c-IAP1 is localized to the Golgi apparatus in differentiated cells. (A) Western blot analysis of c-IAP1 (pAb; Santa Cruz Biotechnology) expression in the mitochondrial (M), cytosolic (C), reticular/microsomal (R), and nuclear (N) fractions obtained from U937 cells before (Co) and after exposure to 20 nM TPA for 72 hours. The expression of poly(ADP-ribose)polymerase (PARP), Golgin 97, and mitochondrial HSP70 was used to assess the enrichment of each cell fraction. (B) THP1 cells were treated with TPA for 48 hours before analyzing the expression of c-IAP1 (red), Golgin 97 (green), or GM130 (green) by confocal microscopy (magnification × 300). (Insets) Increased magnification of Golgi labeling (magnification × 3000). (C) c-IAP1 expression in TPA-differentiated THP1 cells before (Co) and after exposure to either brefeldin A (BFA; 5 μg/mL; 2 h 30 min) or nocodazole (10 μM; 1 h). c-IAP1 expression was observed by fluorescence microscopy (magnification ×700) using an anti-c-IAP1 mAb (Pharmingen). (D) Western blot analysis of c-IAP1 expression under limited proteolytic digestion of the reticular/microsomal fraction of TPA-differentiated U937 cells. Golgin 97 and protein disulfide isomerase (PDI) are used as positive and negative controls, respectively.