Project description:The Nuclear Factor I (NFI) family of DNA binding proteins (also called CAAT box transcription factors or CTF) is involved both in replication of adenoviral DNA and in regulation of gene expression. Using chromatin immuno-precipitation and high throughput sequencing (method termed ChIP-Seq) we performed genome-wide mapping of Nuclear Factor I DNA binding sites in mouse embryonic fibroblasts. We found that in vivo and in vitro NFI binding specificities are identical, since previously established position weight matrix was found to accurately predict binding sites for NFI group of proteins. Positional correlation between + and - strand ChIP-Seq tags revealed that NFI is a nucleosome-binding protein, unlike some other transcription factors. We further found that NFI binding correlates with the specific histone modification H3K4me3, the marker of transcribed promoters. Combining ChIP-Seq with the microarray expression data, we found that NFI associates with promoters with higher transcription level. We estimate that transcribed promoters may be more accessible to transcription factor binding than their nearby regions since NFI preferentially bind their DNA target sites located around transcription start sites. Knocking-out one of the NFI proteins (NFI-C), reduced the occupancy of predicted sites, however at the same time indicating that cells could compensate the missing protein with the other members of the family. Keywords: ChIP-Seq (chromatin immunoprecipitation and high throughput sequencing) ChIP-Seq (chromatin immunoprecipitation and high throughput sequencing) using antibody against NFI family of transcription factors in two cell types (wild type and NFI-C knock-out mouse embryonic fibroblasts MEFs)
Project description:To understand the mechanism underlying the transcriptional regulation by Sox2, we analyzed genome-wide binding sites of Sox2, Tfap2c, and Cdx2 in trophoblast stem (TS) cells by chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq). ZHBTc4- and embryo-derived TS cell lines were maintained in the presence of FGF4 and mouse embryonic fibroblasts (MEFs).
Project description:To understand the mechanism underlying the versatility in transcriptional regulation by Sox2, we compared genome-wide binding sites of Sox2 in embryonic stem (ES) cells and trophoblast stem (TS) cells by chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq). A tetracycline-inducible Oct3/4 knockout ES cell line ZHBTc4 was treated with Tet for 4 days in the presence of FGF4 and mouse embryonic fibroblasts (MEFs).
Project description:ChIP coupled with NGS identifies genome-wide binding sites of a ES cells specific Sin3a/Hdac complex. The aim of these experiments is to study the role of Fam60a in the Sin3a/Hdac complex. ChIP-Seq experiments reveal that Fam60a is required to maintain high levels of Sin3a binding on target genes in mESCs. Depletion of Fam60a causes a drop of Sin3a binding to target sites. Underlining the function of Fam60a in mES cells, ChIP-seq analysis of Sin3a and Fam60a in mouse fibroblasts reveal strikingly low global binding level of Sin3a to its target genes while Fam60a is absent at these sites.
Project description:We performed ChIP-seq on a complex developing tissue (the spinal cord) at mouse embryonic day 12.5. Genome-wide binding sites of Mash1 were generated. ChIP-seq of Mash1/Ascl1 in a heterogeneous tissue.
