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. 2006 Jun 6;34(10):3044–3056. doi: 10.1093/nar/gkl386

Figure 5.

Figure 5

Role of hnRNP K and hnRNP E2/E1 in NO•-Erk1/2-CURE signaling. (A) RNA REMSAs with either a consensus (left panel) or MAP3K7IP2 (right panel) CURE riboprobes. GSNO (400 µM) treatment for 3 h increased complex formation compared to control GSH; anti-hnRNP K and anti-hnRNP E2/E1 both super-shift the complex; the unlabeled CURE riboprobes, but not the mutant of consensus CURE (mutant CURE) compete with the labeled CURE riboprobes. (B) Translocation of hnRNP K and hnRNP E2/E1 to the cytoplasm by western blotting. GSNO (400 µM) treatment for 3 h increased the presence of hnRNP K and hnRNP E2/E1 in the cytoplasm but not in whole-cell lysates compared to control GSH. This effect was further enhanced by the p38 MAPK inhibitor SB202190 (SB; 0.1 µM), but blocked by the Erk1/2 inhibitor PD98059 (PD; 30 µM). A control protein α tubulin is shown for comparison. Experiments in (A and B) were repeated at least twice with similar results. (C) Overexpression of hnRNP K mimicked the effect of NO•, repressing the expression of a chimeric LUC-CURE reporter gene. THP-1 cells were co-transfected with pGL3/CURE, pGL3/CUREmut or control pGL3 and pcDNA3 (empty vector) or phnRNP-K (hnRNP K expression plasmid). After treatment with GSH (400 µM) or GSNO (400 µM) for 20 h, LUC activities were measured. Data, presented as percentage relative to LUC activity with pGL3, are the mean ± SEM of three independent experiments.