Abstract
BackgroundPancreatic ductal adenocarcinoma initiation is most frequently caused by Kras mutations.ResultsHere, we apply biological, biochemical, and network biology methods to validate GEMM-derived cell models using inducible KrasG12D expression. We describe the time-dependent, chromatin remodeling program that impacts function during early oncogenic signaling. We find that the KrasG12D-induced transcriptional response is dominated by downregulated expression concordant with layers of epigenetic events. More open chromatin characterizes the ATAC-seq profile associated with a smaller group of upregulated genes and epigenetic marks. RRBS demonstrates that promoter hypermethylation does not account for the silencing of the extensive gene promoter network. Moreover, ChIP-Seq reveals that heterochromatin reorganization plays little role in this early transcriptional program. Notably, both gene activation and silencing primarily depend on the marking of genes with a combination of H3K27ac, H3K4me3, and H3K36me3. Indeed, integrated modeling of all these datasets shows that KrasG12D regulates its transcriptional program primarily through unique super-enhancers and enhancers, and marking specific gene promoters and bodies. We also report chromatin remodeling across genomic areas that, although not contributing directly to cis-gene transcription, are likely important for KrasG12D functions.ConclusionsIn summary, we report a comprehensive, time-dependent, and coordinated early epigenomic program for KrasG12D in pancreatic cells, which is mechanistically relevant to understanding chromatin remodeling events underlying transcriptional outcomes needed for the function of this oncogene.
Highlights
ResultsWe apply biological, biochemical, and network biology methods to validate Genetically engineered cells and mouse models (GEMMs)-derived cell models using inducible KrasG12D expression
Pancreatic ductal adenocarcinoma initiation is most frequently caused by Kras mutations
In summary, we report a comprehensive, time-dependent, and coordinated early epigenomic program for KrasG12D in pancreatic cells, which is mechanistically relevant to understanding chromatin remodeling events underlying transcriptional outcomes needed for the function of this oncogene
Summary
Cells derived from genetically engineered mouse models recapitulate early oncogenic KrasG12D signaling in vitro First, we sought to better understand how KrasG12D coordinates over time, the reorganization of the genome. Two-thirds of the genes expressed in KrasG12D cells displayed enrichment of these 3 marks at 24 hrs This pattern of histone-based chromatin remodeling correlated with expression of upregulated genes in RNA-seq, hypomethylation of DNA promoters by RRBS, and increased accessible chromatin regions by ATAC-seq (Fig. 9b–d; State 2). We integrated RNA-seq expression patterns with all epigenomic data sets by a hierarchical clustering-based method which illustrated that KrasG12D cells displayed a total of 859 DEGs with upregulated genes mapping to accessible chromatin regions (ATAC-seq) and enrichment for H3K27ac, H3K4me, and H3K36me marks (Fig. 9g). This study integrates knowledge derived from multiple datasets, generated in a controlled and time-dependent manner, to account for many features shown to characterize the KrasG12D phenotype in GEMM-derived pancreatic cells
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