Transcription profiling of human fibroblasts and lymphoblastoid cells derived from individuals with Bipolar disorder vs. unaffected
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ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Maja Bucan
PROVIDER: E-GEOD-11767 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Fibroblasts and lymphoblastoid cells (LCLs) are the most widely used cells in genetic, genomic, and transcriptomic studies in relation to human diseases. Examining the gene expression patterns in these two cell types will provide valuable information regarding the validity of using them to study gene expression related to various human diseases. Fibroblasts and LCLs from four members of the Old Order Amish family 884 were purchased from Coriell cell repositories (Coriell Institute for Medical Research, Camden, NJ). We used microarrays to profile the patterns of gene expression in these eight cell lines. By employing the PennCNV algorithm to the Illumina HumanHap550 SNP genotyping data, we detected 13 Copy Number Variants (CNV) that exist in these four individuals. CNV-expression association analysis revealed that seven of these 13 CNVs were associated with the expression of genes within or near (<2Mb sweep) these CNVs at a nominal regression P value of 0.05.
2008-06-13 | GSE11767 | GEO
Project description:Epstein-Barr virus (EBV) transformed lymphoblastoid cell lines (LCLs) are a widely used renewable resource for functional genomic studies in humans. The ability to accumulate multidimensional data pertaining to the same individual cell lines, from complete genomic sequences to detailed gene regulatory profiles, further enhances the utility of LCLs as a model system. However, the extent to which LCLs are a faithful model system is relatively unknown. We have previously shown that gene expression profiles of newly established LCLs maintain a strong individual component. Here, we extend our study to investigate the effect of freeze-thaw cycles on gene expression patterns in mature LCLs, especially in the context of inter-individual variation in gene regulation. We found a profound difference in the gene expression profiles of newly established and mature LCLs. Once newly established LCLs undergo a freeze-thaw cycle, the individual specific gene expression signatures become much less pronounced as the gene regulatory programs in LCLs from different individuals converge to a more uniform profile, which reflects a mature transformed B cell phenotype. As expected, previously identified eQTLs are enriched among the relatively few genes whose regulations in mature LCLs maintain marked individual signatures. We thus conclude that findings and insight drawn from gene regulatory studies in mature LCLs are generally not affected by artificial nature of the LCL model system and are likely to faithfully reflect regulatory interactions in primary tissues. However, our data indicate that many aspects of primary B cell biology cannot be observed and studied in mature LCL cultures.
2014-09-05 | GSE58942 | GEO
Project description:The following cell lines were used for this study: Epstein-Barr virus (EBV) transformed lymphoblastoid cell lines (LCLs) derived from 5 human (Coriell YRI, NIGMS Human Genetic Cell Repository, GM18505, GM18507, GM18516, GM19193, GM19204), and 5 chimpanzee (Pan troglodytes) individuals (New Iberia Research Center: Min 18358, Min 18359; Coriell/IPBIR: NS03659, NS04973, Arizona State University, Pt91), and rhesus Herpesvirus papio transformed LCLs from 5 rhesus macaque (Macaca mulatta) individuals (Harvard Medical School, NEPRC: 150-99, R181-96, R249-97, 265-95, R290-96). Cells were maintained at identical conditions of 37° with 5% CO2 in RPMI media with 15% FBS, supplemented with 2 mM L-glutamate, 100 IU/ml penicillin, and 100 ug/ml streptomycin. Internal standard LCL (Coriell YRI, NIGMS Human Genetic Cell Repository, GM19238) was grown in RPMI minus L-Lysine and L-Arginine, 15% dialyzed FBS, and L-13C615N4-arginine (Arg-10) and L-13C615N2-lysine (Lys-8) (Cambridge Isotopes, Andover, MA, USA) supplemented with 2 mM L-glutamate, 100 IU/ml penicillin, and 100 ug/ml streptomycin under identical conditions as the unlabeled LCLs. The internal standard LCL was grown for 6 doublings to assure complete SILAC label incorporation. Complete label incorporation was verified by analyzing the protein lysate from the labeled LCL alone by high-resolution LC-MS/MS. LCLs were washed in PBS three times and then lysed using the UPX Universal Protein Extraction Kit (Expedeon Inc., San Diego, CA, USA). Protein quantitation was performed using the Qubit fluorometry assay (Invitrogen, Carlsbad California, USA) and the reducing agent-compatible (RAC) version of the BCA Protein Assay (Thermo Scientific, Waltham, Massachusetts, USA). 12ug of each sample was combined with 12ug of the SILAC labeled lysate from human LCL GM19238. Note that the SILAC lysate was prepared once and used as an internal standard through the quantification of the 15 cell lines. 24ug of each combined sample was then processed by SDS-PAGE using a 4-12% Bis Tris NuPage mini-gel (Invitrogen, Carlsbad California, USA). Calibration was with Thermo PageRuler broad range markers. Each of 40 gel segments were processed by in-gel digestion using a ProGest robot (DigiLab, Marlborough, MA, USA) with the following protocol: wash with 25mM ammonium bicarbonate followed by acetonitrile, reduce with 10mM dithiothreitol at 60°C followed by alkylation with 50mM iodoacetamide at room temperature, digest with trypsin (Promega, Madison, WI, USA) at 37°C for 4h, and quench with formic acid. The supernatant was analyzed directly without further processing. Each of gel digest was analyzed by nano-LC/MS/MS with a Waters NanoAcquity HPLC system interfaced to a ThermoFisher LTQ-Orbitrap Velos Pro. Peptides were loaded on a trapping column and eluted over a 75um analytical column at 350nL/min using a 1-hour LC gradient. Both columns were packed with Jupiter Proteo resin (Phenomenex, Torrance, California, USA). The mass spectrometer was operated in data-dependent mode, with MS performed in the Orbitrap at 60,000 FWHM resolution and MS/MS performed in the LTQ. The fifteen most abundant ions were selected for MS/MS. Low-level data analysis was performed using the open-source proteomics software tool PVIEW (Release December 23, 2012; http://compbio.cs.princeton.edu/pview). As input to PVIEW, we generated in silico translations of coding genes from the UCSC Genome Browser database based on gene models from build hg19 of the human genome. Each protein sequence entry retained the corresponding Ensembl gene identifier and gene symbol. Database searches were performed using +-4 p.p.m. MS1 tolerance and an MS2 window tolerance of +-0.5 Da. Up to 2 missed tryptic cleavages were allowed during search. Carboxyamidomethylation of cysteine was used as fixed modification. Up to two methionine oxidations were allowed as variable modifications of a tryptic peptide. Peptide spectrum matches were obtained at a stringent false discovery rate (FDR) of 1%. We used the median log2(sample/standard) ratio across all independent quantifications of a protein (distinct peptides including duplicate peptide measurements across fractions and for differing charge states). Note that use of the hg19 database was sufficient as SILAC pairs as the expected isotope shift used for quantification were only present for peptides that that had the same underlying sequence in the human internal standard line. Associated RNA-seq data have been deposited to GEO with accession number GSE49682.
2013-10-01 | PXD000419 | Pride
Project description:Renewable in vitro cell cultures, such as lymphoblastoid cell lines (LCLs), have facilitated studies that contributed to our understanding of genetic influence on human traits. However, the degree to which cell lines faithfully maintain differences in donor-specific phenotypes is still debated. We have previously reported that standard cell line maintenance practice results in a loss of donor-specific gene expression signatures in LCLs. An alternative to the LCL model is the induced pluripotent stem cell (iPSC) system, which carries the potential to model tissue-specific physiology through the use of differentiation protocols. Still, existing LCL banks represent an important source of starting material for iPSC generation, and it is possible that the disruptions in gene regulation associated with long-term LCL maintenance could persist through the reprogramming process. To address this concern, we studied the effect of reprogramming mature LCLs to iPSCs on the ensuing gene expression patterns within and between six unrelated donor individuals. We show that the reprogramming process results in a recovery of donor-specific gene regulatory signatures. Since environmental contributions are unlikely to be a source of individual variation in our system of highly passaged cultured cell lines, our observations suggest that the effect of genotype on gene regulation is more pronounced in the iPSCs than in the LCL precursors. Our observations indicate that iPSCs can be a powerful model system for studies of phenotypic variation across individuals in general, and the genetic association with variation in gene regulation in particular. We further conclude that LCLs are an appropriate starting material for iPSC generation.
2014-12-17 | GSE64263 | GEO
Project description:Renewable in vitro cell cultures, such as lymphoblastoid cell lines (LCLs), have facilitated studies that contributed to our understanding of genetic influence on human traits. However, the degree to which cell lines faithfully maintain differences in donor-specific phenotypes is still debated. We have previously reported that standard cell line maintenance practice results in a loss of donor-specific gene expression signatures in LCLs. An alternative to the LCL model is the induced pluripotent stem cell (iPSC) system, which carries the potential to model tissue-specific physiology through the use of differentiation protocols. Still, existing LCL banks represent an important source of starting material for iPSC generation, and it is possible that the disruptions in gene regulation associated with long-term LCL maintenance could persist through the reprogramming process. To address this concern, we studied the effect of reprogramming mature LCLs to iPSCs on the ensuing gene expression patterns within and between six unrelated donor individuals. We show that the reprogramming process results in a recovery of donor-specific gene regulatory signatures. Since environmental contributions are unlikely to be a source of individual variation in our system of highly passaged cultured cell lines, our observations suggest that the effect of genotype on gene regulation is more pronounced in the iPSCs than in the LCL precursors. Our observations indicate that iPSCs can be a powerful model system for studies of phenotypic variation across individuals in general, and the genetic association with variation in gene regulation in particular. We further conclude that LCLs are an appropriate starting material for iPSC generation. Whole genome gene expression data was collected for 34 samples including 17 iPSC lines and 17 LCL lines on the Illumina HT-12 v4 Expression BeadChip array platform. Three biological replicates of each individual were included in the study, except for one individual for which only two replicates were obtained.
2014-12-17 | E-GEOD-64263 | biostudies-arrayexpress
Project description:Copy-number variants (CNVs) are large-scale amplifications or deletions of DNA that can drive rapid adaptive evolution and result in large-scale changes in gene expression. Whereas alterations in the copy number of one or more genes within a CNV can confer a selective advantage, other genes within a CNV can decrease fitness when their dosage is changed. Dosage compensation - in which the gene expression output from multiple gene copies is less than expected - is one means by which an organism can mitigate the fitness costs of deleterious gene amplification. Previous research has shown evidence for dosage compensation at both the transcriptional level and at the level of protein expression; however, the extent of compensation differs substantially between genes, strains, and studies. Here, we investigated sources of dosage compensation at multiple levels of gene expression regulation by defining the transcriptome, translatome and proteome of experimentally evolved yeast (Saccharomyces cerevisiae) strains containing adaptive CNVs.
2023-11-01 | PXD046587 | Pride
Project description:Copy number variants (CNVs) are currently defined as genomic sequences that are polymorphic in copy number and range in length from 1,000 to several million base pairs. Among current array-based CNV detection platforms, long-oligonucleotide arrays promise the highest resolution. However, the performance of currently available analytical tools suffers when applied to these data because of the lower signal:noise ratio inherent in oligonucleotide-based hybridization assays. We have developed wuHMM, an algorithm for mapping CNVs from array comparative genomic hybridization (aCGH) platforms comprised of 385,000 to more than 3 million probes. wuHMM is unique in that it can utilize sequence divergence information to reduce the false positive rate (FPR). We apply wuHMM to 385K-aCGH, 2.1M-aCGH, and 3.1M-aCGH experiments comparing the 129X1/SvJ and C57BL/6J inbred mouse genomes. We assess wuHMM’s performance on the 385K platform by comparison to the higher resolution platforms and we independently validate 10 CNVs. The method requires no training data and is robust with respect to changes in algorithm parameters. At a FPR of less than 10%, the algorithm can detect CNVs with five probes on the 385K platform and three on the 2.1M and 3.1M platforms, resulting in effective resolutions of 24 kb, 2-5 kb, and 1 kb, respectively. Keywords: CNV detection algorithm development and assessment
2008-03-12 | GSE10511 | GEO
Project description:Variation in gene expression is a fundamental aspect of human phenotypic variation. Several studies have analyzed gene expression levels in populations of different continental ancestry, and concluded that there is variation across populations at a fraction of expressed genes. Here we analyze gene expression levels in African American cell lines, which differ from previously analyzed cell lines in that samples from this population have variable proportions of continental ancestry. We show that for most genes examined, gene expression varies with genetic ancestry. Experiment Overall Design: Lymphoblastoid cell lines (LCL) for 60 HapMap CEU, 60 HapMap YRI, and 82 AFA from the Human Variation Panel were obtained from Coriell Cell Repositories. LCLs were grown in culture, total RNA was extracted and hybridized to Affymetrix HG-FOCUS arrays.
2009-01-14 | E-GEOD-10824 | biostudies-arrayexpress
Project description:DNA from Epstein-Barr virus (EBV)-transformed lymphocyte cell lines (LCLs) has proven very useful for studies of genetic sequence polymorphisms. Whether EBV-LCL DNA is suitable for methylation studies is less clear. We conduct a genome-wide methylation investigation using an array set with 45 million probes to investigate the methylome of EBV-LCL DNA and technical duplicates of whole blood (WB) DNA from the same 10 individuals. Methylation sites that show variation between individuals are potentially useful as biomarkers in disease studies. Our comparison is, therefore, focused specifically on these methylation variable sites. The sample correlations (i.e., a measure of whether the rank of the signals remains consistent between two samples from the same individual) for the methylation variable probes ranged from 0.69-0.78 for the WB duplicates and from 0.27-0.72 for WB versus EBV-LCL. To compare the pattern of the methylation signals, we grouped adjacent probes based on their inter-correlations. These analyses showed ~29,000 blocks in WB and ~14,000 blocks in EBV-LCL. Furthermore, merely 31% of the methylated regions detected in WB were detectable in EBV-LCLs. Our study shows that there are substantial differences in the DNA methylation patterns between EBV-LCL and WB. Thus, EBV-LCL DNA cannot be used as a proxy for WB DNA in methylation studies.
2012-03-08 | GSE35204 | GEO
Project description:Epstein-Barr virus (EBV) transformed lymphoblastoid cell lines (LCLs) are a widely used renewable resource for functional genomic studies in humans. The ability to accumulate multidimensional data pertaining to the same individual cell lines, from complete genomic sequences to detailed gene regulatory profiles, further enhances the utility of LCLs as a model system. However, the extent to which LCLs are a faithful model system is relatively unknown. We have previously shown that gene expression profiles of newly established LCLs maintain a strong individual component. Here, we extend our study to investigate the effect of freeze-thaw cycles on gene expression patterns in mature LCLs, especially in the context of inter-individual variation in gene regulation. We found a profound difference in the gene expression profiles of newly established and mature LCLs. Once newly established LCLs undergo a freeze-thaw cycle, the individual specific gene expression signatures become much less pronounced as the gene regulatory programs in LCLs from different individuals converge to a more uniform profile, which reflects a mature transformed B cell phenotype. As expected, previously identified eQTLs are enriched among the relatively few genes whose regulations in mature LCLs maintain marked individual signatures. We thus conclude that findings and insight drawn from gene regulatory studies in mature LCLs are generally not affected by artificial nature of the LCL model system and are likely to faithfully reflect regulatory interactions in primary tissues. However, our data indicate that many aspects of primary B cell biology cannot be observed and studied in mature LCL cultures. We obtained unpurified buffy coats (of a unit of blood) from six unrelated healthy individuals of Caucasian ethnicity (age range: 20-45). We isolated CD20+ B cells and established six independent cultures of LCLs between October 2009 and January 2010. From each of these samples, we obtained genome wide gene expression data using Illumina HumanHT-12 v3 Expression BeadChip arrays. We refer to data from these experiments as M-bM-^@M-^\cycle 0M-bM-^@M-^] to acknowledge the fact that the LCLs from this cycle were newly established and therefore not frozen/thawed. Between February 2011 and October 2012, we thawed each of these LCL cultures every 3 months and cultured them until obtaining ~10 million cells (February 2011=cycle 1, June 2011=cycle 2, October 2011=cycle 3, February 2012=cycle 4, June 2012=cycle 5, and October 2012=cycle 6). We used Illumina HumanHT-12 v4 Expression BeadChip arrays and obtained genome wide gene expression data on cycle 2, cycle 4 and cycle 6 LCLs. At each hybridization batch we included a subset of RNA samples that were also hybridized in a previous batch. This allowed us to effectively consider the batch effect on gene expression profiles. Our final dataset included genome wide gene expression data from 187 samples collected at 4 different time points.
2014-09-05 | E-GEOD-58942 | biostudies-arrayexpress