Hypoxia-induced changes in gene expression are predominantly controlled at the level of transcription. (A) Schematic diagram depicting the experimental procedure and the information provided by each RNA fraction. (B) Venn diagrams showing the overlap between the genes found differentially overexpressed (FDR < 0.05) in the total (Total Up) and newly transcribed (Newly Up) fractions and the differentially down-regulated (FDR < 0.05) in the total (Total Down) and newly transcribed (Newly Down) fractions. (C) The graph represents the proportion of genes in the total mRNA fraction that were also differentially expressed in the newly transcribed fraction for: all regulated genes (black line), significantly down-regulated genes (purple line), and significantly up-regulated genes (orange line). Genes were considered differentially expressed if, in addition to being significantly regulated (FDR < 0.05), showed an absolute value of induction above the indicated threshold of logFC (x-axis). The magnitude of change induced by hypoxia was calculated for each gene as the absolute value of the logarithm of the ratio of the expression level in normoxia over hypoxia (logFC). (D) The magnitude of change (log2-fold change, logFC) induced by hypoxia in 4sU-labeled genes (“Newly tr mRNA”) was represented against the changes in the total mRNA fraction (“Total mRNA”). Genes significantly (FDR < 0.01) regulated by hypoxia in either fraction are labeled in green. Regression analysis of significantly regulated genes showed that total mRNA level increased significantly with transcription (F(1,2225) = 5756, P-value << 0.01).
