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Introduction A vaccine for malaria is urgently required but no vaccine has yet shown satisfactory protective efficacy especially for Plasmodium falciparum. P. falciparum infection can progress to cerebral malaria (CM), a neurological syndrome with exceedingly high mortality. Designing effective P. falciparum vaccines require more understanding of the protective immune response while the host immune response to CM and the mechanisms are still elusive. Here, we aim to identify host gene responses to CM and host gene networks associated with CM pathogenesis. Methods An innovative genomic analysis strategy, the weighted gene coexpression network analysis (WGCNA) combined with differential gene expression analysis, was used in this study. Data for analysis contain 93 whole blood samples, derived from two previous public transcriptome datasets. Results This approach led to the identification of numerous differentially expressed human transcripts and dozens of coexpression gene modules. We further identified nine key genes, including MBP, SAMSN1, PSMF1, SLC39A8, EIF3B, SMPDL3A, FABP5, SPSB3, and SHARPIN, of which the last four genes were first identified to be related to CM in the present study. Conclusion The results provided a comprehensive characterization of host gene expression profiles in CM and offered some new insight into malaria vaccine design. These identified key genes could be potential targets or immune modulators for novel therapeutic interventions of CM., Graphical abstract The present study used an innovative genomic analysis strategy to explored gene coexpression networks associated with cerebral malaria (CM) pathogenesis and identified several key host genes. The findings will be of interest to host–parasite interaction study and would be of assistance to further investigate CM pathology.

To systematically evaluate the whole-transcriptome sequencing data of cholangiocarcinoma (CHOL) to gain more insights into the transcriptomic landscape and molecular mechanism of this cancer, we performed whole-transcriptome sequencing based on the tumorous (C) and their corresponding non-tumorous adjacent to the tumors (CP) from eight CHOL patients. Subsequently, differential expression analysis was performed on the C and CP groups, followed by functional interaction prediction analysis to investigate gene-regulatory circuits in CHOL. In addition, The Cancer Genome Atlas (TCGA) for CHOL data was used to validate the results. In total, 2,895 differentially expressed messenger RNAs (dif-mRNAs), 56 differentially expressed microRNAs (dif-miRNAs), 151 differentially expressed long non-coding RNAs (dif-lncRNAs), and 110 differentially expressed circular RNAs (dif-circRNAs) were found in CHOL samples compared with controls. Enrichment analysis on those differentially expressed genes (DEGs) related to miRNA, lncRNA, and circRNA also identified the function of spliceosome. The downregulated hsa-miR-144-3p were significantly enriched in the competing endogenous RNA (ceRNA) complex network, which also included 7 upregulated and 13 downregulated circRNAs, 7 upregulated lncRNAs, and 90 upregulated and 40 downregulated mRNAs. Moreover, most of the DEGs and a few of the miRNAs (such as hsa-miR-144-3p) were successfully validated by TCGA data. The genes involved in RNA splicing and protein degradation processes and miR-144-3p may play fundamental roles in the pathogenesis of CHOL., Graphical Abstract, The whole-transcriptome sequencing data of cholangiocarcinoma (CHOL) were systematically evaluated to gain more insights into the transcriptomic landscape and molecular mechanism of this cancer.

Retroperitoneal liposarcoma (RLPS) is one of the most common types of retroperitoneal sarcomas, and has a high recurrence rate. There is an urgent need to further explore its pathogenesis and develop more effective treatment strategies. The aim of the present study was to identify potential driver genes of RLPS through bioinformatics analysis and molecular biology to elucidate potential targets that are suitable for further analysis for the treatment of RLPS. Differentially expressed genes (DEGs) between liposarcoma and normal fatty (NF) tissues were identified based on microarray data through bioinformatics analysis, and thymidylate synthase (TYMS) was selected from the DEGs, based on high content screening (HCS). TYMS expression was evaluated in RLPS tumor tissues and cell lines. A total of 21 RLPS tissues and 10 NF frozen tissues were used for reverse transcription-quantitative PCR, and 47 RLPS formalin-fixed specimens were used for immunohistochemical analysis. The effect of TYMS downregulation on cell proliferation, apoptosis, cell cycle progression, and cell migration and invasion were evaluated using lentivirus-mediated short hairpin RNA. The underlying mechanisms of TYMS in RLPS were examined by protein microarray and verified by western blotting. A total of 855 DEGs were identified. TYMS knockdown had the most notable effect on the proliferative capacity of RLPS cells according to the HCS results. TYMS mRNA expression levels were higher in RLPS tissues compared with NF tissues (P

Background & Aims The accumulation of neutral lipids within hepatocytes underlies non-alcoholic fatty liver disease (NAFLD), which affects a quarter of the world’s population and is associated with hepatitis, cirrhosis, and hepatocellular carcinoma. Despite insights gained from both human and animal studies, our understanding of NAFLD pathogenesis remains limited. To better study the molecular changes driving the condition we aimed to generate a humanised NAFLD mouse model. Methods We generated TIRF (transgene-free Il2rg-/-/Rag2-/-/Fah-/-) mice, populated their livers with human hepatocytes, and fed them a Western-type diet for 12 weeks. Results Within the same chimeric liver, human hepatocytes developed pronounced steatosis whereas murine hepatocytes remained normal. Unbiased metabolomics and lipidomics revealed signatures of clinical NAFLD. Transcriptomic analyses showed that molecular responses diverged sharply between murine and human hepatocytes, demonstrating stark species differences in liver function. Regulatory network analysis indicated close agreement between our model and clinical NAFLD with respect to transcriptional control of cholesterol biosynthesis. Conclusions These NAFLD xenograft mice reveal an unexpected degree of evolutionary divergence in food metabolism and offer a physiologically relevant, experimentally tractable model for studying the pathogenic changes invoked by steatosis. Lay summary Fatty liver disease is an emerging health problem, and as there are no good experimental animal models, our understanding of the condition is poor. We here describe a novel humanised mouse system and compare it with clinical data. The results reveal that the human cells in the mouse liver develop fatty liver disease upon a Western-style fatty diet, whereas the mouse cells appear normal. The molecular signature (expression profiles) of the human cells are distinct from the mouse cells and metabolic analysis of the humanised livers mimic the ones observed in humans with fatty liver. This novel humanised mouse system can be used to study human fatty liver disease., Graphical abstract, Highlights • Human liver chimeric mice fed a western diet develop NAFLD. • Within the same chimeric liver, human hepatocytes developed pronounced steatosis while murine hepatocytes remained normal. • Unbiased metabolomics and lipidomics of fatty humanised mouse livers revealed signatures of clinical NAFLD. • Transcriptomic analyses showed that molecular responses diverged sharply between murine and human hepatocytes. • This humanised NAFLD model is a physiologically relevant, experimentally tractable model for the study of steatosis.

Toluene is an industrial raw material and solvent that can be found abundantly in our daily life products. The amount of toluene vapor is one of the most important measurements for evaluating air quality. The evaluation of toluene scavenging ability of different plants has been reported, but the mechanism of plant response to toluene is only partially understood. In this study, we performed RNA sequencing (RNA-seq) analysis to detect differential gene expression in toluene-treated and untreated leaves of Ardisiapusilla. A total of 88,444 unigenes were identified by RNA-seq analysis, of which 49,623 were successfully annotated and 4101 were differentially expressed. Gene ontology analysis revealed several subcategories of genes related to toluene response, including cell part, cellular process, organelle, and metabolic processes. We mapped the main metabolic pathways of genes related to toluene response and found that the differentially expressed genes were mainly involved in glycolysis/gluconeogenesis, starch and sucrose metabolism, glycerophospholipid metabolism, carotenoid biosynthesis, phenylpropanoid biosynthesis, and flavonoid biosynthesis. In addition, 53 transcription factors belonging to 13 transcription factor families were identified. We verified 10 differentially expressed genes related to metabolic pathways using quantitative real-time PCR and found that the results of RNA-seq were positively correlated with them, indicating that the transcriptome data were reliable. This study provides insights into the metabolic pathways involved in toluene response in plants.

Flooding results in significant crop yield losses due to exposure of plants to hypoxic stress. Various studies have reported the effect of flooding stress at seedling establishment or later stages. However, the molecular mechanism prevailing at the germination stage under flooding stress remains enigmatic. The present study highlights the comparative transcriptome analysis in two rapeseed lines, i.e., flooding-tolerant (Santana) and -sensitive (23651) lines under control and 6-h flooding treatments at the germination stage. A total of 1840 up-regulated and 1301 down-regulated genes were shared by both lines in response to flooding. There were 4410 differentially expressed genes (DEGs) with increased expression and 4271 DEGs with reduced expression shared in both control and flooding conditions. Gene ontology (GO) enrichment analysis revealed that “transcription regulation”, “structural constituent of cell wall”, “reactive oxygen species metabolic”, “peroxidase”, oxidoreductase”, and “antioxidant activity” were the common processes in rapeseed flooding response. In addition, the processes such as “hormone-mediated signaling pathway”, “response to organic substance response”, “motor activity”, and “microtubule-based process” are likely to confer rapeseed flooding resistance. Mclust analysis clustered DEGs into nine modules, genes in each module shared similar expression patterns and many of these genes overlapped with the top 20 DEGs in some groups. This work provides a comprehensive insight into gene responses and the regulatory network in rapeseed flooding stress and provides guidelines for probing the underlying molecular mechanisms in flooding resistance.

As one of the most important poultry worldwide, ducks (Anas platyrhynchos) are raised mainly for meat and egg products, and muscle development in ducks is important for meat production. Therefore, an investigation of gene expression in duck skeletal muscle would significantly contribute to our understanding of muscle development. In this study, twenty-four cDNA libraries were constructed from breast and leg muscles of Hanzhong Ma ducks at day 17, 21, 27 of the embryo and postnatal at 6-month-old. High-throughput sequencing and bioinformatics were used to determine the abundances and characteristics of transcripts. A total of 632,172,628 (average 52,681,052) and 637,213,938 (average 53,101,162) reads were obtained from the sequencing data of breast and leg muscles, respectively. Over 71.63% and 77.36% of the reads could be mapped to the Anas platyrhynchos genome. In the skeletal muscle of Hanzhong duck, intron variant (INTRON), synonymous variant (SYNONYMOUS_CODING), and prime 3′ UTR variant (UTR_3_PRIME) were the main single nucleotide polymorphisms (SNP) annotation information, and “INTRON”, “UTR_3_PRIME”, and downstream-gene variant (DOWNSTREAM) were the main insertion-deletion (InDel) annotation information. The predicted number of alternative splicing (AS) in all samples were mainly alternative 5′ first exon (transcription start site)-the first exon splicing (TSS) and alternative 3′ last exon (transcription terminal site)-the last exon splicing (TTS). Besides, there were 292 to 2801 annotated differentially expressed genes (DEGs) in breast muscle and 304 to 1950 annotated DEGs in leg muscle from different databases. It is worth noting that 75 DEGs in breast muscle and 49 DEGs in leg muscle were co-expressed at all developmental points of comparison, respectively. The RNA-Seq data were confirmed to be reliable by qPCR. The identified DEGs, such as CREBL2, RHEB, GDF6, SHISA2, MYLK2, ACTN3, RYR3, and STMN1, were specially highlighted, indicating their strong associations with muscle development in the Hanzhong Ma duck. KEGG pathway analysis suggested that regulation of actin cytoskeleton, oxidative phosphorylation, and focal adhesion were involved in the development of skeletal muscle. The findings from this study can contribute to future investigations of the growth and development mechanism in duck skeletal muscle.

Background Single-cell RNA-Sequencing (scRNA-Seq) has provided single-cell level insights into complex biological processes. However, the high frequency of gene expression detection failures in scRNA-Seq data make it challenging to achieve reliable identification of cell-types and Differentially Expressed Genes (DEG). Moreover, with the explosive growth of single-cell data using 10x genomics protocol, existing methods will soon reach the computation limit due to scalability issues. The single-cell transcriptomics field desperately need new tools and framework to facilitate large-scale single-cell analysis. Results In order to improve the accuracy, robustness, and speed of scRNA-Seq data processing, we propose a generalized zero-inflated negative binomial mixture model, “JOINT,” that can perform probability-based cell-type discovery and DEG analysis simultaneously without the need for imputation. JOINT performs soft-clustering for cell-type identification by computing the probability of individual cells, i.e. each cell can belong to multiple cell types with different probabilities. This is drastically different from existing hard-clustering methods where each cell can only belong to one cell type. The soft-clustering component of the algorithm significantly facilitates the accuracy and robustness of single-cell analysis, especially when the scRNA-Seq datasets are noisy and contain a large number of dropout events. Moreover, JOINT is able to determine the optimal number of cell-types automatically rather than specifying it empirically. The proposed model is an unsupervised learning problem which is solved by using the Expectation and Maximization (EM) algorithm. The EM algorithm is implemented using the TensorFlow deep learning framework, dramatically accelerating the speed for data analysis through parallel GPU computing. Conclusions Taken together, the JOINT algorithm is accurate and efficient for large-scale scRNA-Seq data analysis via parallel computing. The Python package that we have developed can be readily applied to aid future advances in parallel computing-based single-cell algorithms and research in various biological and biomedical fields.

Background: Glioblastoma (GBM) has a high degree of malignancy, aggressiveness and recurrence rate. However, there are limited options available for the treatment of GBM, and they often result in poor prognosis and unsatisfactory outcomes. Materials and methods: In order to identify potential core genes in GBM that may provide new therapeutic insights, we analyzed three gene chips (GSE2223, GSE4290 and GSE50161) screened from the GEO database. Differentially expressed genes (DEG) from the tissues of GBM and normal brain were screened using GEO2R. To determine the functional annotation and pathway of DEG, Gene Ontology (GO) and KEGG pathway enrichment analysis were conducted using DAVID database. Protein interactions of DEG were visualized using PPI network on Cytoscape software. Next, 10 Hub nodes were screened from the differentially expressed network using MCC algorithm on CytoHubba software and subsequently identified as Hub genes. Finally, the relationship between Hub genes and the prognosis of GBM patients was described using GEPIA2 survival analysis web tool. Results: A total of 37 up-regulated and 187 down-regulated genes were identified through microarray analysis. Amongst the 10 Hub genes selected, SV2B appeared to be the only gene associated with poor prognosis in glioblastoma based on the survival analysis. Conclusion: Our study suggests that high expression of SV2B is associated with poor prognosis in GBM patients. Whether SV2B can be used as a new therapeutic target for GBM requires further validation.

Mating triggers substantial changes in gene expression and leads to subsequent physiological and behavioral modifications. However, postmating transcriptomic changes responding to mating have not yet been fully understood. Here, we carried out RNA sequencing (RNAseq) analysis in the sweet potato whitefly, Bemisia tabaci MED, to identify genes in females in response to mating. We compared mRNA expression in virgin and mated females at 24 h. As a result, 434 differentially expressed gene transcripts (DEGs) were identified between the mated and unmated groups, including 331 up- and 103 down-regulated. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that many of these DEGs encode binding-related proteins and genes associated with longevity. An RT-qPCR validation study was consistent with our transcriptomic analysis (14/15). Specifically, expression of P450s (Cyp18a1 and Cyp4g68), ubiquitin-protein ligases (UBR5 and RNF123), Hsps (Hsp68 and Hsf), carboxylase (ACC-2), facilitated trehalose transporters (Tret1-2), transcription factor (phtf), and serine-protein kinase (TLK2) were significantly elevated in mated females throughout seven assay days. These combined results offer a glimpe of postmating molecular modifications to facilitate reproduction in B. tabaci females.

Wheat powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is one of the most destructive fungal diseases threatening global wheat production. Host resistance is well known to be the most efficient method to control this disease. However, the molecular mechanism of wheat powdery mildew resistance (Pm) is still unclear. To analyze the molecular mechanism of Pm, we used the resistant wheat cultivar Jimai 23 to investigate its potential resistance components and profiled its expression in response to powdery mildew infection using bulked segregant RNA-Seq (BSR-Seq). We showed that the Pm of Jimai 23 was provided by a single dominant gene, tentatively designated PmJM23, and assigned it to the documented Pm2 region of chromosome 5DS. 3,816 consistently different SNPs were called between resistant and susceptible parents and the bulked pools derived from the combinations between the resistant parent Jimai23 and the susceptible parent Tainong18. 58 of the SNPs were assigned to the candidate region of PmJM23. Subsequently, 3,803 differentially expressed genes (DEGs) between parents and bulks were analyzed by GO, COG and KEGG pathway enrichment. The temporal expression patterns of associated genes following Bgt inoculation were further determined by RT-qPCR. Expression of six disease-related genes was investigated during Bgt infection and might serve as valuable genetic resources for the improvement of durable resistance to Bgt.

Graphical abstract, Disco-interacting protein 2 homolog B (Dip2B) is a member of Dip2 family encoded by Dip2b gene. Dip2B has been reported to regulate murine epithelial KIT+ progenitor cell expansion and differentiation epigenetically via exosomal miRNA targeting during salivary gland organogenesis. However, its molecular functions, cellular activities and biological process remain unstudied. Here, we investigated the transcriptome of Dip2B-deficient mouse embryonic lung fibroblasts (MELFs) isolated from E14.5 embryos by RNA-Seq. Expression profiling identified 1369 and 1104 differentially expressed genes (DEGs) from Dip2b−/− and Dip2b+/− MELFs in comparisons to wild-type (Dip2b+/+). Functional clustering of DEGs revealed that many gene ontology terms belong to membrane activities such as ‘integral component of plasma membrane’, and ‘ion channel activity’, suggesting possible roles of Dip2B in membrane integrity and membrane function. KEGG pathway analysis revealed that multiple metabolic pathways are affected in Dip2b−/− and Dip2b+/− when compared to Dip2b+/+ MELFs. These include ‘protein digestion and absorption’, ‘pancreatic secretion’ and ‘steroid hormone synthesis pathway’. These results suggest that Dip2B may play important roles in metabolism. Molecular function analysis shows transcription factors including Hox-genes, bHLH-genes, and Forkhead-genes are significantly down-regulated in Dip2b−/− MELFs. These genes are critical in embryo development and cell differentiation. In addition, Dip2B-deficient MELFs demonstrated a reduction in cell proliferation and migration, and an increase in apoptosis. All results indicate that Dip2B plays multiple roles in cell proliferation, migration and apoptosis during embryogenesis and may participate in control of metabolism. This study provides valuable information for further understanding of the function and regulatory mechanisms of Dip2B.

Background Enteroviruses (Picornaviridae family) have been widely detected in the feces from cattle with diarrhea. However, the mechanisms responsible for the pathogenicity of enteroviruses in cattle remain unclear. Recently, we isolated a novel EV-F7 strain called SWUN-AB001 from diarrheal yak (Bos grunniens) feces. To explore the pathogenic mechanisms of this novel virus, we used a transcriptomics approach to find genes with differential expression patterns in Madin-Darby bovine kidney (MDBK) cells during infection with SWUN-AB001 over time. Results MDBK cells were sampled at 12 and 24 h post-infection (hpi) to represent the early and late stages of a SWUN-AB001 infection. Compared with the non-infected cells, 19 and 1050 differentially expressed genes (DEGs) were identified at 12 and 24 hpi, respectively. These DEGs were associated with disease, signal transduction, cellular process and cytokine signaling categories. At 24 hpi, the pathway enrichment analysis revealed that signal pathways such as c-Jun NH2-terminal kinase/ stress-activated protein kinase (JNK/SAPK) and mitogen-activated protein kinase (MAPK) pathways and cytokine-cytokine receptor interactions were associated with the interactions occurring between EV-F7 and MDBK cells. Our additional western blot analysis showed that the phosphorylation levels of JNK/SAPK and p38 MAPK proteins increased significantly in the MDBK cells at 24 hpi. The result indicated that infection with EV-F7 could activate JNK/SAPK and p38 MAPK pathways in MDBK cells, and possibly trigger large-scale cytokine production. Conclusion Our transcriptome analysis provides useful initial data towards better understanding of the infection mechanisms used by EV-F7, while highlighting the potential molecular relationships occurring between the virus and the host’s cellular components. Electronic supplementary material The online version of this article (10.1186/s12917-018-1721-8) contains supplementary material, which is available to authorized users.

As high-strength cotton fibers are critical components of high quality cotton, developing cotton cultivars with high-strength fibers as well as high yield is a top priority for cotton development. Recently, chromosome segment substitution lines (CSSLs) have been developed from high-yield Upland cotton (Gossypium hirsutum) crossed with high-quality Sea Island cotton (G. barbadense). Here, we constructed a CSSL population by crossing CCRI45, a high-yield Upland cotton cultivar, with Hai1, a Sea Island cotton cultivar with superior fiber quality. We then selected two CSSLs with significantly higher fiber strength than CCRI45 (MBI7747 and MBI7561), and one CSSL with lower fiber strength than CCRI45 (MBI7285), for further analysis. We sequenced all four transcriptomes at four different time points postanthesis, and clustered the 44,678 identified genes by function. We identified 2200 common differentially-expressed genes (DEGs): those that were found in both high quality CSSLs (MBI7747 and MBI7561), but not in the low quality CSSL (MBI7285). Many of these genes were associated with various metabolic pathways that affect fiber strength. Upregulated DEGs were associated with polysaccharide metabolic regulation, single-organism localization, cell wall organization, and biogenesis, while the downregulated DEGs were associated with microtubule regulation, the cellular response to stress, and the cell cycle. Further analyses indicated that three genes, XLOC_036333 [mannosyl-oligosaccharide-α-mannosidase (MNS1)], XLOC_029945 (FLA8), and XLOC_075372 (snakin-1), were potentially important for the regulation of cotton fiber strength. Our results suggest that these genes may be good candidates for future investigation of the molecular mechanisms of fiber strength formation and for the improvement of cotton fiber quality through molecular breeding.

Colorectal cancer (CRC) is one of the most frequently occurring malignancies worldwide. The outcomes of patients with similar clinical symptoms or at similar pathological stages remain unpredictable. This inherent clinical diversity is most likely due to the genetic heterogeneity. The present study aimed to create a predicting tool to evaluate patient survival based on genetic profile. Firstly, three Gene Expression Omnibus (GEO) datasets (GSE9348, GSE44076 and GSE44861) were utilized to identify and validate differentially expressed genes (DEGs) in CRC. The GSE14333 dataset containing survival information was then introduced in order to screen and verify prognosis-associated genes. Of the 66 DEGs, the present study screened out 46 biomarkers closely associated to patient overall survival. By Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis, it was demonstrated that these genes participated in multiple biological processes which were highly associated with cancer proliferation, drug-resistance and metastasis, thus further affecting patient survival. The five most important genes, MET proto-oncogene, receptor tyrosine kinase, carboxypeptidase M, serine hydroxymethyltransferase 2, guanylate cyclase activator 2B and sodium voltage-gated channel a subunit 9 were selected by a random survival forests algorithm, and were further made up to a linear risk score formula by multivariable cox regression. Finally, the present study tested and verified this risk score within three independent GEO datasets (GSE14333, GSE17536 and GSE29621), and observed that patients with a high risk score had a lower overall survival (P

Sepsis is a life-threatening disease induced by a systemic inflammatory response, which leads to organ dysfunction and mortality. In sepsis, the host immune response is depressed and unable to cope with infection, no drug is currently available to treat this. The lungs are frequently the starting point for sepsis. This study aimed to identify potential genes for diagnostics and therapeutic purposes in sepsis by a comprehensive bioinformatics analysis. Our criteria are to unravel sepsis-associated signature genes from gene expression datasets. Differentially expressed genes (DEGs) were identified from samples of sepsis patients using a meta-analysis and then further subjected to functional enrichment and protein‒protein interaction (PPI) network analysis for examining their potential functions. Finally, the expression of the topmost upregulated genes (ARG1, IL1R2, ELANE, MMP9) was quantified by reverse transcriptase-PCR (RT-PCR), and myeloperoxidase (MPO) expression was confirmed by immunohistochemistry (IHC) staining in the lungs of a well-established sepsis mouse model. We found that all the four genes were upregulated in semiquantitative RT-PCR studies, however, MMP9 showed a nonsignificant increase in expression. MPO staining showed strong immunoreactivity in sepsis as compared to the control. This study demonstrates the role of significant and widespread immune activation (IL1R2, MMP9), along with oxidative stress (ARG1) and the recruitment of neutrophils, in sepsis (ELANE, MPO).

The dove tree (Davidia involucrata Baill.), a tertiary relic species, is adapted to cool climates. With the progression of global warming, high-temperature stress has become the primary environmental factor restricting geographic distribution, ex situ conservation, and landscape application for D. involucrata resources. However, the detailed molecular events underlying D. involucrata responses to heat stress are poorly understood. Here, we conducted RNA-Seq-based gene expression profiling in D. involucrata seedlings during the time course of a 42 &deg, C heat treatment (0, 1, 6, and 12 h). After de novo assembly, we obtained 138,923 unigenes, of which 69,743 were annotated in public databases. Furthermore, 19,532, 20,497 and 27,716 differentially expressed genes (DEGs) were identified after 1 h (HS1), 6 h (HS6), and 12 h (HS12) of heat treatment in comparison to 0 h (HS0), respectively. Based on a KEGG enrichment analysis, the two pathways &ldquo, protein processing in endoplasmic reticulum&rdquo, and &ldquo, plant hormone signal transduction&rdquo, are hypothesized to play vital roles during heat response in D. involucrata, and their potential interactions during heat stress are also discussed. In addition, 32 genes encoding putative heat shock transcription factors (Hsfs) were found to be associated with the response of D. involucrata to heat stress. Finally, the expression patterns of eight heat-responsive genes derived from qRT-PCR were in agreement with their transcript level alterations, as determined by a transcriptome analysis. Taken together, our transcriptomic data provide the first comprehensive transcriptional profile affected by heat stress in D. involucrata, which will facilitate further studies on the improvement of heat tolerance in this rare and endangered species.

Barley is among four of the most important cereal crops with respect to global production. Increasing barley yields to desired levels can be achieved by the genetic manipulation of cytokinin content. Cytokinins are plant hormones that regulate many developmental processes and have a strong influence on grain yield. Cytokinin homeostasis is regulated by members of several multigene families. CKX genes encode the cytokinin oxidase/dehydrogenase enzyme, which catalyzes the irreversible degradation of cytokinin. Several recent studies have demonstrated that the RNAi-based silencing of CKX genes leads to increased grain yields in some crop species. To assess the possibility of increasing the grain yield of barley by knocking out CKX genes, we used an RNA-guided Cas9 system to generate ckx1 and ckx3 mutant lines with knockout mutations in the HvCKX1 and HvCKX3 genes, respectively. Homozygous, transgene-free mutant lines were subsequently selected and analyzed. A significant decrease in CKX enzyme activity was observed in the spikes of the ckx1 lines, while in the ckx3 lines, the activity remained at a similar level to that in the control plants. Despite these differences, no changes in grain yield were observed in either mutant line. In turn, differences in CKX activity in the roots between the ckx1 and ckx3 mutants were reflected via root morphology. The decreased CKX activity in the ckx1 lines corresponded to greater root length, increased surface area, and greater numbers of root hairs, while the increased CKX activity in the ckx3 mutants gave the opposite results. RNA-seq analysis of the spike and root transcriptomes revealed an altered regulation of genes controlling cytokinin metabolism and signaling, as well as other genes that are important during seed development, such as those that encode nutrient transporters. The observed changes suggest that the knockout of a single CKX gene in barley may be not sufficient for disrupting cytokinin homeostasis or increasing grain yields.

Background The common littoral shrimp Palaemon serratus is an economically important decapod resource in some European communities. Aquaculture practices prevent the genetic deterioration of wild stocks caused by overfishing and at the same time enhance the production. The biotechnological manipulation of sex-related genes has the proved potential to improve the aquaculture production but the scarcity of genomic data about P. serratus hinders these applications. RNA-Seq analysis has been performed on ovary and testis samples to generate a reference gonadal transcriptome. Differential expression analyses were conducted between three ovary and three testis samples sequenced by Illumina HiSeq 4000 PE100 to reveal sex-related genes with sex-biased or sex-specific expression patterns. Results A total of 224.5 and 281.1 million paired-end reads were produced from ovary and testis samples, respectively. De novo assembly of ovary and testis trimmed reads yielded a transcriptome with 39,186 transcripts. The 29.57% of the transcriptome retrieved at least one annotation and 11,087 differentially expressed genes (DEGs) were detected between ovary and testis replicates. Six thousand two hundred seven genes were up-regulated in ovaries meanwhile 4880 genes were up-regulated in testes. Candidate genes to be involved in sexual development and gonadal development processes were retrieved from the transcriptome. These sex-related genes were discussed taking into account whether they were up-regulated in ovary, up-regulated in testis or not differentially expressed between gonads and in the framework of previous findings in other crustacean species. Conclusions This is the first transcriptome analysis of P. serratus gonads using RNA-Seq technology. Interesting findings about sex-related genes from an evolutionary perspective (such as Dmrt1) and for putative future aquaculture applications (Iag or vitellogenesis genes) are reported here. We provide a valuable dataset that will facilitate further research into the reproductive biology of this shrimp.

Gene expression profiling is a useful tool to predict and interrogate mechanisms of toxicity. RNA-Seq technology has emerged as an attractive alternative to traditional microarray platforms for conducting transcriptional profiling. The objective of this work was to compare both transcriptomic platforms to determine whether RNA-Seq offered significant advantages over microarrays for toxicogenomic studies. RNA samples from the livers of rats treated for 5 days with five tool hepatotoxicants (α-naphthylisothiocyanate/ANIT, carbon tetrachloride/CCl4, methylenedianiline/MDA, acetaminophen/APAP, and diclofenac/DCLF) were analyzed with both gene expression platforms (RNA-Seq and microarray). Data were compared to determine any potential added scientific (i.e., better biological or toxicological insight) value offered by RNA-Seq compared to microarrays. RNA-Seq identified more differentially expressed protein-coding genes and provided a wider quantitative range of expression level changes when compared to microarrays. Both platforms identified a larger number of differentially expressed genes (DEGs) in livers of rats treated with ANIT, MDA, and CCl4 compared to APAP and DCLF, in agreement with the severity of histopathological findings. Approximately 78% of DEGs identified with microarrays overlapped with RNA-Seq data, with a Spearman’s correlation of 0.7 to 0.83. Consistent with the mechanisms of toxicity of ANIT, APAP, MDA and CCl4, both platforms identified dysregulation of liver relevant pathways such as Nrf2, cholesterol biosynthesis, eiF2, hepatic cholestasis, glutathione and LPS/IL-1 mediated RXR inhibition. RNA-Seq data showed additional DEGs that not only significantly enriched these pathways, but also suggested modulation of additional liver relevant pathways. In addition, RNA-Seq enabled the identification of non-coding DEGs that offer a potential for improved mechanistic clarity. Overall, these results indicate that RNA-Seq is an acceptable alternative platform to microarrays for rat toxicogenomic studies with several advantages. Because of its wider dynamic range as well as its ability to identify a larger number of DEGs, RNA-Seq may generate more insight into mechanisms of toxicity. However, more extensive reference data will be necessary to fully leverage these additional RNA-Seq data, especially for non-coding sequences.

Background Enormous variability in skin colour and patterning is a characteristic of teleost fish, including Salmonidae fishes, which present themselves as a suitable model for studying mechanisms of pigment patterning. In order to screen for candidate genes potentially involved in the specific skin pigment pattern in marble trout (labyrinthine skin pattern) and brown trout (spotted skin pattern), we conducted comparative transcriptome analysis between differently pigmented dermis sections of the adult skin of the two species. Results Differentially expressed genes (DEGs) possibly associated with skin pigment pattern were identified. The expression profile of 27 DEGs was further tested with quantitative real-time PCR on a larger number of samples. Expression of a subset of ten of these genes was analysed in hybrid (marble x brown) trout individuals and compared with the complexity of their skin pigment pattern. A correlation between the phenotype and the expression profile assessed for hybrid individuals was detected for four (gja5, clcn2, cdkn1a and tjp1) of the ten candidate genes tested. The potential role of these genes in skin pigment pattern maintenance is discussed. Conclusions Our results indicate that the maintenance of different pigment patterns in trout is dependent upon specific communication—involving gap junctions, tight junctions and ion channels—between chromatophores present in differentially pigmented skin regions. Electronic supplementary material The online version of this article (10.1186/s12864-019-5714-1) contains supplementary material, which is available to authorized users.

Alterations in DNA methylation and gene expression have been implicated in the development of human dilated cardiomyopathy (DCM). Differentially methylated probes (DMPs) and differentially expressed genes (DEGs) were identified between the left ventricle (LV, a pathological locus for DCM) and the right ventricle (RV, a proxy for normal hearts). The data in this DiB are for supporting our report entitled “Methylome analysis reveals alterations in DNA methylation in the regulatory regions of left ventricle development genes in human dilated cardiomyopathy” (Bong-Seok Jo, In-Uk Koh, Jae-Bum Bae, Ho-Yeong Yu, Eun-Seok Jeon, Hae-Young Lee, Jae-Joong Kim, Murim Choi, Sun Shim Choi, 2016) [1].

To life scientists, one important feature offered by RNAseq, a next-generation sequencing tool used to estimate changes in gene expression levels, lies in its unprecedented resolution. It can score countable differences in transcript numbers among thousands of genes and between experimental groups, all at once. However, its high cost limits experimental designs to very small sample sizes, usually N = 3, which often results in statistically underpowered analysis and poor reproducibility. All these issues are compounded by the presence of experimental noise, which is harder to distinguish from instrumental error when sample sizes are limiting (e.g., small-budget pilot tests), experimental populations exhibit biologically heterogeneous or diffuse expression phenotypes (e.g., patient samples), or when discriminating among transcriptional signatures of closely related experimental conditions (e.g., toxicological modes of action, or MOAs). Here, we present a leveraged signal-to-noise ratio (LSTNR) thresholding method, founded on generalized linear modeling (GLM) of aligned read detection limits to extract differentially expressed genes (DEGs) from noisy low-replication RNAseq data. The LSTNR method uses an agnostic independent filtering strategy to define the dynamic range of detected aggregate read counts per gene, and assigns statistical weights that prioritize genes with better sequencing resolution in differential expression analyses. To assess its performance, we implemented the LSTNR method to analyze three separate datasets: first, using a systematically noisy in silico dataset, we demonstrated that LSTNR can extract pre-designed patterns of expression and discriminate between “noise” and “true” differentially expressed pseudogenes at a 100% success rate; then, we illustrated how the LSTNR method can assign patient-derived breast cancer specimens correctly to one out of their four reported molecular subtypes (luminal A, luminal B, Her2-enriched and basal-like); and last, we showed the ability to retrieve five different modes of action (MOA) elicited in livers of rats exposed to three toxicants under three nutritional routes by using the LSTNR method. By combining differential measurements with resolving power to detect DEGs, the LSTNR method offers an alternative approach to interrogate noisy and low-replication RNAseq datasets, which handles multiple biological conditions at once, and defines benchmarks to validate RNAseq experiments with standard benchtop assays.

Aim: Co-infection in HIV-1 patients with Mycobacterium tuberculosis poses considerable risk of developing the immune reconstitution inflammatory syndrome (IRIS), especially upon the initiation of antiretroviral therapy (ART). Methodology & results: For transcriptomic analysis, peripheral blood mononuclear cells’ whole gene expression was used from three patient groups: HIV+ (H), HIV-TB+ (HT), HIV-TB+ with IRIS (HTI). Pathway enrichment and functional analysis was performed before and after highly active ART. Genes in the interferon-stimulating and ZNF families maintained tight functional interaction and tilted the balance in favor of TB-IRIS. Discussion & conclusion: The functional impairment of interaction between ZNF genes and interferon-stimulated genes, along with higher expression of S100A8/S100A9 genes possibly forms the genomic basis of TB-IRIS in a subset of HIV patients while on highly active ART.

The present study investigated the transcriptomic and metabolomic changes elicited in tomato plants (Solanum lycopersicum cv. Micro-Tom) following treatments with the biocontrol agent T. harzianum strain M10 or its secondary metabolite harzianic acid (HA), in the presence or absence of the soil-borne pathogen Rhizoctonia solani. Transcriptomic analysis allowed the identification of differentially expressed genes (DEGs) that play a pivotal role in plant growth, development and resistance to biotic stress. Overall, the results support the ability of T. harzianum M10 to activate defense responses in infected tomato plants. We observed an induction of hormone-mediated signaling, as shown by the up-regulation of genes involved in the ethylene and jasmonate (ET/JA) biosynthesis. Further, the protective action of T. harzianum on the host was revealed by the over-expression of genes able to detoxify cells from reactive oxygen species (ROS). Moreover, the over-expression of genes involved in glycolysis, tricarboxylic acid cycle (TCA) and photosynthesis help to understand how this strain promotes plant growth and development. On the other hand, HA treatment also stimulated tomato response to the pathogen by inducing the expression of several genes involved in different defense mechanisms (including protease inhibitors, resistance proteins like CC-NBS-LRR) and activating ET/JA- and salicylic acid (SA)-mediated signaling pathways. The accumulation of steroidal glycoalkaloids caused by either T. harzianum or purified HA, as determined by metabolomic analysis, confirmed the complexity of the plant response to beneficial microbes in terms of secondary metabolism involved in defense mechanisms against pathogens.

The passion fruit (Passiflora edulis Sims), also known as the purple granadilla, is widely cultivated as the new darling of the fruit market throughout southern China. This exotic and perennial climber is adapted to warm and humid climates, and thus is generally intolerant of cold. There is limited information about gene regulation and signaling pathways related to the cold stress response in this species. In this study, two transcriptome libraries (KEDU_AP vs. GX_AP) were constructed from the aerial parts of cold-tolerant and cold-susceptible varieties of P. edulis, respectively. Overall, 126,284,018 clean reads were obtained, and 86,880 unigenes with a mean size of 1449 bp were assembled. Of these, there were 64,067 (73.74%) unigenes with significant similarity to publicly available plant protein sequences. Expression profiles were generated, and 3045 genes were found to be significantly differentially expressed between the KEDU_AP and GX_AP libraries, including 1075 (35.3%) up-regulated and 1970 (64.7%) down-regulated. These included 36 genes in enriched pathways of plant hormone signal transduction, and 56 genes encoding putative transcription factors. Six genes involved in the ICE1–CBF–COR pathway were induced in the cold-tolerant variety, and their expression levels were further verified using quantitative real-time PCR. This report is the first to identify genes and signaling pathways involved in cold tolerance using high-throughput transcriptome sequencing in P. edulis. These findings may provide useful insights into the molecular mechanisms regulating cold tolerance and genetic breeding in Passiflora spp.

Background Osteoarthritis (OA) is one of the most prevalent chronic joint diseases while the precise genetic mechanism remains elusive. In this study, we investigated the gene expression profile in OA by microarray analysis. Results Histopathological characteristics of OA cartilage were examined using a rat model of leptin-induced OA. Gene expression profile of leptin-induced articular cartilage and healthy rat cartilage were compared using genome-wide microarray hybridization. A total of 1857 genes differentially expressed genes (1197 upregulated and 660 downregulated) were identified, some of which are known to be associated with leptin-induced OA phenotype. These included genes related to MMPs, inflammatory factors, growth factors, apoptotic genes and osteogenic genes. In addition, upregulated expressions of some new candidate genes, which have hitherto fore not been linked to OA (such as BCL2L11) were detected in leptin-induced OA cartilage, which suggests that these genes might be important for OA molecular mechanism. Conclusion Our findings suggest that pathogenesis of leptin-induced OA involves modulation of expression of multiple genes, although the underlying molecular mechanisms need to be studied further. Further investigation of leptin-induced gene expression changes is needed to gain new insights into the molecular mechanism of OA pathogenesis. Electronic supplementary material The online version of this article (doi:10.1186/s41065-017-0039-z) contains supplementary material, which is available to authorized users.

Boron (B) deficiency stress is frequently observed in citrus orchards and causes considerable loss of productivity and fruit quality. Carrizo citrange (Cc) has been reported as a rootstock more tolerant to B deficiency than Trifoliate orange (To). The ‘Newhall’ navel orange (Ns) performed better when grafted onto Cc (Ns/Cc) than when grafted onto To (Ns/To) under long-term B deficiency. The present study confirmed that Ns/Cc had higher boron content, leaf fresh weight, lower leaf chlorosis and stronger photosynthesis ability than Ns/To. Moreover, B-deficiency significantly reduced the chlorophyll and carotenoid content in Ns/To. The content of total soluble sugar and lignin were dramatically increased and the expression levels of photosynthesis-related genes were substantially down-regulated in Ns/To by B-deficient treatment. B-deficiency also strongly induced expression levels of chlorophyll decomposition-related genes, glucose synthesis-related genes and lignin synthesis-related genes, and significantly inhibited the expression of carotenoid synthesis-related genes in Ns/To. Overall, these findings suggested that the influence of To on the scion of Ns was worse than that of Cc due to differently regulating these metabolic pathways under the long term of B-deficiency. The transcriptome analysis provided further information for understanding the mechanism of the different responses of scion-rootstock combinations to B-deficiency stress.

Background Teleosts are exposed to a broad range of external stimuli, which may be either of acute or chronic nature. The larval phase of certain fish species offer a unique opportunity to study the interactions between genes and environmental factors during early life. The present study investigates the effects of early-life events, applied at different time points of early ontogeny (first feeding, flexion and development of all fins; Phase 1) as well as on the subsequent juvenile stage after the application of an additional acute stressor (Phase 2) in the gilthead sea bream (Sparus aurata), a commercially important European aquaculture species. Animal performance, the cortisol response and gene expression patterns during early development as well as on the subsequent phases (juveniles) after the application of additional acute stressors were investigated. Results Significant differences on fish performance were found only for juveniles exposed to early-life events at the phase of the formation of all fins. On the transcriptome level distinct expression patterns were obtained for larvae as well as for juveniles with the most divergent expression pattern found to be again at the phase of the development of all fins, which showed to have also an impact later on in the acute stress response of juveniles. Conclusions The present study showed that applying an early-life protocol, characterized by the unpredictable, variable and moderate intensity of the applied stimuli provides a relative realistic model to evaluate the impact of daily aquaculture practices on fish performance. In addition, the power of investigating global gene expression patterns is shown, providing significant insights regarding the response of early-life events during development and as juveniles after the application of extra acute stressors. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-2874-0) contains supplementary material, which is available to authorized users.

The Caenorhabditis elegans DEG/ENaC proteins MEC-4 and MEC-10 transduce gentle touch in the six touch receptor neurons . Gain-of-function mutations of mec-4 and mec-4(d) result in a hyperactive channel and neurodegeneration in vivo. Loss of MEC-6, a putative DEG/ENaC-specific chaperone, and of the similar protein POML-1 suppresses the neurodegeneration caused by a mec-4(d) mutation. We find that mutation of two genes, mec-10 and a new gene mec-19 (previously named C49G9.1), prevents this action of POML-1, allowing the touch receptor neurons to die in poml-1mec-4(d) animals. The proteins encoded by these genes normally inhibit mec-4(d) neurotoxicity through different mechanisms. MEC-10, a subunit of the mechanosensory transduction channel with MEC-4, inhibits MEC-4(d) activity without affecting MEC-4 expression. In contrast, MEC-19, a membrane protein specific to nematodes, inhibits MEC-4(d) activity and reduces MEC-4 surface expression.

Background Pathogenic members of the Fusarium oxysporum species complex are responsible for vascular wilt disease on many important crops including legumes, where they can be one of the most destructive disease causing necrotrophic fungi. We previously developed a model legume-infecting pathosystem based on the reference legume Medicago truncatula and a pathogenic F. oxysporum forma specialis (f. sp.) medicaginis (Fom). To dissect the molecular pathogenicity arsenal used by this root-infecting pathogen, we sequenced its transcriptome during infection of a susceptible and resistant host accession. Results High coverage RNA-Seq of Fom infected root samples harvested from susceptible (DZA315) or resistant (A17) M. truncatula seedlings at early or later stages of infection (2 or 7 days post infection (dpi)) and from vegetative (in vitro) samples facilitated the identification of unique and overlapping sets of in planta differentially expressed genes. This included enrichment, particularly in DZA315 in planta up-regulated datasets, for proteins associated with sugar, protein and plant cell wall metabolism, membrane transport, nutrient uptake and oxidative processes. Genes encoding effector-like proteins were identified, including homologues of the F. oxysporum f. sp. lycopersici Secreted In Xylem (SIX) proteins, and several novel candidate effectors based on predicted secretion, small protein size and high in-planta induced expression. The majority of the effector candidates contain no known protein domains but do share high similarity to predicted proteins predominantly from other F. oxysporum ff. spp. as well as other Fusaria (F. solani, F. fujikori, F. verticilloides, F. graminearum and F. pseudograminearum), and from another wilt pathogen of the same class, a Verticillium species. Overall, this suggests these novel effector candidates may play important roles in Fusaria and wilt pathogen virulence. Conclusion Combining high coverage in planta RNA-Seq with knowledge of fungal pathogenicity protein features facilitated the identification of differentially expressed pathogenicity associated genes and novel effector candidates expressed during infection of a resistant or susceptible M. truncatula host. The knowledge from this first in depth in planta transcriptome sequencing of any F. oxysporum ff. spp. pathogenic on legumes will facilitate the dissection of Fusarium wilt pathogenicity mechanisms on many important legume crops. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-3192-2) contains supplementary material, which is available to authorized users.

Background As the most important yield component in rapeseed (Brassica napus L.), pod number is determined by a series of successive growth and development processes. Pod number shows extensive variation in rapeseed natural germplasm, which is valuable for genetic improvement. However, the genetic and especially the molecular mechanism for this kind of variation are poorly understood. In this study, we conducted QTL mapping and RNA sequencing, respectively, using the BnaZNRIL population and its two parental cultivars Zhongshuang11 and No.73290 which showed significant difference in pod number, primarily due to the difference in floral organ number. Result A total of eight QTLs for pod number were identified using BnaZNRIL population with a high-density SNP linkage map, each was distributed on seven linkage groups and explained 5.8–11.9% of phenotypic variance. Then, they were integrated with those previously detected in BnaZNF2 population (deriving from same parents) and resulted in 15 consensus-QTLs. Of which, seven QTLs were identical to other studies, whereas the other eight should be novel. RNA sequencing of the shoot apical meristem (SAM) at the formation stage of floral bud primordia identified 9135 genes that were differentially expressed between the two parents. Gene ontology (GO) analysis showed that the top two enriched groups were S-assimilation, providing an essential nutrient for the synthesis of diverse metabolites, and polyamine metabolism, serving as second messengers that play an essential role in flowering genes initiation. KEGG analysis showed that the top three overrepresented pathways were carbohydrate (707 genes), amino acid (390 genes) and lipid metabolisms (322 genes). In silico mapping showed that 647 DEGs were located within the confidence intervals of 15 consensus QTLs. Based on annotations of Arabidopsis homologs corresponding to DEGs, nine genes related to meristem growth and development were considered as promising candidates for six QTLs. Conclusion In this study, we discovered the first repeatable major QTL for pod number in rapeseed. In addition, RNA sequencing was performed for SAM in rapeseed, which provides new insights into the determination of floral organ number. Furthermore, the integration of DEGs and QTLs identified promising candidates for further gene cloning and mechanism study. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-3402-y) contains supplementary material, which is available to authorized users.