Upstream Regions Of Genes Research Articles

Soybean (Glycine max) WRINKLED1 transcription factor, GmWRI1a, positively regulates seed oil accumulation.

Soybean is the world's most important leguminous crop producing high-quality protein and oil. Elevating oil accumulation in soybean seed is always many researchers' goal. WRINKLED1 (WRI1) encodes a transcription factor of the APETALA2/ethylene responsive element-binding protein (AP2/EREBP) family that plays important roles during plant seed oil accumulation. In this study, we isolated and characterized three distinct orthologues of WRI1 in soybean (Glycine max) that display different organ-specific expression patterns, among which GmWRI1a was highly expressed in maturing soybean seed. Electrophoretic mobility shift assays and yeast one-hybrid experiments demonstrated that the GmWRI1a protein was capable of binding to AW-box, a conserved sequence in the proximal upstream regions of many genes involved in various steps of oil biosynthesis. Transgenic soybean seeds overexpressing GmWRI1a under the control of the seed-specific napin promoter showed the increased total oil and fatty acid content and the changed fatty acid composition. Furthermore, basing on the activated expressions in transgenic soybean seeds and existence of AW-box element in the promoter regions, direct downstream genes of GmWRI1a were identified, and their products were responsible for fatty acid production, elongation, desaturation and export from plastid. We conclude that GmWRI1a transcription factor can positively regulate oil accumulation in soybean seed by a complex gene expression network related to fatty acid biosynthesis.

Identification and comparative analysis of Brassica juncea pathogenesis-related genes in response to hormonal, biotic and abiotic stresses

Pathogenesis-related proteins (PRs) are the antimicrobial proteins which are commonly used as signatures of defense signaling pathways and systemic acquired resistance. However, in Brassica juncea most of the PR proteins have not been fully characterized and remains largely enigmatic. In this study, full-length cDNA sequences of SA (PR1, PR2, PR5) and JA (PR3, PR12 and PR13) marker genes were isolated from B. juncea and were named as BjPR proteins. BjPR proteins showed maximum identity with known PR proteins of Brassica species. Further, expression profiling of BjPR genes were investigated after hormonal, biotic and abiotic stresses. Pre-treatment with SA and JA stimulators downregulates each other signature genes suggesting an antagonistic relationship between SA and JA in B. juncea. After abscisic acid (ABA) treatment, SA signatures were downregulated while as JA signature genes were upregulated. During Erysiphe cruciferarum infection, SA- and JA-dependent BjPR genes showed distinct expression pattern both locally and systemically, thus suggesting the activation of SA- and JA-dependent signaling pathways. Further, expression of SA marker genes decreases while as JA-responsive genes increases during drought stress. Interestingly, both SA and JA signature genes were induced after salt stress. We also found that BjPR genes displayed ABA-independent gene expression pattern during abiotic stresses thus providing the evidence of SA/JA cross talk. Further, in silico analysis of the upstream regions (1.5 kb) of both SA and JA marker genes showed important cis-regulatory elements related to biotic, abiotic and hormonal stresses.

A Genome-Wide Identification of the WRKY Family Genes and a Survey of Potential WRKY Target Genes in Dendrobium officinale

The WRKY family, one of the largest families of transcription factors, plays important roles in the regulation of various biological processes, including growth, development and stress responses in plants. In the present study, 63 DoWRKY genes were identified from the Dendrobium officinale genome. These were classified into groups I, II, III and a non-group, each with 14, 28, 10 and 11 members, respectively. ABA-responsive, sulfur-responsive and low temperature-responsive elements were identified in the 1-k upstream regulatory region of DoWRKY genes. Subsequently, the expression of the 63 DoWRKY genes under cold stress was assessed, and the expression profiles of a large number of these genes were regulated by low temperature in roots and stems. To further understand the regulatory mechanism of DoWRKY genes in biological processes, potential WRKY target genes were investigated. Among them, most stress-related genes contained multiple W-box elements in their promoters. In addition, the genes involved in polysaccharide synthesis and hydrolysis contained W-box elements in their 1-k upstream regulatory regions, suggesting that DoWRKY genes may play a role in polysaccharide metabolism. These results provide a basis for investigating the function of WRKY genes and help to understand the downstream regulation network in plants within the Orchidaceae.

Correlation between desiccation stress response and epigenetic modifications of genes in Drosophila melanogaster: An example of environment-epigenome interaction

Animals from different phyla including arthropods tolerate water stress to different extent. This tolerance is accompanied by biochemical changes which in turn are due to transcriptional alteration. The changes in transcription can be an indirect effect on some of the genes, ensuing from the effect of stress on the regulators of transcription including epigenetic regulators. Within this paradigm, we investigated the correlation between stress response and epigenetic modification underlying gene expression modulation during desiccation stress in Canton-S. We report altered resistance of flies in desiccation stress for heterozygote mutants of PcG and TrxG members. Pc/+ mutant shows lower survival, while ash1/+ mutants show higher survival under desiccation stress as compared to Canton-S. We detect expression alteration in stress related genes as well the genes of the Polycomb and trithorax complex in Canton-S subjected to desiccation stress. Concomitant with this, there is an altered enrichment of H3K27me3 and H3K4me3 at the upstream regions of the stress responsive genes. The enrichment of activating mark, H3K4me3, is higher in non-stress condition. H3K27me3, the repressive mark, is more pronounced under stress condition, which in turn, can be correlated with the binding of Pc. Our results show that desiccation stress induces dynamic switching in expression and enrichment of PcG and TrxG in the upstream region of genes, which correlates with histone modifications. We provide evidence that epigenetic modulation could be one of the mechanisms to adapt to the desiccation stress in Drosophila. Thus, our study proposes the interaction of epigenome and environmental factors.

Diversity of cis-regulatory elements associated with auxin response in Arabidopsis thaliana.

The phytohormone auxin regulates virtually every developmental process in land plants. This regulation is mediated via de-repression of DNA-binding auxin response factors (ARFs). ARFs bind TGTC-containing auxin response cis-elements (AuxREs), but there is growing evidence that additional cis-elements occur in auxin-responsive regulatory regions. The repertoire of auxin-related cis-elements and their involvement in different modes of auxin response are not yet known. Here we analyze the enrichment of nucleotide hexamers in upstream regions of auxin-responsive genes associated with auxin up- or down-regulation, with early or late response, ARF-binding domains, and with different chromatin states. Intriguingly, hexamers potentially bound by basic helix-loop-helix (bHLH) and basic leucine zipper (bZIP) factors as well as a family of A/T-rich hexamers are more highly enriched in auxin-responsive regions than canonical TGTC-containing AuxREs. We classify and annotate the whole spectrum of enriched hexamers and discuss their patterns of enrichment related to different modes of auxin response.

A Comprehensive Analysis of Transcript-Supported De Novo Genes in Saccharomyces sensu stricto Yeasts.

Novel genes arising from random DNA sequences (de novo genes) have been suggested to be widespread in the genomes of different organisms. However, our knowledge about the origin and evolution of de novo genes is still limited. To systematically understand the general features of de novo genes, we established a robust pipeline to analyze >20,000 transcript-supported coding sequences (CDSs) from the budding yeast Saccharomyces cerevisiae. Our analysis pipeline combined phylogeny, synteny, and sequence alignment information to identify possible orthologs across 20 Saccharomycetaceae yeasts and discovered 4,340 S. cerevisiae-specific de novo genes and 8,871 S. sensu stricto-specific de novo genes. We further combine information on CDS positions and transcript structures to show that >65% of de novo genes arose from transcript isoforms of ancient genes, especially in the upstream and internal regions of ancient genes. Fourteen identified de novo genes with high transcript levels were chosen to verify their protein expressions. Ten of them, including eight transcript isoform-associated CDSs, showed translation signals and five proteins exhibited specific cytosolic localizations. Our results suggest that de novo genes frequently arise in the S. sensu stricto complex and have the potential to be quickly integrated into ancient cellular network.

Genome-wide identification, functional prediction, and evolutionary analysis of the R2R3-MYB superfamily in Brassica napus.

R2R3-MYB transcription factors (TFs) have been shown to play important roles in plants, including in development and in various stress conditions. Phylogenetic analysis showed the presence of 249 R2R3-MYB TFs in Brassica napus, called BnaR2R3-MYB TFs, clustered into 38 clades. BnaR2R3-MYB TFs were distributed on 19 chromosomes of B. napus. Sixteen gene clusters were identified. BnaR2R3-MYB TFs were characterized by motif prediction, gene structure analysis, and gene ontology. Evolutionary analysis revealed that BnaR2R3-MYB TFs are mainly formed as a result of whole-genome duplication. Orthologs and paralogs of BnaR2R3-MYB TFs were identified in B. napus, B. rapa, B. oleracea, and Arabidopsis thaliana using synteny-based methods. Purifying selection was pervasive within R2R3-MYB TFs. Kn/Ks values lower than 0.3 indicated that BnaR2R3-MYB TFs are being functionally converged. The role of gene conversion in the formation of BnaR2R3-MYB TFs was significant. Cis-regulatory elements in the upstream regions of BnaR2R3-MYB genes, miRNA targeting BnaR2R3MYB TFs, and post translational modifications were identified. Digital expression data revealed that BnaR2R3-MYB genes were highly expressed in the roots and under high salinity treatment after 24 h. BnaMYB21, BnaMYB141, and BnaMYB148 have been suggested for improving salt-tolerant B. napus. BnaR2R3-MYB genes were mostly up regulated on the 14th day post inoculation with Leptosphaeria biglobosa and L. maculan. BnaMYB150 is a candidate for increased tolerance to Leptospheria in B. napus.

Comparative study of gene structure in the upstream regulatory region of CDC10 and MYBPC1 genes in Iranian fat-tailed and nonfat-tailed sheep breeds

Marbling is one of the important factors among the characteristics influencing meat quality. Among the candidate genes, MYBPC1 and CDC10 genes are considered as functional genes, responsible for marbling. The present study was conducted to investigate the existence of polymorphism and domain variants in the upstream regulatory region of CDC10 and MYBPC1 genes in Iranian fat-tailed (Lori-Bakhtiari) and nonfat-tailed (Zel) sheep breeds. Blood samples were taken randomly from 300 sheep (150 samples per each breed). DNA extraction was performed by modified salting-out method. Fragments with the length of 291 and 372bp from the upstream regulatory region of MYBPC1 and CDC10 genes were amplified by specific primers and different allelic forms in both marker sites were detected by single-strand conformation polymorphism (SSCP) method. In order to verify observed banding patterns (genotypes) and also to investigate the presence of mutations in the sequence, one sample from each banding pattern was sequenced. MYBPC1 marker site showed monomorphic pattern, but the CDC10 was polymorph in both breeds. The allele A was the most frequent allele (0.90 and 0.77%) in Zel and Lori-Bakhtiari breeds respectively. Genotype frequencies of AA were the highest (83.33 and 59.71%) compared with that of BB genotypes (2.90 and 5.04%) in both Zel and Lori-Bakhtiari breeds respectively. Comparison between genotype frequencies by Chi-square test showed significant differences (P < 0.01) among the two studied breeds. The results of multiple sequence alignment revealed six single nucleotide polymorphisms (SNP) in the amplified fragment of CDC10 marker site. Further bioinformatics analysis revealed 16 motifs for this marker site. This study was the first effort for appraising these genes as a candidate genes affecting marbling in sheep. Therefore, supplementary research in these two studied breeds, as well as more studies on other components of these genes in respect to other sheep breeds, is recommended.

Mediator 1 Is Atherosclerosis Protective by Regulating Macrophage Polarization.

MED1 (mediator 1) interacts with transcription factors to regulate transcriptional machinery. The role of MED1 in macrophage biology and the relevant disease state remains to be investigated. To study the molecular mechanism by which MED1 regulates the M1/M2 phenotype switch of macrophage and the effect on atherosclerosis, we generated MED1/apolipoprotein E (ApoE) double-deficient (MED1ΔMac/ApoE-/-) mice and found that atherosclerosis was greater in MED1ΔMac/ApoE-/- mice than in MED1fl/fl/ApoE-/- littermates. The gene expression of M1 markers was increased and that of M2 markers decreased in both aortic wall and peritoneal macrophages from MED1ΔMac/ApoE-/- mice, whereas MED1 overexpression rectified the changes in M1/M2 expression. Moreover, LDLR (low-density lipoprotein receptor)-deficient mice received bone marrow from MED1ΔMac mice showed greater atherosclerosis. Mechanistically, MED1 ablation decreased the binding of PPARγ (peroxisome proliferator-activated receptor γ) and enrichment of H3K4me1 and H3K27ac to upstream region of M2 marker genes. Furthermore, interleukin 4 induction of PPARγ and MED1 increased the binding of PPARγ or MED1 to the PPAR response elements of M2 marker genes. Our data suggest that MED1 is required for the PPARγ-mediated M2 phenotype switch, with M2 marker genes induced but M1 marker genes suppressed. MED1 in macrophages has an antiatherosclerotic role via PPARγ-regulated transactivation.

Identification and positional distribution analysis of transcription factor binding sites for genes from the wheat fl-cDNA sequences.

The binding sites of transcription factors (TFs) in upstream DNA regions are called transcription factor binding sites (TFBSs). TFBSs are important elements for regulating gene expression. To date, there have been few studies on the profiles of TFBSs in plants. In total, 4,873 sequences with 5' upstream regions from 8530 wheat fl-cDNA sequences were used to predict TFBSs. We found 4572 TFBSs for the MADS TF family, which was twice as many as for bHLH (1951), B3 (1951), HB superfamily (1914), ERF (1820), and AP2/ERF (1725) TFs, and was approximately four times higher than the remaining TFBS types. The percentage of TFBSs and TF members showed a distinct distribution in different tissues. Overall, the distribution of TFBSs in the upstream regions of wheat fl-cDNA sequences had significant difference. Meanwhile, high frequencies of some types of TFBSs were found in specific regions in the upstream sequences. Both TFs and fl-cDNA with TFBSs predicted in the same tissues exhibited specific distribution preferences for regulating gene expression. The tissue-specific analysis of TFs and fl-cDNA with TFBSs provides useful information for functional research, and can be used to identify relationships between tissue-specific TFs and fl-cDNA with TFBSs. Moreover, the positional distribution of TFBSs indicates that some types of wheat TFBS have different positional distribution preferences in the upstream regions of genes.

In the hunt for genomic markers of metabolic resistance to pyrethroids in the mosquito Aedes aegypti: An integrated next-generation sequencing approach.

BackgroundThe capacity of Aedes mosquitoes to resist chemical insecticides threatens the control of major arbovirus diseases worldwide. Until alternative control tools are widely deployed, monitoring insecticide resistance levels and identifying resistance mechanisms in field mosquito populations is crucial for implementing appropriate management strategies. Metabolic resistance to pyrethroids is common in Aedes aegypti but the monitoring of the dynamics of resistant alleles is impeded by the lack of robust genomic markers.Methodology/Principal findingsIn an attempt to identify the genomic bases of metabolic resistance to deltamethrin, multiple resistant and susceptible populations originating from various continents were compared using both RNA-seq and a targeted DNA-seq approach focused on the upstream regions of detoxification genes. Multiple detoxification enzymes were over transcribed in resistant populations, frequently associated with an increase in their gene copy number. Targeted sequencing identified potential promoter variations associated with their over transcription. Non-synonymous variations affecting detoxification enzymes were also identified in resistant populations.Conclusion /SignificanceThis study not only confirmed the role of gene copy number variations as a frequent cause of the over expression of detoxification enzymes associated with insecticide resistance in Aedes aegypti but also identified novel genomic resistance markers potentially associated with their cis-regulation and modifications of their protein structure conformation. As for gene transcription data, polymorphism patterns were frequently conserved within regions but differed among continents confirming the selection of different resistance factors worldwide. Overall, this study paves the way of the identification of a comprehensive set of genomic markers for monitoring the spatio-temporal dynamics of the variety of insecticide resistance mechanisms in Aedes aegypti.

Insights into a key sulfite scavenger enzyme sulfite oxidase (SOX) gene in plants.

Sulfite oxidase (SOX) is a crucial molybdenum cofactor-containing enzyme in plants that re-oxidizes the sulfite back to sulfate in sulfite assimilation pathway. However, studies of this crucial enzyme are quite limited hence this work was attempted to understand the SOXs in four plant species namely, Arabidopsis thaliana, Solanum lycopersicum, Populus trichocarpa and Brachypodium distachyon. Herein studied SOX enzyme was characterized with both oxidoreductase molybdopterin binding and Mo-co oxidoreductase dimerization domains. The alignment and motif analyses revealed the highly conserved primary structure of SOXs. The phylogeny constructed with additional species demonstrated a clear divergence of monocots, dicots and lower plants. In addition, to further understand the phylogenetic relationship and make a functional inference, a structure-based phylogeny was constructed using normalized RMSD values in five superposed models from four modelled plant SOXs herein and one previously characterized chicken SOX structure. The plant and animal SOXs showed a clear divergence and also implicated their functional divergences. Based on tree topology, monocot B. distachyon appeared to be diverged from other dicots, pointing out a possible monocot-dicot split. The expression patterns of sulfite scavengers including SOX were differentially modulated under cold, heat, salt and high light stresses. Particularly, they tend to be up-regulated under high light and heat while being down-regulated under cold and salt stresses. The presence of cis-regulatory motifs associated with different stresses in upstream regions of SOX genes was thus justified. The protein-protein interaction network of AtSOX and network enrichment with gene ontology (GO) terms showed that most predicted proteins, including sulfite reductase, ATP sulfurylases and APS reductases were among prime enzymes involved in sulfite pathway. Finally, SOX-sulfite docked structures indicated that arginine residues particularly Arg374 is crucial for SOX-sulfite binding and additional two other residues such as Arg51 and Arg103 may be important for SOX-sulfite bindings in plants.

Genome-wide exploration of metal tolerance protein (MTP) genes in common wheat (Triticum aestivum): insights into metal homeostasis and biofortification.

Metal transport process in plants is a determinant of quality and quantity of the harvest. Although it is among the most important of staple crops, knowledge about genes that encode for membrane-bound metal transporters is scarce in wheat. Metal tolerance proteins (MTPs) are involved in trace metal homeostasis at the sub-cellular level, usually by providing metal efflux out of the cytosol. Here, by using various bioinformatics approaches, genes that encode for MTPs in the hexaploid wheat genome (Triticum aestivum, abbreviated as Ta) were identified and characterized. Based on the comparison with known rice MTPs, the wheat genome contained 20 MTP sequences; named as TaMTP1-8A, B and D. All TaMTPs contained a cation diffusion facilitator (CDF) family domain and most members harbored a zinc transporter dimerization domain. Based on motif, phylogeny and alignment analysis, A, B and D genomes of TaMTP3-7 sequences demonstrated higher homology compared to TaMTP1, 2 and 8. With reference to their rice orthologs, TaMTP1s and TaMTP8s belonged to Zn-CDFs, TaMTP2s to Fe/Zn-CDFs and TaMTP3-7s to Mn-CDFs. Upstream regions of TaMTP genes included diverse cis-regulatory motifs, indicating regulation by developmental stage, tissue type and stresses. A scan of the coding sequences of 20 TaMTPs against published miRNAs predicted a total of 14 potential miRNAs, mainly targeting the members of most diverged groups. Expression analysis showed that several TaMTPs were temporally and spatially regulated during the developmental time-course. In grains, MTPs were preferentially expressed in the aleurone layer, which is known as a reservoir for high concentrations of iron and zinc. The work identified and characterized metal tolerance proteins in common wheat and revealed a potential involvement of MTPs in providing a sink for trace element storage in wheat grains.

PGMiner reloaded, fully automated proteogenomic annotation tool linking genomes to proteomes

Abstract Improvements in genome sequencing technology increased the availability of full genomes and transcriptomes of many organisms. However, the major benefit of massive parallel sequencing is to better understand the organization and function of genes which then lead to understanding of phenotypes. In order to interpret genomic data with automated gene annotation studies, several tools are currently available. Even though the accuracy of computational gene annotation is increasing, a combination of multiple lines of experimental evidences should be gathered. Mass spectrometry allows the identification and sequencing of proteins as major gene products; and it is only these proteins that conclusively show whether a part of a genome is a coding region or not to result in phenotypes. Therefore, in the field of proteogenomics, the validation of computational methods is done by exploiting mass spectrometric data. As a result, identification of novel protein coding regions, validation of current gene models, and determination of upstream and downstream regions of genes can be achieved. In this paper, we present new functionality for our proteogenomic tool, PGMiner which performs all proteogenomic steps like acquisition of mass spectrometric data, peptide identification against preprocessed sequence databases, assignment of statistical confidence to identified peptides, mapping confident peptides to gene models, and result visualization. The extensions cover determining proteotypic peptides and thus unambiguous protein identification. Furthermore, peptides conflicting with gene models can now automatically assessed within the context of predicted alternative open reading frames.

Nuclear factor-like factor 2-antioxidant response element signaling activation by tert-butylhydroquinone attenuates acute heat stress in bovine mammary epithelial cells.

Nuclear factor (erythroid-derived 2)-like factor 2 (Nrf2) is a transcription factor that binds to the antioxidant response element (ARE) in the upstream promoter region of many antioxidative genes. The Nrf2-ARE signaling plays a key role in the cellular antioxidant-defense system, but whether Nrf2 activation has protective effects against heat shock (HS) stress in mammary epithelial cells (MEC) remains unclear. The objective of this study was to determine whether tert-butylhydroquinone (tBHQ), a well-known Nrf2 activator, could attenuate heat stress-induced cell damage in MAC-T cells of the bovine MEC line. The MAC-T cells were exposed to HS (42.5°C for 1h) followed by recovery at 37°C to mimic HS. Compared with cells that were consistently cultured at normothermia (37°C), the cell viability levels significantly decreased after HS stress. In parallel, heat stress increased the reactive oxygen species levels and induced cellular apoptosis and endoplasmic reticulum stress. The MAC-T cells that were pretreated with tBHQ (10μM) for 2h followed by HS had a reduction in the loss of cell viability. The tBHQ pretreatment significantly decreased cellular reactive oxygen species levels and stress-related marker gene expression. The tBHQ-treated MAC-T cells showed strong Nrf2-ARE signaling activation and a nuclear accumulation of Nrf2 and upregulated expression of Nrf2-ARE downstream genes. Small interfering RNA silencing of Nrf2 in HS-treated MAC-T cells almost completely abolished the cytoprotective effects by tBHQ. Overall, our results demonstrated that HS could cause cell damage in cultured bovine MEC, and that activation of Nrf2 by tBHQ could attenuate HS-induced cell damage.

Identification of a cis-acting element in nitrogen fixation genes recognized by CnfR in the nonheterocystous nitrogen-fixing cyanobacterium Leptolyngbya boryana.

The filamentous cyanobacterium Leptolyngbya boryana has the ability to fix nitrogen without any heterocysts under microoxic conditions. Previously, we identified the cnfR gene for a master transcriptional activator for nitrogen fixation (nif) genes in a 50-kb gene cluster containing nif and nif-related genes in L. boryana. We showed that CnfR activates the transcription of nif genes in response to low oxygen conditions, which allows the oxygen-vulnerable enzyme nitrogenase to function. However, the regulatory mechanism that underlies regulation by CnfR remains unknown. In this study, we identified a conserved cis-acting element that is recognized by CnfR. We established a reporter system in the non-diazotrophic cyanobacterium Synechocystis sp. PCC 6803 using luciferase genes (luxAB). Reporter analysis was performed with a series of truncated and modified upstream regulatory regions of nifB and nifP. The cis-element can be divided into nine motifs I-IX, and it is located 76 bp upstream of the transcriptional start sites of nifB and nifP. Six motifs of them are essential for transcriptional activation by CnfR. This cis-acting element is conserved in the upstream regions of nif genes in all diazotrophic cyanobacteria, including Anabaena and Cyanothece, thereby suggesting that the transcriptional regulation by CnfR is widespread in nitrogen-fixing cyanobacteria.

High-throughput cis-regulatory element discovery in the vector mosquito Aedes aegypti.

BackgroundDespite substantial progress in mosquito genomic and genetic research, few cis-regulatory elements (CREs), DNA sequences that control gene expression, have been identified in mosquitoes or other non-model insects. Formaldehyde-assisted isolation of regulatory elements paired with DNA sequencing, FAIRE-seq, is emerging as a powerful new high-throughput tool for global CRE discovery. FAIRE results in the preferential recovery of open chromatin DNA fragments that are not bound by nucleosomes, an evolutionarily conserved indicator of regulatory activity, which are then sequenced. Despite the power of the approach, FAIRE-seq has not yet been applied to the study of non-model insects. In this investigation, we utilized FAIRE-seq to profile open chromatin and identify likely regulatory elements throughout the genome of the human disease vector mosquito Aedes aegypti. We then assessed genetic variation in the regulatory elements of dengue virus susceptible (Moyo-S) and refractory (Moyo-R) mosquito strains.ResultsAnalysis of sequence data obtained through next generation sequencing of FAIRE DNA isolated from A. aegypti embryos revealed >121,000 FAIRE peaks (FPs), many of which clustered in the 1 kb 5’ upstream flanking regions of genes known to be expressed at this stage. As expected, known transcription factor consensus binding sites were enriched in the FPs, and of these FoxA1, Hunchback, Gfi, Klf4, MYB/ph3 and Sox9 are most predominant. All of the elements tested in vivo were confirmed to drive gene expression in transgenic Drosophila reporter assays. Of the >13,000 single nucleotide polymorphisms (SNPs) recently identified in dengue virus-susceptible and refractory mosquito strains, 3365 were found to map to FPs.ConclusionFAIRE-seq analysis of open chromatin in A. aegypti permitted genome-wide discovery of CREs. The results of this investigation indicate that FAIRE-seq is a powerful tool for identification of regulatory DNA in the genomes of non-model organisms, including human disease vector mosquitoes.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2468-x) contains supplementary material, which is available to authorized users.

Transcriptome analysis of reproductive-stage Arabidopsis plants exposed gamma-ray irradiation at various doses

Purpose: Gamma rays (GR) induce significant changes in the structure and expression of genes involved in the regulation of diverse biochemical and physiological processes. Arabidopsis plants exhibit different growth and development patterns in response to exposure to GR. The effects on gene expression of different radiation doses of GR (100 and 800 Gy) administered to Arabidopsis plants were examined at the reproductive stage.Materials and methods: We irradiated 26-day-old plants with three replications [developmental stages 5.1–6.0, according to Boyes et al. (2001)] using a GR irradiator (60 Co, ca. 150 TBq capacity, Atomic Energy of Canada Limited, Ontario, Canada) at the Korea Atomic Energy Research Institute. Plants were treated with 100, 200, 300, 400, 800, 1200, 1600, or 2000 Gy, and the doses were made from varying the distance to the source.Results: We conducted a high-throughput screening analysis and detected 883 GR-responsive genes that showed significant changes; these were involved in several putative metabolic pathways related to biotic stress. Additionally, five overrepresented cis-regulatory elements were identified in the 1-kb upstream regions of GR-responsive genes by using motif enrichment analysis. We also detected three GR-responsive genes associated with stamen development and confirmed their co-regulation with functionally interacting genes.Conclusions: This finding suggests that a network-based analysis is a viable approach to identify significant GR-responsive genes associated with the reproductive stage of Arabidopsis. Our results provide further insights into the complex biological systems involved in the response to different doses of GR in plants.

PP2C-like Promoter and Its Deletion Variants Are Induced by ABA but Not by MeJA and SA in Arabidopsis thaliana.

Gene expression is mediated through interaction between cis regulatory elements and its cognate transcription factors. Cis regulatory elements are defined as non-coding DNA sequences that provide the binding sites for transcription factors and are clustered in the upstream region of genes. ACGT cis regulatory element is one of the important cis regulatory elements found to be involved in diverse biological processes like auxin response, salicylic acid (SA) response, UV light response, ABA response and jasmonic acid (JA) response. We identified through in silico analysis that the upstream region of protein phosphatase 2C (PP2C) gene has a distinct genetic architecture of ACGT elements. In the present study, the activation of the full length promoter and its deletion constructs like 900 base pair, 500 base pair, 400 base pair and NRM (Nathji Rajesh Mehrotra) were examined by stable transformation in Arabidopsis thaliana using β-glucuronidase as the reporter gene. Evaluation of deletion constructs of PP2C-like promoter was carried out in the presence of phytohormones like abscisic acid (ABA), SA and JA. Our result indicated that the full length and 900 base pair promoter-reporter constructs of PP2C-like promoter was induced in response to ABA but not to methyl jasmonate and SA.

Phosphate starvation induces DNA methylation in the vicinity of cis-acting elements known to regulate the expression of phosphate–responsive genes

ABSTRACTPhosphate (Pi) limitation is a constraint for plant growth in many natural and agricultural ecosystems. Plants possess adaptive mechanisms that enable them to cope with conditions of limited Pi supply, including a highly regulated genetic program controlling the expression of genes involved in different metabolic, signaling and development processes of plants. Recently, we showed that in response to phosphate limitation Arabidopsis thaliana sets specific DNA methylation patterns of genic features that often correlated with changes in gene expression. Our findings included, dynamic methylation changes in response to phosphate starvation and the observation that the expression of genes encoding DNA methyltransferases appear to be directly controlled by the key regulator PHOSPHATE RESPONSE 1 (PHR1). These results provide insight into how epigenetic marks can influence plant genomes and transcriptional programs to respond and adapt to harsh conditions. Here we present an analysis of DNA methylation in the upstream regions of low Pi responsive genes in Arabidopsis seedlings exposed to low Pi conditions. We found that hypo- and hyper-methylation in the vicinity of cognate binding sites for transcription factors known to regulate the phosphate starvation response clearly correlates with increased or decreased expression of low-Pi responsive genes.