Bladder cancer is mostly present in the form of urothelium carcinoma, causing over 150,000 deaths each year. Its histopathological classification as muscle invasive (MIBC) and non-muscle invasive (NMIBC) is the most prominent aspect, affecting the prognosis and progression of this disease. In this study, we defined the active regulatory landscape of MIBC and NMIBC cell lines using H3K27ac ChIP-seq and used an integrative approach to combine our findings with existing data. Our analysis revealed FRA1 and FLI1 as two critical transcription factors differentially regulating MIBC regulatory landscape. We show that FRA1 and FLI1 regulate the genes involved in epithelial cell migration and cell junction organization. Knock-down of FRA1 and FLI1 in MIBC revealed the downregulation of several EMT-related genes such as MAP4K4 and FLOT1. Further, ChIP-SICAP performed for FRA1 and FLI1 enabled us to infer chromatin binding partners of these transcription factors and link this information with their target genes. Finally, we show that knock-down of FRA1 and FLI1 result in significant reduction of invasion capacity of MIBC cells towards muscle microenvironment using IC-CHIP assays. Our results collectively highlight the role of these transcription factors in selection and design of targeted options for treatment of MIBC.
Epigenetic deregulation is a critical theme which needs further investigation in bladder cancer research. One of the most highly mutated genes in bladder cancer is KDM6A, which functions as an H3K27 demethylase and is one of the MLL3/4 complexes. To decipher the role of KDM6A in normal versus tumor settings, we identified the genomic landscape of KDM6A in normal, immortalized, and cancerous bladder cells. Our results showed differential KDM6A occupancy in the genes involved in cell differentiation, chromatin organization, and Notch signaling depending on the cell type and the mutation status of KDM6A. Transcription factor motif analysis revealed HES1 to be enriched at KDM6A peaks identified in the T24 bladder cancer cell line; moreover, it has a truncating mutation in KDM6A and lacks a demethylase domain. Our co-immunoprecipitation experiments revealed TLE co-repressors and HES1 as potential truncated and wild-type KDM6A interactors. With the aid of structural modeling, we explored how truncated KDM6A could interact with TLE and HES1, as well as RUNX and HHEX transcription factors. These structures provide a solid means of studying the functions of KDM6A independently of its demethylase activity. Collectively, our work provides important contributions to the understanding of KDM6A malfunction in bladder cancer.
Epigenetic deregulation is a critical theme which needs further investigation for bladder cancer research. One of the highly mutated genes in bladder cancer is KDM6A, functioning as a H3K27 demethylase and is part of the MLL3/4 complexes. To decipher the role of KDM6A in normal versus tumor setting, we identified the genomic landscape of KDM6A in normal, immortalized and cancer bladder cells. Our results showed differential KDM6A occupancy at the genes involved in cell differentiation, chromatin organization and Notch signaling depending on the cell type and the mutation status of KDM6A. Transcription factor motif analysis revealed HES1 to be enriched at KDM6A peaks identified for T24 bladder cancer cell line, which has a truncating mutation in KDM6A, lacking demethylase domain. Our co-immunoprecipitation experiments reveal TLE co-repressors and HES1 as potential truncated-KDM6A interactors. With the aid of structural modeling, we explored how the truncated KDM6A could interact with TLE, HES1, as well RUNX, HHEX transcription factors. These structures provide a solid mean to study the functions of KDM6A independent of its demethylase activity. Collectively, our work provides important contributions to the understanding of KDM6A malfunction in bladder cancer.
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