Objectives: To identify similarities and differences in gene expression data in the MEK/ERK and PI3K pathways and to determine how histone modification affects these same pathways.
Regulation of gene expression by PI3K in mouse growth plate chondrocytes.
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View SamplesPrimary micromass cultures derived from 11.5 day old mouse embryo limb buds were cultured for 15 days in differentiating conditions (beta-glycerophosphate and ascorbic acid). Total RNA from differentiating chondrocytes was isolated every three days i.e. days 3,6,9,12 and 15 and hybridized to MOE430A chips. Objective: Gain a view of the temporal gene expression changes occuring during chondrocyte differentiation.
Microarray analyses of gene expression during chondrocyte differentiation identifies novel regulators of hypertrophy.
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View SamplesBackground: Glucocorticoids (GCs) are widely used anti-inflammatory drugs. While useful in clinical practice, patients taking GCs often suffer from skeletal side effects including growth retardation and decreased bone quality in adults. On a physiological level, GCs have been implicated in the regulation of chondrogenesis and osteoblast differentiation, as well as maintaining homeostasis in cartilage and bone. We identified the glucocorticoid receptor (GR) as a potential regulator of chondrocyte hypertrophy in a microarray screen of primary limb bud mesenchyme micromass cultures. Some targets of GC regulation in chondrogenesis are known, but the global effects of pharmacological GC doses on chondrocyte gene expression have not been comprehensively evaluated.
Expression profiling of Dexamethasone-treated primary chondrocytes identifies targets of glucocorticoid signalling in endochondral bone development.
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View SamplesA variety of cell cultures models and in vivo approaches have been used to study gene expression during chondrocyte differentiation. The extent to which the in vitro models reflect bona fide gene regulation in the growth plate has not been quantified. In addition, studies that evaluate global gene expression changes among different growth plate zones are limited. To address these issues, we completed a microarray screen of three growth plate zones derived from manually segmented embryonic mouse tibiae. Classification of genes differentially expressed between each respective growth plate zone, functional categorization as well as characterization of gene expression patterns, cytogenetic loci, signaling pathways and functional motifs confirmed documented data and pointed to novel aspects of chondrocyte differentiation. Parallel comparisons of the microdissected tibiae data set to our previously completed micromass culture screen further corroborated the suitability of micromass cultures for modeling gene expression in chondrocyte development. The micromass culture system demonstrated striking similarities to the in vivo microdissected tibiae screen; however, the micromass system was unable to accurately distinguish gene expression differences in the hypertrophic and mineralized zones of the growth plate. These studies will allow us to better understand zone-specific gene expression patterns in the growth plate. Ultimately, this work will help define both the genomic context in which genes are expressed in the long bones and the extent to which the micromass culture system is able to recapitulate chondrocyte development in endochondral ossification.
Genome-wide analyses of gene expression during mouse endochondral ossification.
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View SamplesAffymetrix Mouse Gene 1.0 ST Array profiles were generated from acticular cartilage derived from CBA and Str/ort mice at three ages (8W, 18W, 40W), corresponding to stages prior to, at and late after natural osteoarthritis (OA) onset in OA-prone Str/ort mice.
Time-series transcriptional profiling yields new perspectives on susceptibility to murine osteoarthritis.
Age, Specimen part
View SamplesThe goal of this study is to uncover the changes in the transcriptome of sensory neurons of the liver kinase B1 (LKB1) knockout
Regulation of axonal morphogenesis by the mitochondrial protein Efhd1.
Specimen part
View SamplesGenome-wide mRNA expression in brains of wild-type and eIF2B-R132H/R132H mutant mice (Geva et al., BRAIN 133 (8), 2010) profiled at postnatal (P) days 1, 18 and 21 to reflect the early proliferative stage prior to white matter establishment (P1) and the peak of oligodendrocye differentiation and myelin synthesis (P18 and P21).
A point mutation in translation initiation factor eIF2B leads to function--and time-specific changes in brain gene expression.
Specimen part
View SamplesA series of transfections was performed in Drosophila S2 cells to explore: 1) the types of target sites that Drosophila microRNAs recognize, 2) the relative functional efficacy of these sites in mediating repression, and 3) the determinants that allow some sites to have greater potency than others. 3p-seq was also performed to help reannotate and quantify the landscape of 3'' UTRs in Drosophila S2 cells. Overall design: Nine mRNA profiles were generated, with Drosophila S2 cells transfected with one of 6 microRNAs (miR-1, miR-4, miR-92a, miR-124, miR-263a, and miR-997). These samples were compared to 3 biological replicates of a mock transfection condition. 3p-seq data for S2 cells was also generated to help reannotate and quantify 3'' UTR isoforms.
Predicting microRNA targeting efficacy in Drosophila.
Specimen part, Subject
View SamplesImmunodeficient mouse models have been valuable for studies of human hematopoiesis, but high-fidelity recapitulation of erythropoiesis in most xenograft recipients remains elusive. Recently developed immunodeficient and Kit mutant mice, however, have provided a suitable background to achieve higher-level human erythropoiesis after long-term hematopoietic engraftment. While there has been some characterization of human erythropoiesis in these models, a comprehensive analysis of various developmental stages has not yet been reported. Here, we have utilized cell surface phenotypes, morphologic analyses, and molecular studies to fully characterize human erythropoiesis from multiple developmental stages in immunodeficient and Kit mutant mouse models following long-term hematopoietic stem and progenitor cell engraftment. We show that human erythropoiesis in such models demonstrates complete maturation and enucleation, as well as developmentally appropriate globin gene expression. These results provide a framework for future studies to utilize this model system for interrogating disorders affecting human erythropoiesis and for developing improved therapeutic approaches. Overall design: (mRNA-seq) RNA-seq of human CD235a+ cells isolated 14-16 weeks post-implantation from mouse bone marrow were performed for three biological replicates each of mice xenograted with adult bone marrow-derived human CD34+ cells and cord blood-derived CD34+ cells.
Developmentally-faithful and effective human erythropoiesis in immunodeficient and Kit mutant mice.
Specimen part, Subject
View SamplesThe post-transcriptional fate of messenger RNAs (mRNAs) is largely dictated by their 3'' untranslated regions (3''UTRs), which are defined by cleavage and polyadenylation (CPA) of pre-mRNAs. We used poly(A)-position profiling by sequencing (3P-Seq) to map poly(A) sites at eight developmental stages and tissues in the zebrafish. Analysis of over 60 million 3P-Seq reads substantially increased and improved existing 3''UTR annotations, resulting in confidently identified 3''UTRs for more than 78.79% of the annotated protein-coding genes in zebrafish. Most zebrafish genes undergo alternative CPA with more than a thousand genes using different dominant 3''UTRs at different stages. 3''UTRs tend to be shortest in the ovaries and longest in the brain. Isoforms with some of the shortest 3''UTRs are highly expressed in the ovary yet absent in the maternally contributed RNAs of the embryo, perhaps because their 3''UTRs are too short to accommodate a uridine-rich motif required for stability of the maternal mRNA. At two hours post-fertilization, thousands of unique poly(A) sites appear at locations lacking a typical polyadenylation signal, which suggests a wave of widespread cytoplasmic polyadenylation of mRNA degradation intermediates. Our insights into the identities, formation, and evolution of zebrafish 3''UTRs provide a resource for studying gene regulation during vertebrate development. Overall design: 3P-Seq was used to map the 3'' ends of protein-coding genes in the zebrafish genome
Extensive alternative polyadenylation during zebrafish development.
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