The cascade of molecular events involved in mammalian sex determination has been shown to involve the SRY gene, but specific downstream events have eluded researchers for decades. The current study identifies one of the first direct downstream targets of the male sex-determining factor SRY as the basic-helix-loop-helix (bHLH) transcription factor TCF21. SRY was found to directly associate with the Tcf21 promoter SRY/SOX9 response element both in vitro and in vivo during male sex determination. TCF21 was found to promote an in vitro sex reversal of embryonic ovarian cells to promote precursor Sertoli cell differentiation. Therefore, SRY acts directly on the Tcf21 promoter to, in part, initiate a cascade of events associated with Sertoli cell differentiation and embryonic testis development.
Basic helix-loop-helix transcription factor TCF21 is a downstream target of the male sex determining gene SRY.
Sex, Specimen part, Treatment
View SamplesCoordinated interactions between ovarian granulosa and theca cells are required for female endocrine function and fertility. To elucidate these interactions the regulation of the granulosa and theca cell transcriptomes during bovine antral follicle development were investigated. Granulosa cells and theca cells were isolated from small (<5 mm), medium (5-10 mm), and large (>10 mm) antral bovine follicles. A microarray analysis of 24,000 bovine genes revealed that granulosa cells and theca cells each had gene sets specific to small, medium and large follicle cells. Transcripts regulated (i.e., minimally changed 1.5-fold) during antral follicle development for the granulosa cells involved 446 genes and for theca cells 248 genes. Only 28 regulated genes were common to both granulosa and theca cells. Regulated genes were functionally categorized with a focus on growth factors and cytokines expressed and regulated by the two cell types. Candidate regulatory growth factor proteins mediating both paracrine and autocrine cell-cell interactions include macrophage inflammatory protein (MIP1 beta), teratocarcinoma-derived growth factor 1 (TDGF1), stromal derived growth factor 1 (SDF1; i.e., CXCL12), growth differentiation factor 8 (GDF8), glia maturation factor gamma (GMFG), osteopontin (SPP1), angiopoietin 4 (ANGPT4), and chemokine ligands (CCL 2, 3, 5, and 8). The current study examined granulosa cell and theca cell regulated genes associated with bovine antral follicle development and identified candidate growth factors potentially involved in the regulation of cell-cell interactions required for ovarian function.
Regulation of granulosa and theca cell transcriptomes during ovarian antral follicle development.
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View SamplesThe goal of this study is to compare gene expression levels in the livers of larval Tg(fabp10:nls-mcherry) exposed to 1 mM inorganic arsenic from 4-120 hpf to the unexposed siblings. Samples were collected from Tg(fabp10:nls-mcherry) zebrafish larvae that were derived from incrossed parents of the same strain. The background of transgenic lines were typically from mixed outcrosses of the transgenics to AB, TAB5, and TAB14 strains when regenerating the lines as the working stocks aged. All samples were collected at approximately 120 hpf - natural spawning at 8:30-9:00AM EST on day zero, samples were collected at 8-10AM EST on day 5. Overall design: 10-20 livers from 5dpf embryos were pooled per sample of either control or 1 mM inorganic arsenic exposed Tg(fabp10:nls-mcherry) zebrafish larvae and RNA was extracted using the Zymo Quick-RNA Micro Kit with on-column DNase treatment. Libraries were prepared according to Illumina Truseq RNA sample prep kit, version 2, followed by Ribo-Zero Gold treatment.
Inorganic arsenic causes fatty liver and interacts with ethanol to cause alcoholic liver disease in zebrafish.
No sample metadata fields
View SamplesUHRF1 (Ubiquitin-like, containing PHD and RING finger domains, 1) recruits DNMT1 to hemimethylated DNA during replication, is essential for maintaining DNA methylation patterns during cell division and is suggested to direct additional repressive epigenetic marks. Uhrf1 mutation in zebrafish results in multiple embryonic defects including failed hepatic outgrowth, but the epigenetic basis of these phenotypes is not known. We find that DNA methylation is the only epigenetic mark that is depleted in uhrf1 mutants and make the surprising finding that despite the reduced organ size in uhrf1 mutants, genes regulating DNA replication and S-phase progression were highly upregulated. Further, there is a striking increase in BrdU incorporation in uhrf1 mutant cells, and they retained BrdU labeling over several days, indicating they are arrested in S-phase. Moreover, some of the label retaining nuclei co-localized with TUNEL positive nuclei, suggesting that arrested cells are responsible for apoptosis. Importantly, dnmt1 mutation phenocopies the S-phase arrest and hepatic outgrowth defects in uhrf1 mutants and Dnmt1 knock-down enhances the uhrf1 hepatic phenotype. Together, these data indicate that DNA hypomethylation is sufficient to generate the uhrf1 mutant phenotype by promoting an S-phase arrest. We thus propose that cell cycle arrest is a mechanism to restrict propagation of epigenetically deranged cells during embryogenesis.
DNA hypomethylation induces a DNA replication-associated cell cycle arrest to block hepatic outgrowth in uhrf1 mutant zebrafish embryos.
No sample metadata fields
View Samplescdipt is an essential gene in the synthesis of phosphatidylinositol (PtdIns) in the zebrafish, Danio rerio. The zebrafish mutant cdipt^hi559Tg (ZL782) carries a retroviral insertion which inactivates cdipt. Homozygous mutants exhibit hepatocellular endoplasmic reticulum (ER) stress and non-alcoholic fatty liver disease (NAFLD) pathologies at 5 days post fertilization (dpf). This study reveals a novel link between PtdIns, ER stress, and steatosis.
Lack of de novo phosphatidylinositol synthesis leads to endoplasmic reticulum stress and hepatic steatosis in cdipt-deficient zebrafish.
Age
View SamplesThe enteric nervous system (ENS) can control most essential gut functions owing to its organization into complete neural circuits consisting of a multitude of different neuronal subtypes.
Transcription and Signaling Regulators in Developing Neuronal Subtypes of Mouse and Human Enteric Nervous System.
Specimen part
View SamplesThe goal of this study is to compare gene expression levels in uhrf1 mutants with global DNA hypomethylation to WT siblings Overall design: 10 whole embryos were pooled per sample of either 5 dpf old uhrf1 mutants or phenotypically WT siblings and RNA was extracted. Libraries were prepared according to Illumina Truseq RNA sample prep kit, version 2, followed by Ribo-Zero Gold treatment
Loss of DNA methylation in zebrafish embryos activates retrotransposons to trigger antiviral signaling.
No sample metadata fields
View SamplesStable knockdown of NET1, a RhoGEF, was achieved in AGS Gastric Cancer cells. This gene is known to be overexpressed in the disease.
A functional and transcriptomic analysis of NET1 bioactivity in gastric cancer.
Cell line
View SamplesBACKGROUND & AIMS- More frequent interaction of bacteria with the colonic epithelium is associated with ulcerative colitis (UC). The identities of all proteins which promote bacterial clearance in colonic epithelial cells are unknown. Previously, we discovered that dCAP-D3 (Chromosome Associated Protein-D3), regulates responses to bacterial infection. We examined whether CAP-D3 promotes bacterial clearance in human colonic epithelium. METHODS- Clearance of Salmonella or adherent-invasive Escherichia coli LF82 was assessed by gentamycin protection assays in HT-29 and Caco-2 cells expressing CAP-D3 shRNA. CAP-D3 levels in colonic epithelial cells from healthy and UC patient tissues were analyzed by immunoblot. RNA-sequencing identified bacterially-induced CAP-D3 target genes. The role of CAP-D3 target genes in bacterial clearance was analyzed by gentamycin protection assays, immunofluorescent staining, and by using pharmacologic inhibitors. RESULTS- CAP-D3 expression was reduced in colonic epithelial cells from UC patients with active disease. Reduction of CAP-D3 expression inhibited autophagy and decreased intracellular bacterial clearance. The components of the heterodimeric SLC7A5/SLC3A2 amino acid transporter were identified as CAP-D3 target genes; their levels increased in infected, CAP-D3 deficient cell lines and in cells from UC patients. In HT-29 cells, this resulted in earlier SLC7A5 recruitment to Salmonella-containing vacuoles, increased mTOR activity, and enhanced bacterial survival. Inhibition of SLC7A5/SLC3A2 or mTOR activity rescued the bacterial clearance defect in CAP-D3 deficient cells. CONCLUSIONS- CAP-D3 attenuates amino acid transporter transcription to promote bacterial autophagy in colon epithelial cells. CAP-D3 protein levels are decreased in patients with active UC, suggesting that CAP-D3 is a potential therapeutic target to restore mucosal homeostasis in UC patients. Overall design: Three RNA samples from 3 independent experiments including timepoints taken at 0, 0.5 and 7 hours post-infection were analyzed on a bioanalyzer for quality; one of the 0.5 hour post-infection samples was excluded at this time due to poor RNA purity. Directional, cDNA libraries made from cellular mRNAs were generated from the other 8 samples and sequenced (paired-end sequencing of 100 bp reads) in the Genomics Core at the University of Chicago on an Illumina HiSeq2000.
Chromosome-associated protein D3 promotes bacterial clearance in human intestinal epithelial cells by repressing expression of amino acid transporters.
No sample metadata fields
View SamplesUHRF1 is an essential regulator of DNA methylation that is highly expressed in many cancers. Using transgenic zebrafish, cultured cells and human tumors, we demonstrate that UHRF1 is an oncogene. RNAseq was used to assess the variation in gene expression between control and experimental samples. Overall design: Total small RNA from 2 batches of Tg(fabp10:has.UHRF1-GFP)High and age matched Tg(fabp10:nls-mCherry) control 5 dpf zebrafish livers was purified for preparation of high-throughput sequencing libraries.
UHRF1 overexpression drives DNA hypomethylation and hepatocellular carcinoma.
No sample metadata fields
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