Project description:Chromatin immunoprecipitation using Foxc1 antibodies in second branchial arch (IIBA) cells from E11.5 mouse embryos detected by SOLiD sequencing
Project description:We previously reported that an aggressive subpopulation of highly tumorigenic, drug-resistant bladder cancer cells can arise from the bulk tumor cells without mutational events and that this phenotypic plasticity is driven at least in part by epigenetic mechanisms. In the current work, we analyzed the chromatin accessibility of enhancers to identify transcription factors that contribute to this transition to a drug-resistant state. Comparing the drug resistant side population (SP) cells and the less drug resistant non-side population (NSP) cells in bladder cancer cells, we identified differential accessible enhancers and differentially expressed genes. Transcription factor motif analysis showed that FOX family motif was enriched in accessible enhancers near SP-overexpressed genes. Among FOX family transcription factors, FOXC1 was the only overexpressed transcription factor, and it was also the most significantly overexpressed gene in SP cells. FOXC1 ChIP-seq confirmed that FOXC1 binding sites are more accessible in SP cells and showed a significantly more FOXC1 binding near the overexpressed genes in SP cells. When the FOXC1 is knocked out, bladder cancer cells exhibit decreased cisplatin resistance and less percentage of SP cells. This change in cisplatin resistance is partially related to the FOXC1-driven expression of ABCB1 gene. In summary, our observations suggest that differential expression and enhancer binding of FOXC1 promotes the previously observed, mutation-independent shift towards cisplatin resistance in bladder cancer.
Project description:To investigate FOXC1 chromatin binding, and the effect of FOXC1 CRISPR knockout in triple negative breast cancer cell lines MDA-MB-231, MDA-MB-468, Hs578t, and Bt-549.
Project description:Estrogen Receptor alpha (ERα) is a key driver of most breast cancers, and it is the target of endocrine therapies used in the clinic to treat women with ERα positive (ER+) breast cancer. The two methods ChIP-seq (chromatin immunoprecipitation coupled with deep sequencing) and RIME (Rapid Immunoprecipitation of Endogenous Proteins) have greatly improved our understanding of ERα function during breast cancer progression and in response to anti-estrogens. A critical component of both ChIP-seq and RIME protocols is the antibody that is used to pull down the bait protein. To date, most of the ChIP-seq and RIME experiments for the study of ERα have been performed using the sc-543 antibody from Santa Cruz Biotechnology. However, this antibody has been discontinued, thereby severely impacting the study of ERα in normal physiology as well as diseases such as breast cancer and ovarian cancer. Here, we compare the sc-543 antibody with other commercially available antibodies, and we show that 06-935 (EMD Millipore) and ab3575 (Abcam) antibodies can successfully replace the sc-543 antibody for ChIP-seq and RIME experiments.
Project description:We previously reported that an aggressive subpopulation of highly tumorigenic, drug-resistant bladder cancer cells can arise from the bulk tumor cells without mutational events and that this phenotypic plasticity is driven at least in part by epigenetic mechanisms. In the current work, we analyzed the chromatin accessibility of enhancers to identify transcription factors that contribute to this transition to a drug-resistant state. Comparing the drug resistant side population (SP) cells and the less drug resistant non-side population (NSP) cells in bladder cancer cells, we identified differential accessible enhancers and differentially expressed genes. Transcription factor motif analysis showed that FOX family motif was enriched in accessible enhancers near SP-overexpressed genes. Among FOX family transcription factors, FOXC1 was the only overexpressed transcription factor, and it was also the most significantly overexpressed gene in SP cells. FOXC1 ChIP-seq confirmed that FOXC1 binding sites are more accessible in SP cells and showed a significantly more FOXC1 binding near the overexpressed genes in SP cells. When the FOXC1 is knocked out, bladder cancer cells exhibit decreased cisplatin resistance and less percentage of SP cells. This change in cisplatin resistance is partially related to the FOXC1-driven expression of ABCB1 gene. In summary, our observations suggest that differential expression and enhancer binding of FOXC1 promotes the previously observed, mutation-independent shift towards cisplatin resistance in bladder cancer.
Project description:We previously reported that an aggressive subpopulation of highly tumorigenic, drug-resistant bladder cancer cells can arise from the bulk tumor cells without mutational events and that this phenotypic plasticity is driven at least in part by epigenetic mechanisms. In the current work, we analyzed the chromatin accessibility of enhancers to identify transcription factors that contribute to this transition to a drug-resistant state. Comparing the drug resistant side population (SP) cells and the less drug resistant non-side population (NSP) cells in bladder cancer cells, we identified differential accessible enhancers and differentially expressed genes. Transcription factor motif analysis showed that FOX family motif was enriched in accessible enhancers near SP-overexpressed genes. Among FOX family transcription factors, FOXC1 was the only overexpressed transcription factor, and it was also the most significantly overexpressed gene in SP cells. FOXC1 ChIP-seq confirmed that FOXC1 binding sites are more accessible in SP cells and showed a significantly more FOXC1 binding near the overexpressed genes in SP cells. When the FOXC1 is knocked out, bladder cancer cells exhibit decreased cisplatin resistance and less percentage of SP cells. This change in cisplatin resistance is partially related to the FOXC1-driven expression of ABCB1 gene. In summary, our observations suggest that differential expression and enhancer binding of FOXC1 promotes the previously observed, mutation-independent shift towards cisplatin resistance in bladder cancer.
Project description:To investigate FOXC1 chromatin binding, and the effect of FOXC1 CRISPR knockout in triple negative breast cancer cell lines MDA-MB-231, MDA-MB-468, Hs578t, and Bt-549.
Project description:We report downstream gene expression changes in stem cells of the adult mouse hair follicle upon conditional ablating of the transcription factor Forkhead Box C1 transcription factor (FOXC1). Hair follicles undergo cycles of rest (telogen; Tel) and regeneration (anagen; Ana). As such, we performed our analysis on these two different stages of hair follicles. mRNA-sequencing of WT vs. Foxc1-conditional or inducible KO (Foxc1-cKO or iKO) hair follicle stem cells (HFSCs) purified from mouse dorsal back skin by flow-activated cell sorting (FACS).