Purpose: Klf5 plays a critical role in the mouse ocular surface (Kenchegowda et al., 2011. Dev Biol. 356:5-18). Here, we compare wild-type (WT) and Klf5-conditional null (Klf5CN) corneal gene expression at postnatal day-11 (PN11) and PN56 to identify the Klf5-target genes. Methods: Gene expression was compared using Affymetrix microarrays with QPCR validation. Transient transfection assays examined the effect of Klf5 on selected target gene promoter activities. Whole-mount corneal immunofluorescent staining examined neovascularization and CD45+ macrophage influx. Results: Expression of 714 and 753 genes was increased, and 299 and 210 genes decreased in PN11 and PN56 Klf5CN corneas, respectively, with 366 concordant increases, 72 concordant decreases and 3 discordant changes. Canonical pathway analysis identified 35 and 34 significantly (p<0.001) enriched pathways at PN11 and PN56, respectively, with 24 common pathways. PN56 Klf5CN corneas shared 327 increases and 91 decreases with the previously described Klf4CN corneas (Swamynathan et al., 2008. IOVS 49:3360-70). Angiogenesis and immune response-related genes were affected consistent with lymphangiogenesis and macrophage influx in Klf5CN corneas, respectively. Expression of 1574 genes was increased and 1915 decreased, in the WT PN56 compared with PN11 corneas. Expression of many collagens, matrix metalloproteinases and other extracellular matrix associated genes decreased in WT corneas between PN11 and PN56, while that of solute carrier family members increased. Conclusions: Differences in PN11 and PN56 corneal Klf5-target genes reveal dynamic changes in Klf5 functions during corneal maturation. Klf4- and Klf5-target genes do not overlap, consistent with their non-redundant roles in the mouse cornea.
Critical role of Klf5 in regulating gene expression during post-eyelid opening maturation of mouse corneas.
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View SamplesA number of breast or colon specific genes predictive of the relapse status were used in comparing the outcome from matched fresh frozen and stored in RNAlater preservative.
Prognostic gene expression signatures can be measured in tissues collected in RNAlater preservative.
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View SamplesGenes and pathways in which inactivation dampens tissue inflammation present new opportunities for understanding the pathogenesis of common human inflammatory diseases, including inflammatory bowel disease, rheumatoid arthritis and multiple sclerosis. We identified a mutation in the gene encoding the deubiquitination enzyme USP15 (Usp15L749R) that protected mice against both experimental cerebral malaria (ECM) induced by Plasmodium berghei and experimental autoimmune encephalomyelitis (EAE). Combining immunophenotyping and RNA sequencing in brain (ECM) and spinal cord (EAE) revealed that Usp15L749R-associated resistance to neuroinflammation was linked to dampened type I interferon responses in situ. In hematopoietic cells and in resident brain cells, USP15 was coexpressed with, and functionally acted together with the E3 ubiquitin ligase TRIM25 to positively regulate type I interferon responses and to promote pathogenesis during neuroinflammation. The USP15-TRIM25 dyad might be a potential target for intervention in acute or chronic states of neuroinflammation. Overall design: Sequencing of RNA extracted from target tissue in two experimental neuroinflammation models in wild-type (B6), USP15(L749R) and Trim25 KO mutant mice: (1) brains at day 3 and 5 following Plasmodium berghei ANKA (PbA) infection for the cerebral malaria model (ECM); and (2) spinal cords at day 7 following induction of experimental autoimmune encephalomyelitis (EAE) for B6 and Usp15 mutant mice only.
USP15 regulates type I interferon response and is required for pathogenesis of neuroinflammation.
Sex, Specimen part, Treatment, Subject
View SamplesAsthma is caused by a combination of poorly understood genetic and environmental factors. We found multiple markers on chromosome 17q21 to be strongly and reproducibly associated with childhood onset asthma in family and case-referent panels with a combined P < 10-12. In independent replication studies the 17q21 locus showed strong association with diagnosis of childhood asthma in 2,320 subjects from a cohort of German children (P = 0.0003) and in 3,301 subjects from the British 1958 Birth Cohort (P = 0.0005). We systematically evaluated the relationships between markers of the 17q21 locus and transcript levels of genes in EBV-transformed lymphoblastoid cell lines from children in the asthma family panel used in our association study. The SNPs associated with childhood asthma were consistently and strongly associated (P <10-22) in cis with transcript levels of ORMDL3, a member of a gene family that encode transmembrane proteins anchored in the endoplasmic reticulum. The results indicate that genetic variants regulating ORMDL3 expression are determinants of susceptibility to childhood asthma.
Genetic variants regulating ORMDL3 expression contribute to the risk of childhood asthma.
Sex
View SamplesIn vitro oocyte maturation (IVM) holds great promise as a tool for enhancing clinical treatment of infertility, enhancing availability of non human primates for development of disease models, and facilitating endangered species preservation. However, IVM outcomes have remained significantly below success rates obtained using in vivo matured (VVM) oocytes from humans and non human primates. A cDNA array based analysis is presented, comparing the transcriptomes of VVM oocytes with IVM oocytes. We observe a small set of just 59 mRNAs that are differentially expressed between the two cell types. These mRNAs are related to cellular homeostasis, cell-cell interactions including growth factor and hormone stimulation and cell adhesion, and other functions such as mRNA stability and translation. Additionally, we observe in IVM oocytes overexpression of PLAGL1 and MEST, two maternally imprinted genes, indicating a possible interruption or loss of correct epigenetic programming. These results indicate that, under certain IVM conditions, oocytes that are molecularly highly similar to VVM oocytes can be obtained, however the interruption of normal oocyte-somatic cell interactions during the final hours of oocyte maturation may preclude the establishment of full developmental competence.
Effects of in vitro maturation on gene expression in rhesus monkey oocytes.
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View SamplesGene expression has been proposed as an intermediate phenotype that can increase power in complex trait gene-mapping studies. Psoriasis, an immune-mediated, inflammatory and hyperproliferative disease of the skin and joints, provides an ideal model system to evaluate this paradigm, as conclusive evidence demonstrates that psoriasis has a genetic basis and the disease tissue is readily accessible.
Genome-wide scan reveals association of psoriasis with IL-23 and NF-kappaB pathways.
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View SamplesThe oocytes of many species, both invertebrate and vertebrate, contain a large collection of localized determinants in the form of proteins and translationally inactive maternal mRNAs. However, it is unknown whether mouse oocytes contain localized MmRNA determinants and what mechanisms might be responsible for their control. We collected intact MII oocytes, enucleated MII oocyte cytoplasts (with the spindle removed), and spindle-chromosome complexes which had been microsurgically removed. RNA was extracted, amplified, labeled, and applied to microarrays to determine if any MmRNA determinants were localized to the SCC.
Association of maternal mRNA and phosphorylated EIF4EBP1 variants with the spindle in mouse oocytes: localized translational control supporting female meiosis in mammals.
Sex, Specimen part, Disease
View SamplesTranscriptional activation in mammalian embryos occurs in a stepwise manner. In mice, it begins at the late one-cell stage, followed by a minor wave of activation at the early two-cell stage, and then the major genome activation (MGA) at the late two-cell stage. Cellular homeostasis, metabolism, cell cycle, and developmental events are orchestrated before MGA by time-dependent changes in the array of maternal transcripts being translated (i.e., the translatome). Despite the importance of maternal mRNA and its correct recruitment for development, neither the array of recruited mRNA nor the regulatory mechanisms operating have been well cheracterized. We present the first comprehensive analysis of changes in the maternal component of the zygotic translatome during the transition from oocyte to late one-cell stage embryo, revealing global transitions in the functional classes of translated maternal mRNAs, and apparent changes in the underlying cis-regulatory mechanisms.
Analysis of polysomal mRNA populations of mouse oocytes and zygotes: dynamic changes in maternal mRNA utilization and function.
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View SamplesCumulus oophorus cells play an essential role in oocyte development. CBX4 is a member of the Polycomb complex, which plays a role in regulating cellular differentiation.
Contribution of CBX4 to cumulus oophorus cell phenotype in mice and attendant effects in cumulus cell cloned embryos.
Sex, Specimen part
View SamplesWhile the reprogramming factors OCT4, SOX2, KLF4, and MYC (OSKM) can reactivate the pluripotency network in terminally differentiated cells, they also regulate expression of non-pluripotency genes in other contexts, such as the mouse primitive endoderm. The primitive endoderm is an extraembryonic lineage established alongside the pluripotent epiblast in the blastocyst, and is the progenitor pool for extraembryonic endoderm stem (XEN) cells. Several studies have shown that endodermal genes are upregulated in fibroblasts undergoing reprogramming, although whether endodermal genes promote or inhibit acquisition of pluripotency is unclear. We show that, in fibroblasts undergoing conventional reprogramming, OSKM-induced expression of endodermal genes leads to formation of induced XEN (iXEN) cells, which possess key properties of blastocyst-derived XEN cells, including morphology, transcription profile, self-renewal, and multipotency. Our data show that iXEN cells arise in parallel to iPS cells, indicating that OSKM are sufficient to drive cells to two distinct fates during reprogramming. Overall design: Sequence-based mRNA transcriptional profiling of three different cell lines (MEF, XEN, iXEN) with multiple biological replicates, under two different growth medium conditions (ESC medium, XEN medium) for XEN and iXEN cells.
OSKM Induce Extraembryonic Endoderm Stem Cells in Parallel to Induced Pluripotent Stem Cells.
Specimen part, Treatment, Subject
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