Linking Proteomic and Transcriptional Data through the Interactome and Epigenome Reveals a Map of Oncogene-induced Signaling
A. The number of nodes in networks constructed from multiple approaches and their overlap with the PCST solution. NN of pY termini: the proteins containing phosphorylated tyrosine residues reported by mass-spectrometry and their direct interactors (nearest neighbors) in the interactome. NN of TF termini: transcription factor candidates selected by the expression regression procedure and their direct interactors in the interactome. NN of all termini: the union of pY termini, TF termini and their direct interactors in the interactome. RN: a network constructed by using a flow based approach ResponseNet  to connect the pY termini to the TF termini. B. GBM gene ranker scores for nodes included in the PCST solution were significantly higher than the nodes excluded from the PCST solution (labeled as “Interactome excl. PCST”; p<2.2E-16 by Wilcoxon rank-sum test) and compared favorably to the nearest neighbor networks. Higher GBM scores indicate greater relevance to the disease. C. Scoring proteins by connectivity to the PCST solution representing a disease network. The score of each protein, whether the protein is inside or outside of the original PCST network, is the sum of the scores of all its interactions with the nodes in the PCST. Thus a node in the interactome (deep red) with many high confidence interactions to the nodes in the PCST disease network receives a higher score than a node in the interactome (light red) that has fewer or lower confidence interactions to the nodes in the PCST. D. Proteins with EGFRvIII regulated tyrosine phosphorylation in mouse xenografts (red bars) are more closely connected to the PCST solution than the proteins on which the tyrosine phosphorylation levels do not change significantly (green bars). Each protein in the interactome was scored then ranked by its connectivity to the PCST solution constructed from the U87 cell line data as described in B and in Materials and Methods. P-value was computed by Wilcoxon rank-sum test comparing the ranks of EGFRvIII-specific and not EGFRvIII specific phosphorylated proteins. The number of proteins in each category is indicated in parentheses. E. The targets for transcription factors identified in condition-specific DNaseI hypersensitive regions are enriched for genes differentially expressed in response to EGFRvIII. U87H TF: transcription factors that have motif matches in regions with increased DNaseI hypersensitivity in the U87H cells and within 40 kb of transcription start sites. U87DK TF: transcription factors that have motif matches in the regions with higher DNaseI hypersensitivity in the U87DK cells and within 40 kb of transcription start sites. EGFRvIII up- and down-regulated genes: genes that are up- or down- regulated in the TCGA GBM exon array dataset comparing EGFRvIII positive samples to wild-type EGFR samples. For each TF, we computed a minimum hypergeometric (mHG) p-value that tested for the probability that the set of target genes are differentially expressed in the TCGA GBM samples by chance. Top panel: U87H TF targets are more enriched (smaller mHG values) in EGFRvIII up-regulated genes than in EGFRvIII down-regulated genes. Bottom panel: U87DK TF targets are more enriched in EGFRvIII down-regulated genes than in EGFRvIII up-regulated genes. P-values were computed by Student's t-test comparing the mHG p-values on EGFRvIII up- and down-regulated genes for each set of TF. F. The transcription factors included in the PCST solution are more enriched in EGFRvIII-induced differential gene expression than the transcription factors excluded from the PCST. Each set of U87H TF and U87DK TF were further divided into whether they were included in the PCST solution, denoted by the “Yes” and “No” categories. First panel: targets of U87H TF included in the PCST solution have stronger enrichment in EGFRvIII up-regulated genes than targets of the TF excluded from the solution. Fourth panel: targets of U87DK TF included in the PCST solution have stronger enrichment in EGFRvIII down-regulated genes than targets of the TF excluded from the PCST. Second and third panel: with respect to the comparison between U87H TF targets and EGFRvIII down-regulated genes, or between U87DK TF targets and EGFRvIII up-regulated genes, the TF included in the PCST do not show significantly stronger enrichment than the TF excluded from the PCST. P-values were computed by Student's t-test comparing the mHG scores of TF included in the PCST and TF excluded from the PCST.