Project description:We examine the role of Klf6 in oligodendrocyte progenitor cells and determine that Klf6 acts as a gp130-sensitive transactivator of the nuclear import factor importin-α5 (Impα5), a key controller of nuclear trafficking in oligodendrocytes. Examination of expression profiles of 2 different cell stages exposed to siRNA vs. control
Project description:We examine the role of Klf6 in oligodendrocyte progenitor cells and determine that Klf6 acts in part through direct regulation of gp130 signaling and nuclear import via importin-α5 (Impα5), a key controller of nuclear trafficking. Examination of Klf6 DNA binding in two different stages of differentiation
Project description:We performed genome-wide profiling of Tcf7l2 occupancy during oligodendrocyte differentiation and identified the key enzymes involved in cholesterol metabolism and essential for CNS myelination. Examination of Tcf7l2 chIP-seq in oligodendrocyte progenitor cell and 2 differentiation oligodendrocytes.
Project description:Establishment and maintenance of CNS glial cell identity ensures proper brain development and function, yet the epigenetic mechanisms underlying glial fate control remain poorly understood. Here we show that the histone deacetylase Hdac3 controls oligodendrocyte-specification gene Olig2 expression, and functions as a molecular switch for oligodendrocyte and astrocyte lineage determination. Our data suggest that Hdac3 cooperates with p300 to prime and maintain oligodendrogenic programs while inhibiting Stat3-mediated astrogliogenesis, and thereby regulate phenotypic commitment at the point of oligodendrocyte-astrocytic fate decision. Examination of Hdac3 and p300 genomewide occupancy in differentiating oligodendrocytes
Project description:Gene-expression profiles of liver and hepatocellular carcinoma induced by diethylnitrosamine (DEN) in KLF6 +/- and wild type KLF6 mice. Inactivation of the KLF6 tumor suppressor is common in HCC due to hepatitis C virus (HCV), consistent with its anti-proliferative activity in HCC-derived cell lines and in hepatocytes of transgenic mice. We have evaluated the impact of KLF6 depletion on human HCC and experimental hepatocarcinogenesis. In patients with surgically resected HCC, those with significantly reduced tumor expression of KLF6 had a significantly decreased survival. We modeled this event in KLF6 +/- mice, which displayed significantly more tumorigenicity than KLF6 +/+ animals in response to the hepatic carcinogen DEN, associated with recapitulation of gene signatures in both surrounding tissue and tumors that are associated with aggressive human HCCs. In DNA microarrays, mdm2 mRNA expression was increased in tumors from KLF6 +/- compared to KLF6 +/+ mice, which was validated by realtime qPCR and Western blot in both human HCC and DEN-induced murine tumors. Moreover, chromosomal immunoprecipitation and co-transfection assays established the P2 intronic promoter of mdm2 as a bona fide transcriptional target repressed by KLF6. Whereas KLF6 over-expression in HCC cell lines led to reduced MDM2 levels and increased p53 protein and transcriptional activity, reduction in KLF6 by siRNA led to increased MDM2 and reduced p53. Our findings indicate that KLF6 deficiency contributes significantly to the carcinogenic milieu in human and murine HCC, and uncover a novel tumor suppressor activity of KLF6 in HCC, by linking its transcriptional repression of MDM2 to stabilization of p53. Keywords: Liver, Hepatocellular carcinoma, Expression array, Exon array, Affymetrix KLF6 +/- mice were previously generated by homologous recombination in which exon 2 was targeted using an 11-kb targeting construct, and replaced with neomycin/lacZ cassette. After selection with neomycin, the ES clones were injected into C57BL/6 mouse blastocysts and implanted into pseudo pregnant females; two lines of KLF6 +/- mice were generated from the resulting chimeric animals (Blood 107;1357, Oncogene 26;4428). Whereas KLF6 -/- mice are embryonic lethal, KLF6 +/- animals had no demonstrable abnormalities in the absence of any stressor. Male KLF6 +/- mice were bred with wild type C57BL/6 to generate mixed litters of KLF6 +/- and KLF6 +/+ animals. Progeny were genotyped using PCR-amplified tail DNA, using primers as previously described (Oncogene 26;4428). Amplified fragments were separated on a 2.5% agarose gel, revealing bands of ~200 bp (wild type KLF6) and ~100 bp (Neo), as expected. At 2 weeks of age, KLF6 +/+ and KLF6 +/- mice were injected intraperitoneally with either a single dose of diethyl nitrosamine (DEN), 5 µg/g body weight in 100 µl of saline, or vehicle alone. Vehicle and DEN-treated mice were maintained on standard chow, and then sacrificed 3, 6 or 9 months later. At the time of sacrifice the animals were weighed, and blood and liver samples were harvested for analysis and tumor quantification.
Project description:We investigate the role of Brg1 and Olig2 during oligodendrocyte differentiation by combining gene conditional knockout and next generation sequencing technology. We generate genome-wide maps of RNA polymerase II (RPolII), Brg1 (Smarca4), Olig2 and histone modifications in primary rat oligodendrocyte precursor cells (iOLs), differentiating oligodendrocytes and mature oligodendrocytes.We found that Brg1 is intensely regulated by RPolII at the initiation of oligodedrocyte differentiation. The genomic distribution of Brg1 in differentiating oligodendrocytes is pre-directed by Olig2 in iOLs. The dynamic interaction of Brg1 and chromatin is correlate with the distinct stages of gene expression during maturation. Finally, we show that Brg1 and Olig2 localization predict critical genes controling CNS myeliantion. Our study represents the first detailed analysis of genomic landscape during the oligodendrocyte development and provides a framework for further understanding of molecular mechanisms underlying oligodendrocyte lineage progression. Genomic distribution of Brg1, Olig2, RPolII and three different histone modifications in three oligodendrocyte developemtal stages were examined using primary cells by ChIP-sequencing. Spinal cord mRNA profiles of 14-day old control and Brg1c/c;Olig1-Cre mice were generated by RNA-sequencing.
Project description:We sequenced DNA isolated from performing ChIP of full-length KLF6 and an Input sample in an HCC cell line. The goal is to determine KLF6 binding sites in a mouse-derived HCC cell line. Determination of KLF6 binding sites in an HCC cell line using 2 control input libraries and 2 KLF6-ChIP libraries