Specific interactions of HeLa cell proteins with Coxsackievirus B3 RNA: La autoantigen binds differentially to multiple sites within the 5' untranslated region

Abstract

Translation initiation of the coxsackievirus B3 (CVB3) RNA occurs by internal ribosomal entry. The internal ribosomal entry site (IRES) of the virus has been mapped to the 5' untranslated region (5' UTR) of the genome. As well, the 5' UTR has been suggested to play roles in determining the tissue tropism and infectivity of the virus. In this study, we investigated interactions between HeLa cell protein extracts and radiolabeled RNA of CVB3 5' UTR by competitive UV cross-linking. We have observed a number of proteins that specifically interact with the three sub-cloned regions of the 5' UTR. In particular, the molecular weights of five of these proteins resemble those of the eukaryotic translation initiation factors 4A, 4B and 4G, as well as the La autoantigen and the polypyrimidine tract binding protein. Based on this data, we focused on the interaction of the 5' UTR with the La autoantigen, which was purified by the glutathione-S-transferase affinity method. We have confirmed the highly specific interaction of the La autoantigen with the 5' UTR sequence nt 210-529. The core IRES (nt 530-630) and nt 1-209 also appear to bind to the La protein at moderate and weak affinities, respectively. A functional role of the La autoantigen in translation initiation is suggested.