Hypothalamic-Pituitary-Gonadal(HPG)Axis and Transcriptional Regulatory Elements Regulate piwil2 Gene Expression During Gametogenesis and Gonadal Development in Japanese Flounder(Paralichthys olivaceus)

The P-element induced wimpy testis(Piwi)proteins,which are associated with PIWI-interacting RNAs(piRNAs),play important roles in meiosis,germ cell division,and germline maintenance.In this study,we identified and characterized the Paralichthys olivaceus piwil2 gene,a constituent factor of the piRNA pathways involved in the biogenesis of reproductive development.The biological analysis indicated that piwil2,which contains PAZ and PIWI domains,was highly conserved between teleosts and tetrapods.The piwil2 distribution profile in different tissues confirmed a sexually dimorphic expression pattern,with a higher expression level in testis.In situ hybridization demonstrated that piwil2 was expressed in the oogonia and oocytes of the ovaries as well as in the Sertoli cells and spermatocytes of the testes.Gene piwil2 showed a maternally inherited expression pattern during embryonic development,and was highly expressed during the early embryonic development.Different luciferase reporters were constructed to determine the transcriptional regulatory mechanisms of piwil2.The piwil2 core promoter region was located at−360 bp to−60 bp.Furthermore,some representative sex hormones,including human chorionic gonadotropin,17α-methyltestosterone,and estradiol-17βhad distinct regulatory effects on piwil2.In a summery,these results indicate that piwil2,regulated by sex hormones and transcriptional elements,has vital functions in the reproductive cycle and gonadal development.

Regulatory elements and transcriptional control of chicken vasa homologue(CVH)promoter in chicken primordial germ cells

Background： Primordial germ cells（PGCs）, the precursors of functional gametes, have distinct characteristics and exhibit several unique molecular mechanisms to maintain pluripotency and germness in comparison to somatic cells. They express germ cel-specific RNA binding proteins（RBPs） by modulating tissue-specific cis-and trans-regulatory elements. Studies on gene structures of chicken vasa homologue（CVH）, a chicken RNA binding protein, involved in temporal and spatial regulation are thus important not only for understanding the molecular mechanisms that regulate germ cel fate, but also for practical applications of primordial germ cells. However, very limited studies are available on regulatory elements that control germ cel-specific expression in chicken. Therefore, we investigated the intricate regulatory mechanism（s） that governs transcriptional control of CVH.Results： We constructed green fluorescence protein（GFP） or luciferase reporter vectors containing the various 5′ flanking regions of CVH gene. From the 5′ deletion and fragmented assays in chicken PGCs, we have identified a CVH promoter that locates at-316 to ＋275 base pair fragment with the highest luciferase activity. Additional y, we confirmed for the first time that the 5′ untranslated region（UTR） containing intron 1 is required for promoter activity of the CVH gene in chicken PGCs. Furthermore, using a transcription factor binding prediction, transcriptome analysis and siR NA-mediated knockdown,we have identified that a set of transcription factors play a role in the PGC-specific CVH gene expression.Conclusions： These results demonstrate that cis-elements and transcription factors localizing in the 5′ flanking region including the 5′ UTR and an intron are important for transcriptional regulation of the CVH gene in chicken PGCs. Final y,this information wil contribute to research studies in areas of reproductive biology, constructing of germ cel-specific synthetic promoter for tracing primordial germ cells as wel as understanding the transcriptional regulation for maintaining germness in PGCs.

Preliminary identification and analysis of point mutations corre lated with response to interferon-α in hepatitis B virus post-transcriptional regulatory elements

Background It is still unclear whether viral genetic variability influences response to interferon(IFN) α treatment Recent reports suggest that IFN α effects may be associated with hepatitis B virus(HBV) post transcriptional regulation This study was designed to explore the heterogeneity of HBV post transcriptional regulatory elements (HPRE) and the relationship between the diversity of HPRE and the response to IFN α treatment Methods The HPRE sequences from 31 Chinese patients infected with HBV were determined by directly sequencing of polymerase chain reaction (PCR) product, and comparing them to those from Caucasian patients Subsequently, eukaryotic expression vectors containing HPRE at various points were constructed and transfected into HepG2 cells, which were then exposed to recombinant human cytokines Results The T to C point mutation at nt 1504 and the C to T (G) at nt 1508 in HPRE were found in 21 and 19 patients with chronic hepatitis B, respectively; the C to T point mutation at nt 1509 was found in 17 patients These point mutations did not exist in the HPRE of the Caucasian patients The activity of the CAT gene obviously increased in the case of T to C point mutation at nt 1504, but did not change in the case of the C to T (G) mutations at nt 1508 and 1509 The activity of the CAT gene at these point mutations of HPRE could be inhibited by IFN α/γ and tumor necrosis factor (TNF) α except for the point mutations at nt 1508 of HPRE which may escape the suppression role of IFN α on HPRE Conclusions There are point mutations between the HPRE of Chinese and Caucasian HBV patients, which might be correlated with response to IFN α The variation of HPRE might affect the function of HPRE and influence the regulative function of IFN α other than that of IFN γ or TNF α on HPRE

Stochastic resonance induced by a multiplicative periodic signal in the gene transcriptional regulatory system with correlated noises

This paper investigates the stochastic resonance （SR） induced by a multiplicative periodic signal in the gene transcriptional regulatory system with correlated noises. The expression of the signal-to-noise ratio （SNR） is derived. The results indicate that the existence of a maximum in SNR vs. the additive noise intensity α the multiplicative noise intensity D and the cross-correlated noise intensity λ is the identifying characteristic of the SR phenomenon and there is a critical phenomenon in the SNR as a function of λ, i.e., for the case of smaller values of noise intensity （α or D）, the SNR decreases as λ increases; however, for the case of larger values of noise intensity （α or D）, the SNR increases as λ increases.

Stochastic resonance in the gene transcriptional regulatory system subjected to noises

We have investigated in the adiabatic limit the phenomenon of stochastic resonance in the gene transcriptional regulatory system subjected to an additive noise, a multiplicative noise, and a weakly periodic signal. Using the general two-state approach for the asymmetry system, the analytic expression of signal-to-noise ratio is obtained. The effects of the additive noise intensity a, the multiplicative noise intensity D and the amplitude of input periodic signal A on the signal-to-noise ratio are analysed by numerical calculation. It is found that the existence of a maximum in the RSNR a and RSNR D plots is the identifying characteristic of the stochastic resonance phenomenon in the weakened noise intensity region. The stochastic resonance phenomena are restrained with increasing a and D, and enhanced with increasing A.

Identifying changes in punitive transcriptional factor binding sites from regulatory single nucleotide polymorphisms that are significantly associated with disease or sickness

AIM To identify punitive transcriptional factor binding sites(TFBS) from regulatory single nucleotide polymorphisms(rS NPs) that are significantly associated with disease.METHODS The genome-wide association studies have provided us with nearly 6500 disease or trait-predisposing SNPs where 93% are located within non-coding regions such as gene regulatory or intergenic areas of the genome. In the regulatory region of a gene, a SNP can change the DNA sequence of a transcriptional factor(TF) motif and in turn may affect the process of gene regulation. SNP changes that affect gene expression and impact gene regulatory sequences such as promoters, enhancers, and silencers are known as rS NPs. Computational tools can be used to identify unique punitive TFBS created by rS NPs that are associated with disease or sickness. Computational analysis was used to identify punitive TFBS generated by the alleles of these rS NPs.RESULTS r SNPs within nine genes that have been significantly associated with disease or sickness were used to illustrate the tremendous diversity of punitive unique TFBS that can be generated by their alleles. The genes studied are the adrenergic, beta, receptor kinase 1, the v-akt murine thymoma viral oncogene homolog 3, the activating transcription factor 3, the type 2 demodkinase gene, the endothetal Per-Arnt-Sim domain protein 1, the lysosomal acid lipase A, the signal Transducer and Activator of Transcription 4, the thromboxane A2 receptor and the vascular endothelial growth factor A. From this sampling of SNPs among the nine genes, there are 73 potential unique TFBS generated by the common alleles comparedto 124 generated by the minor alleles indicating the tremendous diversity of potential TFs that are capable of regulating these genes.CONCLUSION From the diversity of unique punitive binding sites for TFs, it was found that some TFs play a role in the disease or sickness being studied.

Nonchaoticity of Ordinary Differential Equations Describing Autonomous Transcriptional Regulatory Circuits

Gene transcriptional regulation （TR） processes are often described by coupled nonlinear ordinary differential equations （ODEs）. When the dimension of TR circuits is high （e.g. n≥3） the motions of the corresponding ODEs may, very probably, show self-sustained oscillations and chaos. On the other hand, chaoticity may be harmful for the normal biological functions of TR processes. In this letter we numerically study the dynamics of 3-gene TR ODEs in great detail, and investigate many 4-, 5-, and lO-gene TR systems by randomly choosing figures and parameters in the conventionally accepted ranges. And we find that oscillations are very seldom and no chaotic motion is observed, even if the dimension of systems is sufficiently high （n≥3）. It is argued that the observation of nonchaoticity of these ODEs agrees with normal functions of actual TR processes.

Detection and characterization of regulatory elements using probabilistic conditional random field and hidden Markov models

By altering the electrostatic charge of histones or providing binding sites to protein recognition molecules, Chromatin marks have been proposed to regulate gene expression, a property that has motivated researchers to link these marks to cis-regulatory elements. With the help of next generation sequencing technologies, we can now correlate one specific chromatin mark with regulatory elements (e.g. enhancers or promoters) and also build tools, such as hidden Markov models, to gain insight into mark combinations. However, hidden Markov models have limitation for their character of generative models and assume that a current observation depends only on a current hidden state in the chain. Here, we employed two graphical probabilistic models, namely the linear conditional random field model and multivariate hidden Markov model, to mark gene regions with different states based on recurrent and spatially coherent character of these eight marks. Both models revealed chromatin states that may correspond to enhancers and promoters, transcribed regions, transcriptional elongation, and low-signal regions. We also found that the linear conditional random field model was more effective than the hidden Markov model in recognizing regulatory elements, such as promoter-, enhancer-, and transcriptional elongation-associated regions, which gives us a better choice.

The abscisic acid-responsive transcriptional regulatory module CsERF110-CsERF53 orchestrates citrus fruit coloration

Carotenoid biosynthesis is closely associated with abscisic acid(ABA)during the ripening process of non-climacteric fruits,but the regulatory mechanism that links ABA signaling to carotenoid metabolism remains largely unclear.Here,we identified two master regulators of ABA-mediated citrus fruit coloration,CsERF110 and CsERF53,which activate the expression of carotenoid metabolism genes(CsGGPPS,CsPSY,CsPDS,CsCRTISO,CsLCYB2,CsLCYE,CsHYD,CsZEP,and CsNCED2)to facilitate carotenoid accumulation.Further investigations showed that CsERF110 not only activates the expression of CsERF53 by binding to its promoter but also interacts with CsERF53 to form the transcriptional regulatory module CsERF110-CsERF53.We also discovered a positive feedback regulatory loop between the ABA signal and carotenoid metabolism regulated by the transcriptional regulatory module CsERF110-CsERF53.Our results reveal that the CsERF110-CsERF53 module responds to ABA signaling,thereby orchestrating citrus fruit coloration.Considering the importance of carotenoid content for citrus and many other carotenoid-rich crops,the revelation of molecular mechanisms that underlie ABA-mediated carotenoid biosynthesis in plants will facilitate the development of transgenic/gene-editing approaches,further contributing to improving the quality of citrus and other carotenoid-rich crops.

Differential transcriptional regulation of the NANOG gene in chicken primordial germ cells and embryonic stem cells

Background:NANOG is a core transcription factor(TF)in embryonic stem cells(ESCs)and primordial germ cells(PGCs).Regulation of the NANOG gene by TFs,epigenetic factors,and autoregulatory factors is well characterized in ESCs,and transcriptional regulation of NANOG is well established in these cells.Although NANOG plays a key role in germ cells,the molecular mechanism underlying its transcriptional regulation in PGCs has not been studied.Therefore,we investigated the mechanism that regulates transcription of the chicken NANOG(cNANOG)gene in PGCs and ESCs.Results:We first identified the transcription start site of cNANOG by 5′-rapid amplification of cDNA ends PCR analysis.Then,we measured the promoter activity of various 5′flanking regions of cNANOG in chicken PGCs and ESCs using the luciferase reporter assay.cNANOG expression required transcriptional regulatory elements,which were positively regulated by POU5F3(OCT4)and SOX2 and negatively regulated by TP53 in PGCs.The proximal region of the cNANOG promoter contains a positive transcriptional regulatory element(CCAAT/enhancer-binding protein(CEBP)-binding site)in ESCs.Furthermore,small interfering RNA-mediated knockdown demonstrated that POU5F3,SOX2,and CEBP played a role in cell type-specific transcription of cNANOG.Conclusions:We show for the first time that different trans-regulatory elements control transcription of cNANOG in a cell type-specific manner.This finding might help to elucidate the mechanism that regulates cNANOG expression in PGCs and ESCs.

Single-cell RNA-Seq reveals transcriptional regulatory networks directing the development of mouse maxillary prominence

During vertebrate embryonic development,neural crest-derived ectomesenchyme within the maxillary prominences undergoes precisely coordinated proliferation and differentiation to give rise to diverse craniofacial structures,such as tooth and palate.However,the transcriptional regulatory networks underpinning such an intricate process have not been fully elucidated.Here,we perform single-cell RNA-Seq to comprehensively characterize the transcriptional dynamics during mouse maxillary development from embryonic day(E)10.5eE14.5.Our single-cell transcriptome atlas of~28,000 cells uncovers mesenchymal cell populations representing distinct differentiating states and reveals their developmental trajectory,suggesting that the segregation of dental from the palatal mesenchyme occurs at E11.5.Moreover,we identify a series of key transcription factors(TFs)associated with mesenchymal fate transitions and deduce the gene regulatory networks directed by these TFs.Collectively,our study provides important resources and insights for achieving a systems-level understanding of craniofacial morphogenesis and abnormality.

Pig H3K4me3,H3K27ac,and gene expression profiles reveal reproductive tissue-specific activity of transposable elements

Regulatory sequences and transposable elements(TEs)account for a large proportion of the genomic sequences of species;however,their roles in gene transcription,especially tissue-specific expression,remain largely unknown.Pigs serve as an excellent animal model for studying genomic sequence biology due to the extensive diversity among their wild and domesticated populations.Here,we conducted an integrated analysis using H3K27ac ChIP-seq,H3K4me3 ChIP-seq,and RNA-seq data from 10 different tissues of seven fetuses and eight closely related adult pigs.We aimed to annotate the regulatory elements and TEs to elucidate their associations with histone modifications and mRNA expression across different tissues and developmental stages.Based on correlation analysis between mRNA expression and H3K27ac and H3K4me3 peak activity,results indicated that H3K27ac exhibited stronger associations with gene expression than H3K4me3.Furthermore,1.45%of TEs overlapped with either the H3K27ac or H3K4me3 peaks,with the majority displaying tissue-specific activity.Notably,a TE subfamily(LTR4C_SS),containing binding motifs for SIX1 and SIX4,showed specific enrichment in the H3K27ac peaks of the adult and fetal ovaries.RNA-seq analysis also revealed widespread expression of TEs in the exons or promoters of genes,including 4688 TE-containing transcripts with distinct development stage-specific and tissue-specific expression.Of note,1967 TE-containing transcripts were enriched in the testes.We identified a long terminal repeat(LTR),MLT1F1,acting as a testis-specific alternative promoter in SRPK2(a cell cycle-related protein kinase)in our pig dataset.This element was also conserved in humans and mice,suggesting either an ancient integration of TEs in genes specifically expressed in the testes or parallel evolutionary patterns.Collectively,our findings demonstrate that TEs are deeply embedded in the genome and exhibit important tissue-specific biological functions,particularly in the reproductive organs.

A step-by-step protocol for formaldehyde-assisted isolation of regulatory elements from Arabidopsis thaliana

The control of gene expression by transcriptional regulators and other types of functional y relevant DNA transactions such as chromatin remodeling and replication underlie a vast spectrum of biological processes in al organisms. DNA transactions require the control ed interaction of proteins with DNA sequence motifs which are often located in nucleosome-depleted regions （NDRs） of the chromatin. Formaldehyde-assisted isolation of regulatory elements （FAIRE） has been established as an easy-to-implement method for the isolation of NDRs from a number of eukaryotic organisms, and it has been successful y employed for the discovery of new regulatory segments in genomic DNA from, for example, yeast, Drosophila, and humans. Until today, however, FAIRE has only rarely been employed in plant research and currently no detailed FAIRE protocol for plants has been published. Here, we provide a step-by-step FAIRE protocol for NDR discovery in Arabidopsis thaliana. We demonstrate that NDRs isolated from plant chromatin are readily amenable to quantitative polymerase chain reaction and next-generation sequencing. Only minor modification of the FAIRE protocol wil be needed to adapt it to other plants, thus facilitating the global inventory of regulatory regions across species.

Large-scale determination and characterization of cell type-specific regulatory elements in the human genorne

Histone modifications have been widely elucidated to play vital roles in gene regulation and cell identity. The Roadmap Epigenomics Consortium generated a reference catalog of several key histone modifications across 〉lOOs of human cell types and tissues. Decoding these epJgenomes into functional regulatory elements is a challenging task in computational biology. To this end, we adopted a differential chromatin modification analysis framework to comprehensively determine and characterize cell type-specific regulatory elements （CSREs） and their histone modification codes in the human epigenomes of five histone modifications across 127 tissues or cell types. The CSREs show significant relevance with cell type-specific biological functions and diseases and cell identity. Clustering of CSREs with their specificity signals reveals distinct histone codes, demonstrating the diversity of functional roles of CSREs within the same cell or tissue. Last but not least, dynamics of CSREs from close cell types or tissues can give a detailed view of developmental processes such as normal tissue development and cancer occurrence.

Transcriptional Regulatory Networks Activated by PI3K and ERK Transduced Growth Signals in Human Glioblastoma Cells

Determining how cells regulate their transcriptional response toextracellular signals is key to the understanding of complex eukaryotic systems. This study wasinitiated with the goals of furthering the study of mammalian transcriptional regulation andanalyzing the relative benefits of related computational methodologies. One dataset available forsuch an analysis involved gene expression profiling of the early growth factor response to plateletderived growth factor (PDGF) in a human glioblastoma cell line; this study differentiated geneswhose expression was regulated by signaling through the phosphoinositide-3-kinase (PI3K) versus theextracellular-signal regulated kinase (ERK) pathways. We have compared the inferred transcriptionfactors from this previous study with additional predictions of regulatory transcription factorsusing two alternative promoter sequence analysis techniques. This comparative analysis, in which thealgorithms predict overlapping, although not identical, sets of factors, argues for meticulousbenchmarking of promoter sequence analysis methods to determine the positive and negative attributesthat contribute to their varying results. Finally, we inferred transcriptional regulatory networksderiving from various signaling pathways using the CARRIE program suite. These networks not onlyincluded previously described transcriptional features of the response to growth signals, but alsopredicted new regulatory features for the propagation and modulation of the growth signal.

The future of genome-scale modeling of yeast through integration of a transcriptional regulatory network

Metabolism is regulated at multiple levels in response to the changes of internal or external conditions. Transcriptional regulation plays an important role in regulating many metabolic reactions by altering the concentrations of metabolic enzymes. Thus, integration of the transcriptional regulatory information is necessary to improve the accuracy and predictive ability of metabolic models. Here we review the strategies for the reconstruction of a transcriptional regulatory network （TRN） for yeast and the integration of such a reconstruction into a flux balance analysis-based metabolic model. While many large-scale TRN reconstructions have been reported for yeast, these reconstructions still need to be improved regarding the functionality and dynamic property of the regulatory interactions. In addition, mathematical modeling approaches need to be further developed to efficiently integrate transcriptional regulatory interactions to genome-scale metabolic models in a quantitative manner.

The transcriptional regulators of virulence for Pseudomonas aeruginosa:Therapeutic opportunity and preventive potential of its clinical infections

In Pseudomonas aeruginosa(P.aeruginosa),transcription factors(TFs)are important mediators in the genetic regulation of adaptability and pathogenicity to respond to multiple environmental stresses and host defences.The P.aeruginosa genome harbours 371 putative TFs;of these,about 70 have been shown to regulate virulence-associated phenotypes by binding to the promoters of their target genes.Over the past three decades,several techniques have been applied to identify TF binding sites on the P.aeruginosa genome,and an atlas of TF binding patterns has been mapped.The virulence-associated regulons of TFs show complex crosstalk in P.aeruginosa's regulatory network.In this review,we summarise the recent literature on TF regulatory networks involved in the quorum-sensing system,biofilm formation,pyocyanin synthesis,motility,the type III secretion system,the type VI secretion system,and oxidative stress responses.We discuss future perspectives that could provide insights and targets for preventing clinical infections caused by P.aeruginosa based on the global regulatory network of transcriptional regulators.

Cis-acting regulatory elements： from random screening to quantitative design

The cis-acting regulatory elements, e.g., promoters and ribosome binding sites （RBSs） with various desired properties, are building blocks widely used in synthetic biology for fine tuning gene expression. In the last decade, acquisition of a controllable regulatory element from a random library has been established and applied to control the protein expression and metabolic flux in different chassis cells. However, more rational strategies are still urgently needed to improve the efficiency and reduce the laborious screening and multifaceted characterizations. Building precise computational models that can predict the activity of regulatory elements and quantitatively design elements with desired strength have been demonstrated tremendous potentiality. Here, recent progress on construction of cis- acting regulatory element library and the quantitative predicting models for design of such elements are reviewed and discussed in detail.

A new method for splice site prediction based on the sequence patterns of splicing signals and regulatory elements

It is of significance for splice site prediction to develop novel algorithms that combine the sequence patterns of regulatory elements such as enhancers and silencers with the patterns of splicing signals.In this paper,a statistical model of splicing signals was built based on the entropy density profile(EDP) method,weight array method(WAM) and κ test;moreover,the model of splicing regulatory elements was developed by an unsupervised self-learning method to detect motifs associated with regulatory elements.With two models incorporated,a multi-level support vector machine(SVM) system was de-vised to perform ab initio prediction for splice sites originating from DNA sequence in eukaryotic ge-nome.Results of large scale tests on human genomic splice sites show that the new method achieves a comparative high performance in splice site prediction.The method is demonstrated to be with at least the same level of performance and usually better performance than the existing SpliceScan method based on modeling regulatory elements,and shown to have higher accuracies than the traditional methods with modeling splicing signals such as the GeneSplicer.In particular,the method has evident advantage over splice site prediction for the genes with lower GC content.

The contribution of cis-regulatory elements to head-to-head gene pairs' co-expression pattern

Transcription regulation is one of the most critical pipelines in biological process,in which cis-elements play the role as gene expression regulators.We attempt to deduce the principles underlying the co-expression of "head-to-head" gene pairs by analyzing activities or behaviors of the shared cis-elements.A network component analysis was performed to estimate the impact of cis-elements on gene promoters and their activities under different conditions.Our discoveries reveal how biological system uses those regulatory elements to control the expression pattern of "head-to-head" gene pairs and the whole transcription regulation system.