Project description:Androgen receptor (AR) signaling is a key driver of prostate cancer (PCa) growth and progression. Understanding the factors influencing AR-mediated transcription provides new opportunities for therapeutic intervention. We have previously discovered that a genetic variant in one of the DNA repair genes, PARP2, is associated with aggressive PCa. Here, we show that a high expression level of PARP2 in PCa tumors is associated with high Gleason scores and biochemical recurrence using The Cancer Genome Atlas (TCGA) dataset. Functional studies reveal that PARP2 enhances AR-mediated gene expression through interacting with the pioneer factor FOXA1 and facilitating AR recruitment to the enhancer regions genome-wide in PCa cells. Selective targeting of PARP2, but not PARP1, by genetic or pharmacological means blocks interaction between PARP2 and FOXA1, which in turn attenuates AR-mediated transcription and inhibits AR-positive PCa growth. SIGNIFICANCE: Current anti-androgens act through blocking ligand binding or inhibiting androgen synthesis. Selective targeting of PARP2 may provide a novel therapeutic approach for AR inhibition by disruption of FOXA1 function, which may be beneficial to patients, irrespective of their DNA repair defect status and, more importantly, when direct AR-targeted therapies fail. Overall design: Genome-wide occupation of AR, FOXA1, and H3K27AC, by ChIP-seq, in LNCaP cells treated with UPF-1069 or vehicle control, with biological duplicates.
Project description:We profiled the genome-wide occupancy of three tissue-specific transcription factors, HNF4A, CEBPA and FOXA1, as well as the genome-wide occurrence of the histone mark, H3K4me3 in the livers of two inbred parental mouse strains (C57BL/6J and CAST/EiJ) and their F1 crosses. We also included H3K27ac data generated from F1 hybrids as well as the profiling of HNF4A, CEBPA and FOXA1 in both CEBPA and HNF4a heterozygous knock-outs.
Project description:Our data suggested that FKHD-MSs impair the chromatin binding of FOXA1 to AR-dependent enhancers and thus suppress AR transcriptional activity, and promote PCa progression through increasing FOXA1 binding to a subset of AR-independent enhancers that regulates transcription of genes mediating EMT and metastasis Overall design: We generated LNCaP stable cells with tetracycline-inducible expression of V5-tagged FKHD-MSs or WT FOXA1. H3K27ac ChIP-seq experiments were performed on these cell lines
Project description:Prostate cancer is one of the most common types of cancer in men and the leading cause of death in developed countries. With the aid of molecular and genetic profiling of cancers, cancer molecular subtypes are paving the way for tailored cancer therapy. FOXA1 has been identified as one of the seven molecular subtypes of prostate cancer. FOXA1 is involved in a variety of metabolic process such as glucose homeostasis and deregulation of its expression is crucial in prostate cancer progression. In this study, we investigated the effects of FOXA1 gene knock-out on the expression levels of various cancer cell metabolism and cell cycle-related protein expressions. FOXA1 gene was knocked-out by using CRISPR/Cas9 technique. While FOXA1 gene knock-out significantly altered Casp-9, Bax, CCND1, CDK4, and fibronectin protein expressions (P?<?0.05, fold change: ?40, 4.5, 2.5, 4.5, and 4, respectively), it did not affect the protein expression levels of Casp-3, Bcl-2, survivin, ?-catenin, c-Myc, and GSK-3B. Knocking-out FOXA1 gene in androgen-dependent LNCaP prostate cancer cells inhibited CCND1 protein expression. Our pre-clinical results demonstrate the importance of FOXA1 as a drug target in the treatment of prostate cancer. Impact statement Knock-out studies offer a unique way of studying the function of genes especially for developmentally lethal genes. FOXA1 has prominent roles both in breast and prostate cancer pathogenesis due to its role in ER receptor signaling pathway. FOXA1 has also been identified as one of the seven molecular subtypes of primary prostate cancer. In the present study, we used an efficient gene knock-out method, CRISPR/Cas9, in order to investigate FOXA1 function on LNCaP prostate cancer cells in vitro. FOXA1 knock-out altered cell-cycle regulator CCND1 protein expression levels. Therefore, our results suggest that FOXA1 might be a plausible drug target for prostate cancer treatment.
Project description:AR is tightly regulated by many transcriptional cofactors, including key pioneer factor such as FOXA1 and GATA2. While FOXA1 was recently shown to be able to redistribute AR across the genome, how GATA2 regulates AR cistrome has not been carefully investigated. Here, we report that, unlike FOXA1, GATA2 is unable to reprogram AR, but instead it enhances AR program by inducing AR expression and augmenting AR co-occupancy, thereby acting as a pure AR coactivator, rather than a pioneer factor. On the other hand, AR co-occupancy enhances both GATA2 and FOXA1 binding on the chromatin, forming a positive feedback loop. Importantly, we found that FOXA1 is also capable of reprogramming GATA2 by recruiting GATA2 from GATA motif to FKHD-containing regions, being analogous to its pioneering effect on AR signaling. By contrast, GATA2 simply acts as a co-activator of FOXA1 with a lack of pioneering ability. ChIP_Seq examination of AR, FoxA1 and GATA2 binding sites in LNCaP and DU145 cells
Project description:A transcription factor functions differentially and/or identically in multiple cell types. However, the mechanism for cell-specific regulation of a transcription factor remains to be elucidated. We address how a single transcription factor, forkhead box protein A1 (FOXA1), forms cell-specific genomic signatures and differentially regulates gene expression in four human cancer cell lines (HepG2, LNCaP, MCF7, and T47D). FOXA1 is a pioneer transcription factor in organogenesis and cancer progression. Genomewide mapping of FOXA1 by chromatin immunoprecipitation sequencing annotates that target genes associated with FOXA1 binding are mostly common to these cancer cells. However, most of the functional FOXA1 target genes are specific to each cancer cell type. Further investigations using CRISPR-Cas9 genome editing technology indicate that cell-specific FOXA1 regulation is attributable to unique FOXA1 binding, genetic variations, and/or potential epigenetic regulation. Thus, FOXA1 controls the specificity of cancer cell types. We raise a "flower-blooming" hypothesis for cell-specific transcriptional regulation based on these observations.
Project description:High androgen receptor (AR) level in primary tumour predicts increased prostate cancer-specific mortality. However, the mechanisms that regulate AR function in prostate cancer are poorly known. We report here a new paradigm for the forkhead protein FoxA1 action in androgen signalling. Besides pioneering the AR pathway, FoxA1 depletion elicited extensive redistribution of AR-binding sites (ARBs) on LNCaP-1F5 cell chromatin that was commensurate with changes in androgen-dependent gene expression signature. We identified three distinct classes of ARBs and androgen-responsive genes: (i) independent of FoxA1, (ii) pioneered by FoxA1 and (iii) masked by FoxA1 and functional upon FoxA1 depletion. FoxA1 depletion also reprogrammed AR binding in VCaP cells, and glucocorticoid receptor binding and glucocorticoid-dependent signalling in LNCaP-1F5 cells. Importantly, FoxA1 protein level in primary prostate tumour had significant association to disease outcome; high FoxA1 level was associated with poor prognosis, whereas low FoxA1 level, even in the presence of high AR expression, predicted good prognosis. The role of FoxA1 in androgen signalling and prostate cancer is distinctly different from that in oestrogen signalling and breast cancer.
Project description:HNF4G is a gastrointestinal tissue enriched master transcriptional regulator seen overexpressed in a subset of prostate cancer. Here we have mapped binding sites of HNF4G, AR, Foxa1, H3K4me1, H3K27acetyl upon knockdown and overexpression of HNF4G in in 22Rv1 and LNCaP cells respectively Overall design: ChIPSeq mapping of HNF4G, AR, Foxa1, H3K4me1, H3K27acetyl upon knockdown and overexpression of HNF4G in in 22Rv1 and LNCaP cells respectively
Project description:Our data suggested that FKHD-MSs impair the chromatin binding of FOXA1 to AR-dependent enhancers and thus suppress AR transcriptional activity, and promote PCa progression through increasing FOXA1 binding to a subset of AR-independent enhancers that regulates transcription of genes mediating EMT and metastasis Overall design: We generated LNCaP stable cells with tetracycline-inducible expression of V5-tagged FKHD-MSs or WT FOXA1. FOXA1 ChIP-seq experiments were performed on these cell lines