Incorporation of an internal ribosome entry site-dependent mechanism in arsenic-induced GADD45α expression

We have previously shown that trivalent arsenic (arsenite, As3+) is able to induce GADD45 alpha expression in human bronchial epithelial cells through activation of c-Jun NH2-terminal kinase and nucleolin-dependent mRNA stabilization. In the present report, we show that As3+ is capable of inducing translation of the GADD45 alpha protein through a cap-independent, or rather, an internal ribosome entry site (IRES)-dependent mechanism. In growth-arrested cells, As 3, elevated the GADD45 alpha protein level in a dose- and time-dependent manner which did not correlate with the GADD45 alpha mRNA expression. Pretreatment of the cells with rapamycin, an inhibitor for the cap-dependent translation machinery through the suppression of mTOR and p70S6 kinase, failed to affect the induction of the GADD45 alpha protein induced by AS. Sequence analysis revealed a potential IRES element in the 5 '-untranslated region of the GADD45 alpha mRNA. This IRES element in the 5 '-untranslated region of the GADD45 alpha mRNA is functional in mediating As3+-induced translation of the GADD45 alpha protein in a dicistronic reporter gene activity assay. Immunoprecipitation and proteomic studies suggest that AS impairs the assembly of the cap-dependent initiating complex for general protein translation but increases the association of human elongation factor 2 and human heterogeneous nuclear ribonucleoprotin with this complex. Thus, these results suggest that in growth-arrested cells, As3+ is still capable of inducing GADD45 alpha expression through an IRES-dependent translational regulation.