Fig. 5

Selective interaction of CYFIP1/2 with FMRP, FXR1P, and FXR2P. (A) Interaction between the CYFIP proteins and the members of the FXR family, determined by yeast two-hybrid system. Diagram illustrating activation of the β-gal reporter in yeast coexpressing the following proteins (from Left to Right): FMRP + CYFIP1, FXR1P + CYFIP1, FXR2P + CYFIP1, FMRP + CYFIP2, FXR1P + CYFIP2, and FXR2P + CYFIP2, measured by liquid ONPG assay. At the foot of each bar, a X-gal filter lift assay is shown. Blue (black) color indicates β-gal activity induced by interaction of the two transformed fusion proteins. CYFIP2 interacts also with the FMRP-related proteins FXR1P/2P, whereas CYFIP1 interaction is restricted to FMRP. (B) The His₆ pull-down assay (His-pd) assay using His₆-tagged full-length CYFIP1 and in vitro translated proteins FMRP iso7 (lane 1), FXR1P iso a/P₇₀ (lane 2), FXR2P (lane 3), and luciferase (lane 4). His-pd assays were performed in a buffer containing 200 mM sodium chloride. Loadings were performed as indicated in the legend to Fig. 1. FMRP bound specifically to CYFIP1, whereas FXR1P, FXR2P, and the negative control luciferase were not retained. None of the proteins is retained by beads incubated with extract from SF9 cells infected by wild-type baculovirus.