Project description:To analyze global gene-expression changes caused by IRF2 loss in ISCs, ISCs were prepared from Irf2fl/fl: Lgr5-EGFP-Ires-CreERT2(Irf2fl/fl: Lgr5ki) mice or Irf2fl/fl: Ah-Cre: Lgr5-EGFP-Ires-CreERT2 (Irf2AhcKO: Lgr5ki) mice 1 month after 5 consecutive days of βNF treatment. The Gene Ontology (GO) analysis indicated that the GO terms overrepresented among the genes upregulated in ISCs of βNF treated Irf2AhcKO: Lgr5ki mice compared with those of Irf2fl/fl: Lgr5ki mice included “immune system process”, “immune response”, and “cellular response to Interferon-beta”, all from the GOTERM_Biological Processes (BP) category. Because these GO_term inculded many IFN-inducibl genes, IFN signaling augumented in Irf2 deleted ISCs.
Project description:To check gene expression signatures for LGR5hi and LGR5lo ISCs in response to gamma-irradiation, we performed whole genome microarray on LGR5hi and LGR5lo ISCs purified from irradiated mice or non-irradiated mice Radiation induced gene expression in LGR5hi and LGR5lo ISCs of LGR5-GFPki mice was measured at 3 hours after exposure to 12Gy γ-irradiation or non-irradiation.
Project description:To examine whether IRF2, a negative regulator of IFN signaling, constitutively represses IFN signaling by binding IFN-inducible gene loci in ISCs, we performed a genome-wide chromatin immunoprecipitation/DNA sequencing (ChIP-Seq) analysis of IRF2 in Lgr5 ISCs. We identified 381 binding peaks in these cells, including well-known IFN-inducible genes. Motif analysis showed significant enrichment of consensus-binding motifs for IRF transcription factors within these peaks. Within IRF2-occupied genes in ISCs, we identified 204 of experimentally validated IFN-inducible genes from Interferome database, and 10.8% of them were overlapped with the genes upregulated by type I IFN stimulation in ISCs. These findings indicated for the first time that IRF2 constitutively bound and repressed the sterile IFN signaling at the level of ISCs.
Project description:To explore the influence of IFNR-mediated singnaling in ISCs, we performed microarray analysis of the ISCs from control Lgr5ki mice versus Ifnar1-/-Lgr5ki mice after treating for 1 weeks with low doses of poly(I:C), which potentially induce type I and -III IFNs. The Gene Ontology (GO) analysis indicated that the GO terms overrepresented among the genes upregulated in ISCs from poly(I:C)-treated WT mice compared with those from poly(I:C)-treated Ifnar1−/− mice included “defense response to virus”, “immune system process”, “response to virus”, and “innate immune response”, all from the GOTERM_Biological Processes (BP) category. We compared these results against the Interferome database, which contains genes regulated by type I, II, or III IFN, compiled by analyzing the expression data of IFN-treated cells (http://www.interferome.org), and found that most of the genes found for the GO term “defense response to virus” were IFN-inducible genes, indicating that IFNs act directly on ISCs. The GO analysis also showed an enrichment of defense response to bacteria. In the GOTERM_Cellular Component (CC) category, “extracellular space” and “extracellular region” ranked at the top, indicating a dramatic change in the expression pattern of secretory molecules. Notably, these GO terms included genes encoding antibacterial proteins and endocrine hormones such as angiogenin, defensin, lysozyme, chromogranin, and glucagon, all of which are produced by secretory IECs. Thus, we hypothesized that excess IFN signals force ISCs to lose stemness and differentiate into secretory progenitors.
Project description:Purpose: To analysis impacts of groucho and notch on transcription in Drosophila ISCs by mRNA-seq. Methods: mRNA-seq of RNA extracted from FACS sorted Drosophila ISCs. Results: Groucho loss in ISCs leads to disruption of lateral inhibition,and knocking down notch in ISCs down-regulates expression of e(spl) factors.