Figure 3.

Competition of I-RNA with CVB3 5′-UTR for binding to La and rescue of CVB3 IRES-mediated translation by recombinant La protein. (A) [³²P]UTP labeled CVB3 5′-UTR RNA probe was UV crosslinked to His-tagged La protein in the absence (lane 1) and presence (lanes 2–4) of 200-, 400- and 600-fold molar excess of unlabeled I-RNA. (B) [³²P]UTP labeled CVB3 5′-UTR RNA probe was UV crosslinked to His-tagged La protein in the absence (lane 1) and presence (lanes 2 and 3) of 400- and 600-fold molar excess of a non-specific RNA. (C) CVB3 5′-UTR–GFP RNA was translated in vitro in RRL supplemented with 0.5 µg HeLa S10 (lanes 1–4), 50-fold excess of I-RNA (lanes 2–4) and two concentrations (200 and 400 ng) of La protein (lanes 3 and 4). The positions of the La–CVB3 RNA complex (A and B) and the translation product GFP (C) are indicated by arrows.