The effect of RAP55 knock-down on P-bodies. (Panel I) Immunoblotting was used to determine the effect of siRNAs on the levels of RAP55 and DCP1a in Hep-2 cells. Compared with control siRNA, RAP55 siRNA decreased the level of (A) RAP55 but not (B) DCP1a in Hep-2 cells. As indicated, siRNA knock-down was performed in duplicate. RAP55 and DCP1a were detected using patient serum 0080 and rabbit anti-DCP1a antiserum, respectively. The human serum reacted weakly with E2 PDC (70 kDa). (Panel II) RAP55 siRNA decreased the level of RAP55, DCP1a, and Ge-1 in P-bodies. After transfection of RAP55 siRNA into Hep-2 cells, neither (A) RAP55 nor (B) DCP1a was detected in P-bodies. (D,E) Control siRNA did not alter the P-body location of either protein. (G,H) Transfection of RAP55 siRNA into Hep-2 cells also displaced Ge-1 from P-bodies. (J,K) Control siRNA had no effect on the P-body localization of Ge-1. Patient 0012 serum or rabbit anti-RAP55 antiserum was used to detect RAP55, as indicated. Patient 0050 serum was used to detect Ge-1. DAPI staining indicates the location of nuclei in C, F, I, and L.
