scholarly journals Comparability of reference-based and reference-free transcriptome analysis approaches at the gene expression level

2021 ◽  
Vol 22 (S11) ◽  
Author(s):  
Sung-Gwon Lee ◽  
Dokyun Na ◽  
Chungoo Park
Keyword(s):  
Gene Expression ◽  
De Novo ◽  
Expression Profiles ◽  
Gene Families ◽  
Cell Types ◽  
Model Organisms ◽  
Expression Levels ◽  
Large Gene ◽  
Gene Expression Quantification ◽  
Gene Expression Levels

Abstract Background Lately, high-throughput RNA sequencing has been extensively used to elucidate the transcriptome landscape and dynamics of cell types of different species. In particular, for most non-model organisms lacking complete reference genomes with high-quality annotation of genetic information, reference-free (RF) de novo transcriptome analyses, rather than reference-based (RB) approaches, are widely used, and RF analyses have substantially contributed toward understanding the mechanisms regulating key biological processes and functions. To date, numerous bioinformatics studies have been conducted for assessing the workflow, production rate, and completeness of transcriptome assemblies within and between RF and RB datasets. However, the degree of consistency and variability of results obtained by analyzing gene expression levels through these two different approaches have not been adequately documented. Results In the present study, we evaluated the differences in expression profiles obtained with RF and RB approaches and revealed that the former tends to be satisfactorily replaced by the latter with respect to transcriptome repertoires, as well as from a gene expression quantification perspective. In addition, we urge cautious interpretation of these findings. Several genes that are lowly expressed, have long coding sequences, or belong to large gene families must be validated carefully, whenever gene expression levels are calculated using the RF method. Conclusions Our empirical results indicate important contributions toward addressing transcriptome-related biological questions in non-model organisms.

scholarly journals Single-cell RNA sequencing reveals the mesangial identity and species diversity of glomerular cell transcriptomes

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Bing He ◽  
Ping Chen ◽  
Sonia Zambrano ◽  
Dina Dabaghie ◽  
Yizhou Hu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Human Kidney ◽  
Cell Types ◽  
Model Organisms ◽  
Mural Cell ◽  
Glomerular Cell ◽  
Individual Gene ◽  
The Individual

AbstractMolecular characterization of the individual cell types in human kidney as well as model organisms are critical in defining organ function and understanding translational aspects of biomedical research. Previous studies have uncovered gene expression profiles of several kidney glomerular cell types, however, important cells, including mesangial (MCs) and glomerular parietal epithelial cells (PECs), are missing or incompletely described, and a systematic comparison between mouse and human kidney is lacking. To this end, we use Smart-seq2 to profile 4332 individual glomerulus-associated cells isolated from human living donor renal biopsies and mouse kidney. The analysis reveals genetic programs for all four glomerular cell types (podocytes, glomerular endothelial cells, MCs and PECs) as well as rare glomerulus-associated macula densa cells. Importantly, we detect heterogeneity in glomerulus-associated Pdgfrb-expressing cells, including bona fide intraglomerular MCs with the functionally active phagocytic molecular machinery, as well as a unique mural cell type located in the central stalk region of the glomerulus tuft. Furthermore, we observe remarkable species differences in the individual gene expression profiles of defined glomerular cell types that highlight translational challenges in the field and provide a guide to design translational studies.

scholarly journals Evaluation of TGF-Beta 2 and VEGFα Gene Expression Levels in Epiretinal Membranes and Internal Limiting Membranes in the Course of Retinal Detachments, Proliferative Diabetic Retinopathy, Macular Holes, and Idiopathic Epiretinal Membranes

2018 ◽  
Vol 2018 ◽  
pp. 1-6
Author(s):  
Joanna Stafiej ◽  
Karolina Kaźmierczak ◽  
Katarzyna Linkowska ◽  
Paweł Żuchowski ◽  
Tomasz Grzybowski ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Profiles ◽  
Rna Extraction ◽  
Retinal Diseases ◽  
Epiretinal Membranes ◽  
Expression Levels ◽  
Macular Holes ◽  
Retinal Disorders ◽  
Pars Plana ◽  
Gene Expression Levels

Purpose. To evaluate the expression profiles of the VEGFα and TGFβ in the ERMs and ILMs in retinal disorders. Methods. In this nonrandomized prospective study, 75 patients (34 females and 41 males) referred to pars plana vitrectomy (PPV) due to different retinal diseases were enrolled to the study. The samples of ERMs and ILMs collected during PPV were immediately put in TRIzol® Reagent (Life Technologies, USA) and stored at −70°C until RNA extraction. Gene expression analysis was done with TaqMan® Gene Expression Assays (Applied Biosystems, USA) following the manufacturer’s instructions. Results. The gene expression levels of VEGFα as well as of TGFβ2 were significantly higher in ERMs than in ILMs in all studied groups. The level of TGFβ2 expression exhibits a significantly lower values in iERMs as compared with the RRD group (p=0.043). There were differences in TGFβ2 expression in ILM in groups studied: DR versus RRD, p=0.003; DR versus iERM, p=0,047; and iERM versus RRD, p=0.004. Conclusions. Our results revealed that factors associated with angiogenesis and wound healing processes in eyes with RRD, PDR, iERM, and MH were more upregulated in ERMs than in ILMs. This may indicate that ILM is not responsible for reproliferation and its peeling should be avoided in routine PPV.

scholarly journals High-throughput mRNA sequencing of stromal cells from endometriomas and endometrium

Reproduction ◽  
2017 ◽  
Vol 154 (1) ◽  
pp. 93-100 ◽  
Author(s):  
Kadri Rekker ◽  
Merli Saare ◽  
Elo Eriste ◽  
Tõnis Tasa ◽  
Viktorija Kukuškina ◽  
...  
Keyword(s):  
Gene Expression ◽  
High Throughput ◽  
Complement System ◽  
Stromal Cells ◽  
Cell Types ◽  
Coagulation Cascade ◽  
Expression Levels ◽  
Mrna Sequencing ◽  
Standard Design ◽  
Gene Expression Levels

The aetiology of endometriosis is still unclear and to find mechanisms behind the disease development, it is important to study each cell type from endometrium and ectopic lesions independently. The objective of this study was to uncover complete mRNA profiles in uncultured stromal cells from paired samples of endometriomas and eutopic endometrium. High-throughput mRNA sequencing revealed over 1300 dysregulated genes in stromal cells from ectopic lesions, including several novel genes in the context of endometriosis. Functional annotation analysis of differentially expressed genes highlighted pathways related to cell adhesion, extracellular matrix–receptor interaction and complement and coagulation cascade. Most importantly, we found a simultaneous upregulation of complement system components and inhibitors, indicating major imbalances in complement regulation in ectopic stromal cells. We also performed in vitro experiments to evaluate the effect of endometriosis patients’ peritoneal fluid (PF) on complement system gene expression levels, but no significant impact of PF on C3, CD55 and CFH levels was observed. In conclusion, the use of isolated stromal cells enables to determine gene expression levels without the background interference of other cell types. In the future, a new standard design studying all cell types from endometriotic lesions separately should be applied to reveal novel mechanisms behind endometriosis pathogenesis.

Expression profiles of metamorphosis-related genes during natural transformations in tadpoles of wild Wood Frogs (Lithobates sylvaticus)

2012 ◽  
Vol 90 (9) ◽  
pp. 1059-1071 ◽  
Author(s):  
Laia Navarro-Martín ◽  
Chantal Lanctôt ◽  
Christopher Edge ◽  
Jeff Houlahan ◽  
Vance L. Trudeau
Keyword(s):  
Gene Expression ◽  
Expression Profiles ◽  
Rana Sylvatica ◽  
Fold Increase ◽  
Gonadal Development ◽  
Specific Expression ◽  
Wood Frogs ◽  
Lithobates Sylvaticus ◽  
Expression Levels ◽  
Gene Expression Levels

Numerous studies using laboratory-reared tadpoles have shown the importance of thyroid hormones (TH), thyroid receptors (TR), and deiodinase (Dio) enzymes during anuran metamorphosis. Our study focuses on the analysis of thyroid-related genes in tadpoles of wild Wood Frogs ( Lithobates sylvaticus (LeConte, 1825); also known as Rana sylvatica (Cope, 1889)) during metamorphosis. Results showed that, in concordance with laboratory-reared studies, thyroid receptor beta (trb) gene expression profiles presented the most marked changes. At climax and compared with premetamorphic stages, brains, tails, and gonad–mesonephros complex (GMC) tissues increased trb expression levels 5-, 21-, and 41-fold, respectively (p < 0.05). In addition, gene expression levels of brain deiodinase type II and III showed opposite trends, where 3-fold decrease and 10-fold increase were, respectively, found. This finding supports the idea that thyroid hormone, as it has been demonstrated in laboratory-reared tadpoles, is also involved in natural metamorphosis in wild tadpoles. Interestingly, and contrary to our predictions, we observed that whole brain corticotropin-releasing factor (crf) and crf receptor 1 (crfr1) gene expression levels significantly decrease through metamorphosis in wild L. sylvaticus tadpoles. Further analyses are required to determine if a role of TH in the timing of anuran gonadal development exists, as well as the importance of cell-specific and tissue-specific expression of crf and crfr1 to metamorphosis.

Identification of Aberrantly Regulated Genes through Comparison of Genome-Wide Expression Profiles and Karyotype in Hyperdiploid >50 Chromosomes Pediatric Acute Lymphoblastic Leukemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 420-420
Author(s):  
Christian Flotho ◽  
Susana C. Raimondi ◽  
James R. Downing
Keyword(s):  
Gene Expression ◽  
Copy Number ◽  
Expression Profiles ◽  
Chromosome 21 ◽  
Small Subset ◽  
Specific Gene ◽  
Expression Levels ◽  
Number Of Genes ◽  
The Mean ◽  
Gene Expression Levels

Abstract We have demonstrated that expression profiling of leukemic blasts can accurately identify the known prognostic subtypes of ALL, including T-ALL, E2A-PBX1, TEL-AML1, MLL rearrangements, BCR-ABL, and hyperdiploid &gt;50 chromosomes (HD&gt;50). Interestingly, almost 70% of the genes that defined HD&gt;50 ALL localized to chromosome 21 or X. To further explore the relationship between gene expression and chromosome dosage, we compared the expression profiles obtained using the Affymetrix U133A&B microarrays of 17 HD&gt;50 ALLs to 78 diploid or pseudodiploid ALLs. Our analysis demonstrated that the average expression level for all genes on a chromosome could be used to predict chromosome copy numbers. Specifically, the copy number for each chromosome calculated by gene expression profiling predicted the numerical chromosomal abnormalities detected by standard cytogenetics. For chromosomes that were trisomic in HD&gt;50 ALL, the mean chromosome-specific gene expression level was increased approximately 1.5-fold compared to that observed in diploid or pseudodiploid ALL cases. Similarly, for chromosome 21 and X, the mean chromosome-specific gene expression levels were increased approximately 2-fold, consistent with a duplication of the active X chromosome and tetrasomy of chromosome 21, a finding verified by standard cytogenetics in &gt;90% of the HD&gt;50 cases. These finding indicate that the aberrant gene expression levels seen in HD&gt;50 ALL primarily reflect gene dosages. Importantly, we did not observe any clustering of aberrantly expressed genes across the duplicated chromosomes, making regional gain or loss of genomic material unlikely. Paradoxically, however, a more detailed analysis revealed a small but statistically significant number of genes on the trisomic/tetrasomic chromosomes whose expression levels were markedly reduced when compared to that seen in diploid or pseudodiploid leukemic samples. Using the Statistical Analysis of Microarrays (SAM) algorithm we identified 20 genes whose expression was reduced &gt;2-fold despite having an increase in copy number. Interestingly, included within this group are several known tumor suppressors, including AKAP12, which is specifically silenced by methylation in fos-transformed cells, and IGF2R and IGFBP7, negative regulators of insulin-like growth factor signaling. In addition to the silencing of a small subset of genes, we also identified 21 genes on these chromosomes whose expression levels were markedly higher (&gt;3-fold) than would be predicted solely based on copy number. Although the mechanism responsible for their increased expression remains unknown, included in this group are four genes involved in signal transduction (IL3RA, IL13RA1, SNX9, and GASP) and a novel cytokine, C17, whose expression is normally limited to CD34+ hematopoietic progenitors. Taken together, these data suggest that aberrant growth in HD&gt;50 ALL is in part driven by increased expression of a large number of genes secondary to chromosome duplications, coupled with a further enhanced expression of a limited number of growth promoting genes, and the specific silencing of a small subset of negative growth regulatory genes. Understanding the mechanisms responsible for the non-dosage related changes in gene expression should provide important insights into the pathology of HD&gt;50 ALL.

Genetic control of global gene expression levels in the intestinal mucosa: a human twin study

2009 ◽  
Vol 38 (1) ◽  
pp. 73-79 ◽  
Author(s):  
Robert Häsler ◽  
Alexander Begun ◽  
Sandra Freitag-Wolf ◽  
Martin Kerick ◽  
Nancy Mah ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mrna Expression ◽  
Genetic Control ◽  
Intestinal Mucosa ◽  
Expression Profiles ◽  
Global Gene Expression ◽  
Model Organisms ◽  
Expression Levels ◽  
Molecular Phylogenetic ◽  
Health And Disease

Phenotypic variation between individuals, such as different mRNA expression levels, is influenced by genetic and nongenetic factors. Although several studies have addressed the interplay between genotypes and expression profiles in various model organisms in the recent years, the detailed and relative contributions of genetic and nongenetic factors in regulating plasticity of gene expression in barrier organs (e.g., skin, gut), which are exposed to continuous environmental challenge, are still poorly understood. Here we systematically monitored the level of genetic control over genomewide mRNA expression profiles in the healthy intestinal mucosa of 10 monozygotic and 10 dizygotic human twin pairs with microarray analyses. Our results, which are supported by real-time PCR and analysis of molecular phylogenetic conservation, indicate that genes associated with energy metabolism and cell and tissue regeneration pathways are under strong genetic control. Conversely, genes associated with immune response seem to be mainly controlled by exogenous factors. Further insights into the relative extent of genetic and nongenetic determinants of transcriptomal profiles and their influence on physiological and pathophysiological mechanisms are crucial to understanding the key role played by gene-environment interactions in health and disease.

scholarly journals Porcine colonoids and enteroids keep the memory of their origin during regeneration.

2021 ◽  
Author(s):  
Alicia M. Barnett ◽  
Jane A. Mullaney ◽  
Charlotte Hendriks ◽  
Lisa Le Borgne ◽  
Warren C. McNabb ◽  
...  
Keyword(s):  
Gene Expression ◽  
Glucose Transporter ◽  
Expression Profiles ◽  
Cell Types ◽  
Specific Cell ◽  
Stem Cell Markers ◽  
Culture Methods ◽  
Glucose Transporter 1 ◽  
Expression Levels

The development of alternative in vitro culture methods has increased in the last decade as three-dimensional organoids of various tissues, including those of the small and large intestines. Due to their multicellular composition, organoids offer advantages over traditionally used immortalized or primary cell lines. However, organoids must be accurate models of their tissues of origin. This study compared gene expression profiles with respect to markers of specific cell-types (stem-cells, enterocytes, goblet and enteroendocrine cells) and barrier maturation (tight junctions) of colonoid and enteroid cultures with their tissues of origin, and colonoids with enteroids. Colonoids derived from three healthy pigs formed multi-lobed structures with a monolayer of cells similar to the crypt structures in colonic tissue. Colonoid and enteroid gene expression signatures were more similar to those found for the tissues of their origin than to each other. However, relative to their derived tissues, organoids had increased gene expression levels of stem-cell markers Sox9 and Lgr5 encoding Sex determining region Y-box 9 and leucine-rich repeat-containing G-protein coupled rector 5, respectively. In contrast, expression levels of Occl and Zo1 encoding occludin and zonula occludens 1 respectively, were decreased. Expression levels of the cell lineage markers Atoh1, Cga and Muc2 encoding atonal homolog 1, chromogranin A and mucin 2 respectively, were decreased in colonoids, while Sglt1 and Apn encoding sodium-glucose transporter 1 and aminopeptidase A respectively, were decreased in enteroids. These results indicate colonoid and enteroid cultures were predominantly comprised of undifferentiated cell-types with decreased barrier maturation relative to their tissues of origin.

scholarly journals Gene expression is encoded in all parts of a co-evolving interacting gene regulatory structure

2019 ◽  
Author(s):  
Jan Zrimec ◽  
Filip Buric ◽  
Azam Sheikh Muhammad ◽  
Rhongzen Chen ◽  
Vilhelm Verendel ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Expression Regulation ◽  
Model Organisms ◽  
Regulatory Structure ◽  
Expression Levels ◽  
Coding Regions ◽  
Dna Regulatory Motifs ◽  
Gene Regulatory ◽  
The Individual ◽  
Gene Expression Levels

AbstractUnderstanding the genetic regulatory code that governs gene expression is a primary, yet challenging aspiration in molecular biology that opens up possibilities to cure human diseases and solve biotechnology problems. However, the fundamental question of how each of the individual coding and non-coding regions of the gene regulatory structure interact and contribute to the mRNA expression levels remains unanswered. Considering that all the information for gene expression regulation is already present in living cells, here we applied deep learning on over 20,000 mRNA datasets in 7 model organisms ranging from bacteria to Human. We show that in all organisms, mRNA abundance can be predicted directly from the DNA sequence with high accuracy, demonstrating that up to 82% of the variation of gene expression levels is encoded in the gene regulatory structure. Coding and non-coding regions carry both overlapping and orthogonal information and additively contribute to gene expression levels. By searching for DNA regulatory motifs present across the whole gene regulatory structure, we discover that motif interactions can regulate gene expression levels in a range of over three orders of magnitude. The uncovered co-evolution of coding and non-coding regions challenges the current paradigm that single motifs or regions are solely responsible for gene expression levels. Instead, we show that the correct combination of all regulatory regions must be established in order to accurately control gene expression levels. Therefore, the holistic system that spans the entire gene regulatory structure is required to analyse, understand, and design any future gene expression systems.

scholarly journals Ampliconic Genes on the Great Ape Y Chromosomes: Rapid Evolution of Copy Number but Conservation of Expression Levels

2020 ◽  
Vol 12 (6) ◽  
pp. 842-859
Author(s):  
Rahulsimham Vegesna ◽  
Marta Tomaszkiewicz ◽  
Oliver A Ryder ◽  
Rebeca Campos-Sánchez ◽  
Paul Medvedev ◽  
...  
Keyword(s):  
Gene Expression ◽  
Y Chromosome ◽  
Copy Number ◽  
Gene Families ◽  
Great Apes ◽  
Published Data ◽  
Great Ape ◽  
Data Set ◽  
Expression Levels ◽  
Gene Expression Levels

Abstract Multicopy ampliconic gene families on the Y chromosome play an important role in spermatogenesis. Thus, studying their genetic variation in endangered great ape species is critical. We estimated the sizes (copy number) of nine Y ampliconic gene families in population samples of chimpanzee, bonobo, and orangutan with droplet digital polymerase chain reaction, combined these estimates with published data for human and gorilla, and produced genome-wide testis gene expression data for great apes. Analyzing this comprehensive data set within an evolutionary framework, we, first, found high inter- and intraspecific variation in gene family size, with larger families exhibiting higher variation as compared with smaller families, a pattern consistent with random genetic drift. Second, for four gene families, we observed significant interspecific size differences, sometimes even between sister species—chimpanzee and bonobo. Third, despite substantial variation in copy number, Y ampliconic gene families’ expression levels did not differ significantly among species, suggesting dosage regulation. Fourth, for three gene families, size was positively correlated with gene expression levels across species, suggesting that, given sufficient evolutionary time, copy number influences gene expression. Our results indicate high variability in size but conservation in gene expression levels in Y ampliconic gene families, significantly advancing our understanding of Y-chromosome evolution in great apes.

scholarly journals CellExpress: a comprehensive microarray-based cancer cell line and clinical sample gene expression analysis online system

Database ◽  
2018 ◽  
Vol 2018 ◽  
Author(s):  
Yi-Fang Lee ◽  
Chien-Yueh Lee ◽  
Liang-Chuan Lai ◽  
Mong-Hsun Tsai ◽  
Tzu-Pin Lu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Lines ◽  
Cancer Cell ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Comprehensive Analysis ◽  
Next Generation Sequencing Data ◽  
Clinical Samples ◽  
Expression Levels ◽  
Gene Expression Levels

Abstract With the advancement of high-throughput technologies, gene expression profiles in cell lines and clinical samples are widely available in the public domain for research. However, a challenge arises when trying to perform a systematic and comprehensive analysis across independent datasets. To address this issue, we developed a web-based system, CellExpress, for analyzing the gene expression levels in more than 4000 cancer cell lines and clinical samples obtained from public datasets and user-submitted data. First, a normalization algorithm can be utilized to reduce the systematic biases across independent datasets. Next, a similarity assessment of gene expression profiles can be achieved through a dynamic dot plot, along with a distance matrix obtained from principal component analysis. Subsequently, differentially expressed genes can be visualized using hierarchical clustering. Several statistical tests and analytical algorithms are implemented in the system for dissecting gene expression changes based on the groupings defined by users. Lastly, users are able to upload their own microarray and/or next-generation sequencing data to perform a comparison of their gene expression patterns, which can help classify user data, such as stem cells, into different tissue types. In conclusion, CellExpress is a user-friendly tool that provides a comprehensive analysis of gene expression levels in both cell lines and clinical samples. The website is freely available at http://cellexpress.cgm.ntu.edu.tw/. Source code is available at https://github.com/LeeYiFang/Carkinos under the MIT License. Database URL: http://cellexpress.cgm.ntu.edu.tw/

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