Project description:Genome-wide association studies (GWASs) have identified thousands of single nucleotide polymorphisms (SNPs) associated with human traits and diseases. But because the vast majority of these SNPs are located in the noncoding regions of the genome their risk promoting mechanisms are elusive. Employing a new methodology combining cistromics, epigenomics and genotype imputation we annotate the noncoding regions of the genome in breast cancer cells and systematically identify the functional nature of SNPs associated with breast cancer risk. Our results demonstrate that breast cancer risk-associated SNPs are enriched in the cistromes of FOXA1 and ESR1 and the epigenome of H3K4me1 in a cancer and cell-type-specific manner. Furthermore, the majority of these risk-associated SNPs modulate the affinity of chromatin for FOXA1 at distal regulatory elements, which results in allele-specific gene expression, exemplified by the effect of the rs4784227 SNP on the TOX3 gene found within the 16q12.1 risk locus. Examination of histone modification H3K4me2 in untreated and E2 treated cells
Project description:Understanding the regulatory landscape of the human genome is a central question in complex trait genetics. Most single nucleotide polymorphisms (SNPs) associated with cancer risk lie in non protein-coding regions, implicating regulatory DNA elements as functional targets of susceptibility variants. Here, we describe genome-wide annotation of regions of open chromatin and histone modification in fallopian tube and ovarian surface epithelial cells (FTSECs, OSECs), the debated cellular origins of high-grade serous ovarian cancers (HGSOCs), and in endometriosis epithelial cells (EECs), the likely precursor of clear cell ovarian carcinomas (CCOCs). The regulatory architecture of these cell types was compared to normal human mammary epithelial cells (HMECs) and LNCaP prostate cancer cells. We observed similar positional patterns of global enhancer signatures across the three different ovarian cancer precursor cell types, and evidence of tissues specific regulatory signatures to non-gynecological cell types. We found significant enrichment for risk-associated SNPs intersecting regulatory biofeatures at 17 known HGSOC susceptibility loci in FTSECs (P=3.8x10-30) OSECs (P=2.4x10-23) and HMECs (P=6.7x10-15) but not for EECs (P=0.45) or LNCaP cells (P=0.88). Hierarchical clustering of risk SNPs conditioned on the six different cell types indicates FTSECs and OSECs are highly related (96% of samples using multi-scale bootstrapping) indicating both cell types may be precursors of HGSOC. These data represent the first description of regulatory catalogues of normal precursor cells for different ovarian cancer subtypes, and provide unique insights into the tissue specific regulatory variation with respect to the likely functional targets germline genetic susceptibility variants for ovarian cancer FAIRE-Seq and ChIP-Seq of 2 different histone modifications in 5 cell types.
Project description:We adapted the DiR barcode-based parallel reporter assay systems strategy to systematically identify the breast cancer related SNPs that affect gene expression by modulating activities of regulatory elements. Among 293 SNPs linked with GWAS-identified breast cancer-risk SNPs, we found seven functional regulatory SNPs in MCF7 cells. Further mechanism study indicates that one SNP regulates gene expression in breast cancer malignancy. The DiR system has great potential to advance the functional study of risk SNPs that have associations with polygenic diseases. Our findings hold great promise in benefiting breast cancer patients with prognostic prediction.
Project description:We have sequenced miRNA libraries from human embryonic, neural and foetal mesenchymal stem cells. We report that the majority of miRNA genes encode mature isomers that vary in size by one or more bases at the 3’ and/or 5’ end of the miRNA. Northern blotting for individual miRNAs showed that the proportions of isomiRs expressed by a single miRNA gene often differ between cell and tissue types. IsomiRs were readily co-immunoprecipitated with Argonaute proteins in vivo and were active in luciferase assays, indicating that they are functional. Bioinformatics analysis predicts substantial differences in targeting between miRNAs with minor 5’ differences and in support of this we report that a 5’ isomiR-9-1 gained the ability to inhibit the expression of DNMT3B and NCAM2 but lost the ability to inhibit CDH1 in vitro. This result was confirmed by the use of isomiR-specific sponges. Our analysis of the miRGator database indicates that a small percentage of human miRNA genes express isomiRs as the dominant transcript in certain cell types and analysis of miRBase shows that 5’ isomiRs have replaced canonical miRNAs many times during evolution. This strongly indicates that isomiRs are of functional importance and have contributed to the evolution of miRNA genes