FIG. 3.

Second RRM of GRSF-1 is required for stimulation of translation in vitro. Equal moles (2.5 × 10⁻¹² mol) of GST, wild-type, and deletion mutant GST-GRSF-1 proteins were incubated with in vitro translation-competent influenza virus-infected HeLa cell extracts containing 200 ng of capped NP-luciferase or NP-A-luciferase chimeric mRNAs in a 12-μl final volume, as described in Materials and Methods. Reactions were prepared in triplicate and incubated for 60 min at room temperature, and the reactions were terminated by the addition of 9 volumes of 1× PBS, pH 7.5. Luciferase activity in 25-μl aliquots of each reaction was determined with the luciferase assay kit (Promega). These data are representative of two independent experiments.