Fig. 3.

Pelp1 is the core component of the 5FMC complex. A, Pelp1 is required for interaction between Senp3 and Wdr18. 293T cells were transiently transfected with expression vectors encoding T7_Pelp1, HA_Wdr18 and Myc_Senp3. Cell lysates were analyzed by IP and Western blotting with the antibodies indicated. B, Pelp1 is needed for the stability of the 5FMC complex. MEL_BirA cells were treated with the indicated shRNAs. Nuclear lysates were analyzed by Western blotting with the indicated antibodies. Actin staining serves as a loading control. Total RNA was extracted from MEL_BirA cells transduced with the indicated shRNA and analyzed by RT-QPCR for Las1L, Pelp1, Senp3, Tex10, and Wdr18. Error bars: S.D. of triplicate experiment. C, Mapping the interaction regions of Pelp1. Schematic representation of Pelp1 deletion constructs. GST fused Pelp1 domains and GST alone (GST_empty) were immobilized onto glutathione beads (lower panel stained with Coomassie) and used to pull down nuclear cell lysates from 293T cells expressing HA_Las1L, HA_Senp3 and HA_Wdr18. Asterisks indicate GST fusion proteins. D, Schematic representation of interaction between Pelp1, Las1L, Senp3, and Wdr18 protein domains.