SEX-LETHAL INTERACTIONS WITH PROTEIN AND RNA - ROLES OF GLYCINE-RICH AND RNA-BINDING DOMAINS

Sex-lethal (Sxl) is an RNA-binding protein, containing two conserved R NA binding domains (RBDs) and a glycine-rich region, which functions a s a regulator of alternative splicing in Drosophila sex determination, Previous work demonstrated that Sxl monomers interact cooperatively u pon binding to target RNAs and that the cooperativity depends on the g lycine-rich N terminus, Here we use band shift experiments to show tha t RNA binding patterns are altered when Sxl is combined with other pro teins having similar glycine-rich domains, including mammalian heterog eneous nuclear (hn) RNP L and Drosophila Hrb87F (an hnRNP A/B homolog) . Direct involvement of the Sxl glycine-rich region in protein interac tions was verified by Far-Western analysis, Two interaction domains, t he Sxl N terminus and the Sxl first RNA binding domain, were suggested by the yeast two-hybrid assay, In a systematic examination of the RNA binding properties of Sxl domains, it was found that the Sxl termini as well as the RBDs influence RNA binding specificity, Finally, select ion of the Sxl optimal binding site (SELEX) confirms the importance of U-runs in the Sxl binding site and suggests a second type of non-U-ru n target that may be associated with RNA secondary structure.