Fig. 4.

GAr-induced mRNA translation stress stimulates E2F1 synthesis. a Autoradiograph shows the effect of GAr on newly synthesised E2F1 levels following ³⁵S-Met pulse labelling in the presence of a proteasome inhibitor (20 µM MG132) followed by E2F1 immunoprecipitation. b Western blots showing GAr-dependent induction of E2F1 from constructs carrying the untranslated (UTR) regions of the E2F1 mRNA (E2F1 UTR), or the coding sequence alone (E2F1 CDS). c Polysome profiling and RT-qPCR analysis of E2F1 mRNA in cells expressing the GAr or EV. The upper panels show polysome profiles. The lower panel shows the relative E2F1 mRNA levels (%) from total mRNA normalised against actin. The values represent the mean data from three independent experiments with s.d. d Schematic representation of functional domains of E2F1 deleted from the 5′ of the CDS. The +1 indicated the first AUG of the CDS. e Western blots showing the expression of E2F1 deletion mutants with and without GAr expression. Western blots represent n ≥ 3, actin was used as a loading control