In Vivo Association of the Stability Control Protein αCP with Actively Translating mRNAs

Posttranscriptional controls play a major role in eucaryotic gene expression. These controls are mediated by sequence-specific interactions of cis-acting determinants in target mRNAs with one or more protein factors. The positioning of a subset of these mRNA-protein (RNP) complexes within the 3' untranslated region (3' UTR) may allow them to remain associated with the mRNA during active translation. Robust expression of human alpha-globin mRNA during erythroid differentiation has been linked to formation of a binary complex between a KH-domain protein, alphaCP, and a 3' UTR C-rich motif. Detection of this "alpha-complex" has been limited to in vitro studies, and the functional state of the alpha-globin mRNA targeted by alphaCP has not been defined. In the present study we demonstrate that a significant fraction of alphaCP is associated with polysomal mRNA. Targeted analysis of the polysomal RNP complexes revealed that alphaCP is specifically bound to actively translating alpha-globin mRNA. The bound alphaCP is restricted to the poly(C)-rich 3' UTR motif and is dislodged when ribosomes are allowed to enter this region. These data validate the general importance of the 3' UTR as a sheltered site for RNP complexes and support a specific model in which the stabilizing function of alphaCP is mediated on actively translating target mRNAs.