Fig. 8.

CIF1 structural motifs required for maintaining CIF2 protein stability. (A) Western blotting to examine the level of CIF2, which was tagged with an N-terminal PTP epitope from one of its endogenous loci, and the level of CIF1 and its mutants, which were tagged with a C-terminal triple HA epitope and ectopically overexpressed in cells containing the CIF1-3′UTR RNAi construct. Cells were induced with tetracycline for 72 h, and time-course samples were collected for western blotting with anti-Protein-A and anti-HA antibodies. TbPSA6 served as the loading control. Asterisks indicate a non-specific band detected by the anti-HA antibody. (B) Quantification of the level of CIF2 protein in CIF1-3′UTR RNAi cell line and CIF1-3′UTR RNAi cell lines expressing CIF1 and its mutants. CIF2 band intensity was determined by ImageJ and normalized with the intensity of TbPSA6. Error bars indicate s.d. calculated from three independent experiments.