Conformational-Dependent and Independent RNA Binding to the Fragile X Mental Retardation Protein
Figure 7
Annealed BC1 RNA binds pleiotropically to the FXRPs. (a) Equimolar amounts of 35S-FMRP, 35S-FMRP15c, 35S-FXR1P, 35S-FXR2P, 35S-eIF4A, 35S-FMRP280, 35S-FMRP204, and 35S-Luciferase were incubated with 2 μM annealed BC1 RNA. The percent binding corrected for background of 6 reactions per protein is plotted. The relative levels of FMRP and FMRP15c, FMRP and FXR1P and FMRP and FXR2P were not significantly different (, and , resp., ANOVA). However, the relative levels of FMRP and eIF4A and FMRP and NTD204 were significantly different ( and , resp., ANOVA). Since no binding above background was observed for NTD280 and Luciferase relative differences were not assessed. (b) Pull-down assays between 35S-luciferase and poly (rG), poly (rA), eEF1A RNA (90 nM) and annealed BC1 RNA (5 μM) showing the unbound (U) and bound (B) fractions. Binding in the absence of RNA is shown for the negative control. (c) Pull-down assays between 35S-NTD280 or 35S-NTD204 and poly (rG), poly (rA), poly (rI : rC) and annealed BC1 RNA (5 μM) showing the unbound (U) and bound (B) fractions. Binding in the absence of RNA is shown for the negative control.
