Pathway enrichment analysis was conducted via gene-set enrichment analysis (GSEA) and over-representation analysis (ORA)-based approaches. For each tissue, GSEA was performed on the DESeq2 normalized expression signals via a running-sum statistic procedure [8 ,9 (link)] to determine the enrichment of a priori defined biological pathways from the Kyoto Encyclopedia of Genes and Genomes (KEGG) repository [10 (link)], obtained from the Molecular Signature Database (MSigDB)[11 (link)]. Statistical significance of pathway enrichment was ascertained by permutation testing over size-matched random gene-sets. Adjustments for multiple testing were performed via control of the FDR [12 (link)]. Overlap between significant pathways were visualized via the EnrichmentMap Cytoscape plugin [13 (link)] using the following filters–pathway p-value ≤ 0.005, q-value ≤ 0.1, overlap ≥ 50%.
ORA was conducted using Qiagen’s Ingenuity Pathway Analysis (IPA) tool (Qiagen, USA) on differentially expressed genes with p < 0.01 and absolute fold-change ≥ 1.5-fold, for each tissue. Statistical significance of over-represented pathways was ascertained via Fisher's exact test and adjusted for multiple testing via the FDR according to Benjamin-Hochberg [8 ].
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