FIG 8.

Imp7/9 but not Imp3 directly interact with JNK/p38 MAPKs. (A) An in vitro binding assay demonstrated a direct Imp7/9 interaction with JNK1/2 and p38α/β. Imp3/7/9 were IPed from either stimulated or untreated cells (NT) and extensively washed with RIPA buffer following LiCl (0.5 M) and buffer A. To examine the association, 500 ng of each indicated GST-MAPK was incubated with the indicated IPed Imps (2 h with rotation) following washing and resuspension with sample buffer. Interacting MAPKs were detected using Western blotting (IB) with the indicated Abs. (B) Imp3 is not required for Imp7/9 interactions with MAPKs. siRNAs of Imp3 and the siRNA control were transfected into HeLa cells, as described in Materials and Methods. Cell extracts were then subjected to CoIP with the indicated Abs. The amount of CoIPed MAPKs, Imp, and input Imps was detected by Western blotting with the indicated Abs. To confirm the efficiency of the siRNA, cells were transfected with siRNAs and then subjected to a Western blot analysis with the indicated Abs (lower panels).