seedlings.However,its function in regulating plant architecture remains unclear.Here,we show that mutation in CRY1 resulted in increased branching of Arabidopsis plants.To investigate the underlying mechanism,we analyzed the expression profiles of branching-related genes and found that the mRNA levels of Phytochrome Interaction Factor 4(PIF4)and PIF5 are significantly increased in the cry1 mutant.Genetic analysis showed that the pif4 or pif4pif5 mutant is epistatic to the cry1 mutant,and overexpression of PIF4 conferred increased branching.Moreover,we demonstrated that PIF4 proteins physically associate with the G-box motif within the PIF4 promoter to form a self-activated transcriptional feedback loop,while CRY1 represses this process in response to blue light.Taken together,this study suggests that the CRY1–PIF4 module regulates gene expression via forming a regulatory loop and shoot branching in response to ambient light conditions.

Novel Nuclear Protein ALC-INTERACTING PROTEIN1 is Expressed in Vascular and Mesocarp Cells in Arabidopsis

Pod shattering is an agronomical trait that is a result of the coordinated action of cell differentiation and separation. In Arabidopsis, pod shattering is controlled by a complex genetic network in which ALCATRAZ （ALC）, a member of the basic helix-loop-helix family, is critical for cell separation during fruit dehiscence. Herein, we report the identification of ALC-INTERACTING PROTEIN1 （ACI1） via the yeast two-hybrid screen. ACll encodes a nuclear protein with a lysine-rich domain and a C-terminal serine-rich domain. ACI1 is mainly expressed in the vascular system throughout the plant and mesocarp of the valve in siliques. Our data showed that ACI1 interacts strongly with the N-terminal portion of ALC in yeast cells and in plant cells in the nucleus as demonstrated by bimolecular fluorescence complementaUon assay. Both ACI1 and ALC share an overlapping expression pattern, suggesting that they likely function together in planta. However, no detectable phenotype was found in plants with reduced ACll expression by RNA interference technology, suggesting that ACll may be redundant. Taken together, these data indicate that ALC may interact with ACI1 and its homologs to control cell separation during fruit dehiscence in Arabidopsis.