Transcriptomic profiling analysis to identify genes associated with PA biosynthesis and insolubilization in the late stage of fruit development in C-PCNA persimmon
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Abstract PA-enhanced content causes astringency in persimmon fruit. PCNA persimmons can lose their astringency naturally and they become edible when still on the tree, which allows for conserves of physical and financial resources. C-PCNA persimmon originates in China. Its deastringency trait primarily depends on decreased PA biosynthesis and PA insolubilization at the late stage of fruit development. Although some genes and transcription factors that may be involved in the deastringency of C-PCNA persimmon have been reported, the expression patterns of these genes during the key deastringency stage are reported less. To investigate the variation in PA contents and the expression patterns of deastringency-related genes during typical C-PCNA persimmon ‘Xiaoguo-tianshi’ fruit development and ripening, PA content and transcriptional profiling were carried out at five late stages from 70 to 160 DAF. The combinational analysis phenotype, PA content, and DEG enrichment revealed that 120–140 DAF and 140–160 DAF were the critical phases for PA biosynthesis reduction and PA insolubilization, respectively. The expression of PA biosynthesis-associated genes indicated that the downregulation of the ANR gene at 140–160 DAF may be associated with PA biosynthesis and is decreased by inhibiting its precursor cis-flavan-3-ols. We also found that a decrease in acetaldehyde metabolism-associated ALDH genes and an increase in ADH and PDC genes might result in C-PCNA persimmon PA insolubilization. In addition, a few MYB-bHLH-WD40 (MBW) homologous transcription factors in persimmon might play important roles in persimmon PA accumulation. Furthermore, combined coexpression network analysis and phylogenetic analysis of MBW suggested that three putative transcription factors WD40 ( evm.TU.contig1.155 ), MYB ( evm.TU.contig8910.486 ) and bHLH ( evm.TU.contig1398.203 ), might connect and co-regulate both PA biosynthesis and its insolubilization in C-PCNA persimmon. The present study elucidated transcriptional insights into PA biosynthesis and insolubilization during the late development stages based on the C-PCNA D. kaki genome (unpublished). Thus, we focused on PA content variation and the expression patterns of genes involved in PA biosynthesis and insolubilization. Our work has provided additional evidence on previous knowledge and a basis for further exploration of the natural deastringency of C-PCNA persimmon.Keywords:
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Matrix metalloproteinase inhibitor
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AIM:To investigate the molecular pathways involved in human cholangiocarcinogenesis by gene expression profiling. METHODS:Oligonucleotide arrays (Affymetrix U133A) were used to establish a specific gene expression profile of intrahepatic CCC in comparison to corresponding nonmalignant liver tissue.To validate the expression values of the most overexpressed genes, RT-PCR experiments were performed. RESULTS:Five hundred and fifty-two statistically differentially expressed genes/ESTs (221 probes significantly up-regulated, 331 probes down-regulated; P < 0.05; fold change > 2; ≥ 70%) were identified.Using these data and two-dimensional cluster analysis, a specific gene expression profile was obtained allowing fast and reproducible differentiation of CCC, which was confirmed by supervised neuronal network modelling.The most consistently overexpressed gene (median fold change 33.5, significantly overexpressed in 100%) encoded osteopontin.Furthermore, an association of various genes with the histopathological grading could be demonstrated. CONCLUSION:A highly specific gene expression profile for intrahepatic CCC was identified, allowing for its fast and reproducible discrimination against nonmalignant liver tissue and other liver masses.The most overexpressed gene in intrahepatic CCC was the gene encoding osteopontin.These data may lead to a better understanding of human cholangiocarcinogenesis.
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In broilers, high ambient temperature can result in reduced feed consumption, digestive inefficiency, impaired metabolism, and even death. The broiler sector of the U.S. poultry industry incurs approximately $52 million in heat-related losses annually. The objective of this study is to characterize the effects of cyclic high ambient temperature on the transcriptome of a metabolically active organ, the liver. This study provides novel insight into the effects of high ambient temperature on metabolism in broilers, because it is the first reported RNA-seq study to characterize the effect of heat on the transcriptome of a metabolic-related tissue. This information provides a platform for future investigations to further elucidate physiologic responses to high ambient temperature and seek methods to ameliorate the negative impacts of heat. Transcriptome sequencing of the livers of 8 broiler males using Illumina HiSeq 2000 technology resulted in 138 million, 100-base pair single end reads, yielding a total of 13.8 gigabases of sequence. Forty genes were differentially expressed at a significance level of P-value < 0.05 and a fold-change ≥ 2 in response to a week of cyclic high ambient temperature with 27 down-regulated and 13 up-regulated genes. Two gene networks were created from the function-based Ingenuity Pathway Analysis (IPA) of the differentially expressed genes: "Cell Signaling" and "Endocrine System Development and Function". The gene expression differences in the liver transcriptome of the heat-exposed broilers reflected physiological responses to decrease internal temperature, reduce hyperthermia-induced apoptosis, and promote tissue repair. Additionally, the differential gene expression revealed a physiological response to regulate the perturbed cellular calcium levels that can result from high ambient temperature exposure. Exposure to cyclic high ambient temperature results in changes at the metabolic, physiologic, and cellular level that can be characterized through RNA-seq analysis of the liver transcriptome of broilers. The findings highlight specific physiologic mechanisms by which broilers reduce the effects of exposure to high ambient temperature. This information provides a foundation for future investigations into the gene networks involved in the broiler stress response and for development of strategies to ameliorate the negative impacts of heat on animal production and welfare.
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Background In psoriasis, only limited overlap between sets of genes identified as differentially expressed (psoriatic lesional vs. psoriatic non-lesional) was found using statistical and fold-change cut-offs. To provide a framework for utilizing prior psoriasis data sets we sought to understand the consistency of those sets. Methodology/Principal Findings Microarray expression profiling and qRT-PCR were used to characterize gene expression in PP and PN skin from psoriasis patients. cDNA (three new data sets) and cRNA hybridization (four existing data sets) data were compared using a common analysis pipeline. Agreement between data sets was assessed using varying qualitative and quantitative cut-offs to generate a DEG list in a source data set and then using other data sets to validate the list. Concordance increased from 67% across all probe sets to over 99% across more than 10,000 probe sets when statistical filters were employed. The fold-change behavior of individual genes tended to be consistent across the multiple data sets. We found that genes with <2-fold change values were quantitatively reproducible between pairs of data-sets. In a subset of transcripts with a role in inflammation changes detected by microarray were confirmed by qRT-PCR with high concordance. For transcripts with both PN and PP levels within the microarray dynamic range, microarray and qRT-PCR were quantitatively reproducible, including minimal fold-changes in IL13, TNFSF11, and TNFRSF11B and genes with >10-fold changes in either direction such as CHRM3, IL12B and IFNG. Conclusions/Significance Gene expression changes in psoriatic lesions were consistent across different studies, despite differences in patient selection, sample handling, and microarray platforms but between-study comparisons showed stronger agreement within than between platforms. We could use cut-offs as low as log10(ratio) = 0.1 (fold-change = 1.26), generating larger gene lists that validate on independent data sets. The reproducibility of PP signatures across data sets suggests that different sample sets can be productively compared.
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PURPOSE: We sought to investigate the effect of unloading and physical inactivity typical of the bed rest model by identifying leukocyte transcriptome changes in participants that underwent 60 days of bed rest followed by reambulation. Previous work from our lab utilized a time course analysis and identified temporal expression changes in 2,415 protein-coding transcripts (Stratis et al., 2022). Our current work is focused on selective time-point comparisons to reveal expression changes in both coding and non-coding genes specific to the bed rest and reambulation study phases. METHODS: This longitudinal study design collected ten blood samples from twenty healthy male participants. We measured gene expression through RNA sequencing of leukocytes and applied linear mixed modelling to assess differential expression at the following time-points: model 1, baseline data collection (BDC) (BDC-12 and BDC-11 combined) vs head-down tilt (HDT) bed rest (HDT1, HDT2, HDT30, HDT60); and model 2, HDT60 vs reambulation (R1, R2, R12, R30). RESULTS: Model 1 found 30/44 (68%) differentially expressed genes (α < 0.05 & log fold change>|1|) were between baseline and early bed rest (BDC-12/-11 vs HDT2). Model 2 found 24/37 (65%) differentially expressed genes were between late bed rest and early reambulation (HDT60 vs R1). CONCLUSIONS: Major transcriptome changes occurred early at the transitions to and from bed rest. Current findings can guide future work on the complex responses and adaptation mechanisms experienced during physical inactivity and unloaded environments.
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Temperatures worldwide have been rising due to global warming. The Korean grapevine (Vitis labruscana) cultivars are sensitive to high temperature during the vegetative growing period and are challenged with various physiological disorders, resulting in decreased production and poor quality of fruit. This study was conducted to identify genes that are differentially expressed in response to high temperature to help elucidate the mechanism of tolerance to temperatures in grapevines. ‘Kyoho’ grapevine leaves were maintained at 25°C and 35°C for 24 h in a plastic house and then used for transcriptome analysis by RNA-seq. More than 92 million transcripts were used for transcriptome analysis, with a total length of 7 billion bp and an average length of 89.6 bp; an average of 85.5% of the data were used in the final analysis. Investigation of differentially expressed genes (DEG) in response to high temperature discovered 243 induced genes and 378 inhibited genes. The selected DEGs were related to metabolism (49 genes), cell structure (22 genes), and molecular functions (15 genes). The selected DEGs clustered in the pathways of carbohydrate metabolism, amino acid metabolism, and the citrate cycle in the KEGG metabolic pathway. Among the selected DEGs, we selected the top 10 highly up-regulated and the top 10 down-regulated genes and confirmed their expression levels by reverse transcription quantitative PCR. At high temperature, the top 10 genes, including BCL-2-associated athanogene 6, heat shock protein 21, HAD superfamily, and subfamily IIIB acid phosphatase, showed induced expression, whereas the other 10 genes, including APS reductase3, ribosomal protein S7e family protein, and ribosomal protein L2 family, showed inhibited expression pattern. This is consistent with the results from the transcriptome analysis. We propose that these genes can potentially be used for the development of molecular markers to elucidate defense mechanisms against high temperature stress, and for breeding grapevine varieties tolerant to high temperature.
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Background The brown planthopper (BPH) Nilaparvata lugens (Stål) is one of the most serious insect pests of rice in Asia. However, little is known about the mechanisms responsible for the development, wing dimorphism and sex difference in this species. Genomic information for BPH is currently unavailable, and, therefore, transcriptome and expression profiling data for this species are needed as an important resource to better understand the biological mechanisms of BPH. Methodology/Principal Findings In this study, we performed de novo transcriptome assembly and gene expression analysis using short-read sequencing technology (Illumina) combined with a tag-based digital gene expression (DGE) system. The transcriptome analysis assembles the gene information for different developmental stages, sexes and wing forms of BPH. In addition, we constructed six DGE libraries: eggs, second instar nymphs, fifth instar nymphs, brachypterous female adults, macropterous female adults and macropterous male adults. Illumina sequencing revealed 85,526 unigenes, including 13,102 clusters and 72,424 singletons. Transcriptome sequences larger than 350 bp were subjected to Gene Orthology (GO) and KEGG Orthology (KO) annotations. To analyze the DGE profiling, we mainly compared the gene expression variations between eggs and second instar nymphs; second and fifth instar nymphs; fifth instar nymphs and three types of adults; brachypterous and macropterous female adults as well as macropterous female and male adults. Thousands of genes showed significantly different expression levels based on the various comparisons. And we randomly selected some genes to confirm their altered expression levels by quantitative real-time PCR (qRT-PCR). Conclusions/Significance The obtained BPH transcriptome and DGE profiling data provide comprehensive gene expression information at the transcriptional level that could facilitate our understanding of the molecular mechanisms from various physiological aspects including development, wing dimorphism and sex difference in BPH.
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Based on the assumption that severe alterations in the expression of genes known to be involved in high-density lipoprotein (HDL) metabolism may affect the expression of other genes, we screened an array of >5000 mouse expressed sequence tags for altered gene expression in the livers of two lines of mice with dramatic decreases in HDL plasma concentrations. Labeled cDNA from livers of apolipoprotein AI (apoAI)-knockout mice, scavenger receptor BI (SR-BI) transgenic mice, and control mice were cohybridized to microarrays. Two-sample t statistics were used to identify genes with altered expression levels in the knockout or transgenic mice compared with control mice. In the SR-BI group we found nine array elements representing at least five genes that were significantly altered on the basis of an adjusted P value < 0.05. In the apoAI-knockout group, eight array elements representing four genes were altered compared with the control group (adjusted P < 0.05). Several of the genes identified in the SR-BI transgenic suggest altered sterol metabolism and oxidative processes. These studies illustrate the use of multiple-testing methods for the identification of genes with altered expression in replicated microarray experiments.
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