Incorporation of an Internal Ribosome Entry Site–Dependent Mechanism in Arsenic-Induced GADD45α Expression

Abstract

We have previously shown that trivalent arsenic (arsenite, As 3+ ) is able to induce GADD45A expression in human bronchial epithelial cells through activation of c-Jun NH 2 -terminal kinase and nucleolin-dependent mRNA stabilization. In the present report, we show that As 3+ is capable of inducing translation of the GADD45A protein through a cap-independent, or rather, an internal ribosome entry site (IRES)-dependent mechanism. In growth-arrested cells, As 3+ elevated the GADD45A protein level in a dose-and time-dependent manner which did not correlate with the GADD45A 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 GADD45A protein induced by As 3+ . Sequence analysis revealed a potential IRES element in the 5 ¶-untranslated region of the GADD45A mRNA. This IRES element in the 5 ¶-untranslated region of the GADD45A mRNA is functional in mediating As 3+ -induced translation of the GADD45A protein in a dicistronic reporter gene activity assay. Immunoprecipitation and proteomic studies suggest that As 3+ 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, As 3+ is still capable of inducing GADD45A expression through an IRES-dependent translational regulation. [Cancer Res 2007;67(13):6146-54]

Analysis of GADD45A sequence variations in French Canadian families with high risk of breast cancer

“…In addition, GADD45A expression has been recently shown to be regulated through a cap-independent and internal ribosome entry site (IRES)-dependent mechanism in response to cellular stress signals such as arsenic-induced cytotoxic and genotoxic damage (Chang et al 2007). This expression is dependent on an IRES element located in the 5 0 UTR region proximal to the start codon.…”

A Novel Post-translational Modification of Nucleolin, SUMOylation at Lys-294, Mediates Arsenite-induced Cell Death by Regulating gadd45α mRNA Stability

“…tion (18), arsenite (28,47,48,69), and inflammatory cytokines (70). Our previous findings showed that GADD45␣ was one of the crucial mediators of arsenite-induced apoptosis (47,48).…”

“…The cell growing conditions seem critical for the gadd45␣ mRNA induction by arsenite (69). When the cells were in a logarithmically growing condition, arsenite resulted in a timedependent increase in gadd45␣ mRNA in the late phase of exposure (47,69).…”

“…The cell growing conditions seem critical for the gadd45␣ mRNA induction by arsenite (69). When the cells were in a logarithmically growing condition, arsenite resulted in a timedependent increase in gadd45␣ mRNA in the late phase of exposure (47,69). However, if the cells were growing in the arrested condition, in which the cells reached almost confluence with a high density, gadd45␣ mRNA was not induced by arsenite (48); rather, it was decreased with prolonged exposure time (69).…”

“…When the cells were in a logarithmically growing condition, arsenite resulted in a timedependent increase in gadd45␣ mRNA in the late phase of exposure (47,69). However, if the cells were growing in the arrested condition, in which the cells reached almost confluence with a high density, gadd45␣ mRNA was not induced by arsenite (48); rather, it was decreased with prolonged exposure time (69). Our current studies showed that arsenite exposure did increase the gadd45␣ mRNA level; however, the transcriptional regulation is unlikely to be the main mechanism of arsenite-induced expression of GADD45␣.…”