Characterization of epigenetic changes and their connection to gene expression abnormalities in clear cell renal cell carcinoma
"Clear cell renal cell carcinoma (ccRCC) is the most common kidney cancer subtype comprising 70-75% of all adult kidney malignancies. Although the genetic mechanisms of ccRCC have been widely studied, the mechanisms underlying its epigenetic modifications are not yet well understood, despite the evidence on genome-wide epigenome abnormalities in ccRCC. Therefore, we identified and investigated the ccRCC-associated epigenetic alterations on cis-regulatory elements, including distal enhancers and proximal regulatory elements that are close to transcription start sites (TSSs), and their contribution to gene expression changes in ccRCC. Using a machine-learning approach to combine ChIP-seq and whole genome bisulphite sequencing data of tumors and adjacent normal kidney tissues of ccRCC patients, we identified thousands of gained and lost cis-regulatory elements. Amongst them, ~71% are novel elements that had not been reported in ccRCC. Using a second machine-learning classifier, we showed that the dysregulated cis-regulatory elements are predictive of the genes with abnormal expression patterns in ccRCC (minimum area under ROC curve: 0.88, P-value <2.59 x 10-73). As such, gained and lost enhancers are significantly associated with up-regulated and down-regulated genes in ccRCC, respectively. Our analysis of the binding sites of 161 regulatory factors, including transcription factors and specific epigenome modifiers, revealed several regulatory factors whose binding sites were enriched in gained or lost enhancers. The target genes of these regulatory factors were enriched among up or down regulated genes in ccRCC consistent with the status of the associated enhancers (gained or lost). Among the top-ranked activated regulatory factors identified in our analysis, FOXM1, SPI1, IKZF1, JUNB, JUN, FOS, BCL11A and STAT3 are significantly enriched in gained enhancers, while EZH2, FOXA1, FOXA2, GATA3, ESR1 are significantly associated with lost enhancers (FDR <0.05). The target genes of these activated regulatory factors are involved in biological pathways that are central to the biology and function of ccRCC cells. For example, HIF1α and HIF2α transcription factor network, VEGF and VEGFR signaling network and immune system-related pathways are enriched in up regulated target genes in ccRCC, associated with gained enhancers (FDR <0.01). Finally, we sought to examine potential involvement of von Hippel-Lindau (VHL), the most frequently mutated gene in ccRCC, in these epigenome alterations, given the recent reports on VHL function as a regulator of the epigenome. Our analysis of VHL-deficient ccRCC cell lines and their wild-type VHL- reconstituted counterparts revealed that ~10% of gene expression changes in ccRCC can be recovered by VHL-driven epigenome alterations on enhancers. We uncovered several regulatory factors that could be regulated by VHL-mediated DNA methylation and that are associated with dysregulated enhancers, including BATF, BCL11A, IKZF1, JUN, SPI1, STAT3 and EZH2. In conclusion, our study proposes a new method to identify active (gained) and inactive (lost) cis-regulatory elements by combining histone modifications and DNA methylation data through machine-learning. In addition, our results provide new insights into the functional consequences of dysregulation of cis-regulatory elements on gene expression patterns in ccRCC. These findings will improve our understanding of the epigenetic mechanisms underlying transcriptome aberrations in ccRCC, which may eventually lead to the development of new preventive or therapeutic interventions"-- Author's abstract
Thesis, Dissertation, English, 2019
McGill University Libraries, [Montreal], 2019