Hotspots of aberrant enhancer activity in fibrolamellar carcinoma reveal molecular mechanisms of oncogenesis and intrinsic drug resistance

Timothy A. Dinh; Ramja Sritharan; F. Donelson Smith; Adam B. Francisco; Rosanna K. Ma; Rodica P. Bunaciu; Matt Kanke; Charles G. Danko; Andrew P. Massa; John D. Scott; Praveen Sethupathy

TRE annotation was performed using the annotatePeaks.pl function from HOMER (Heinz et al., 2010) based on GENCODE v25 annotations. Transcription factor motif enrichment analysis was performed using the findMotifsGenome.pl function from HOMER using "given" as the size parameter. The input (and background) peaks run were FLC-specific TREs (all TREs that were not identified as FLCspecific as background), NML-specific TREs (all TREs not identified as NMLspecific), FLC-specific TREs without FOSL2/JUN or CREB motifs (all TREs not identified as FLC-specific), FLC-specific promoters (all promoters not identified as FLC-specific), and FLC-specific enhancers (all enhancers not identified as FLC-specific). CpG island annotations were downloaded from the UCSC Table Browser. Hierarchical clustering was performed following DESeq2 normalization of read counts quantified from each TRE or gene body as described above. Clustering was performed in a pairwise manner using a correlation-based distance metric (1 -Spearman's rho) using Ward's minimum variance method. Proximal (-1000 to +100 bp from TSS) and distal TREs (the remaining TREs) were classified based on GENCODE v25 transcript annotations. Principal components analysis was performed following Variance Stabilizing Transformation from DESeq2.