2002). The Golgi complex plays an important role in the maturation, sorting, and transport of newly synthesized secreted lysosomal and plasma membrane proteins. In mammalian cells, this organelle selleck inhibitor is built up of stacks of flattened cisternae grouped together in the pericentrosomal region. Adjacent stacks are connected to each other by lateral tubules, forming a continuous ribbon-like structure (Rambourg and Clermont 1990). Each stack has a polarized structure and may be considered to be composed of at least three compartments: the cis-Golgi network (CGN)/cis-Golgi, the medial-Golgi cisternae, and the trans-Golgi/trans-Golgi network (TGN). The CGN/cis-Golgi, which receives newly synthesized proteins from the ERGIC, is where the first Golgi-specific glycosylation reactions occur.

Each medial-Golgi cisterna contains specific enzymes that sequentially act to allow the addition or trimming of carbohydrate moieties. The trans-Golgi/TGN ensures the final glycosylation reactions and the sorting of proteins to the plasma membrane and lysosomes. The structural integrity of the Golgi complex is indispensable in the accurate sorting and transport of proteins to their final destination. Thus, during mitosis, the Golgi complex breaks down into small vesicles and the protein transport along the secretory pathway is arrested (Lowe et al. 1998; Thyberg and Moskalewski 1998). Various drugs, such as brefeldin A and okadaic acid, have a profound effect on the integrity of Golgi stacks, causing perturbations in protein maturation, sorting, and trafficking (Fujiwara et al. 1988; Lucocq et al.

1991; Tamaki and Yamashina 2002). Microtubule-disrupting agents, such as nocodazole, also induce dispersal of Golgi stacks (Rogalski and Singer 1984; Thyberg and Moskalewski 1985; Cole et al. 1996), often leading to a retardation in the transport of secreted and plasma membrane proteins (Matter et al. 1990; Robin et al. 1995; Cole et al. 1996). To determine whether the perturbations of CA IV traffic observed in CF cells result from changes in the Golgi complex, we analyzed the structure and distribution of this organelle with regard to the microtubule network in the pancreatic duct cell line CFPAC-1. We found a dispersal of Golgi stacks associated with disorganization of the microtubule cytoskeleton and an increase in the number of microtubule-organizing centers (MTOCs).

Moreover, the reversion of CF cells by the wild-type CFTR led to the restoration of Golgi complex and microtubule distribution that allowed the correct trafficking of CA IV. Materials and Methods Cell Lines We conducted this study using the cancerous human pancreatic duct cell lines CFPAC-1, CFPAC-PLJ-CFTR6, and CFPAC-PLJ6. The CFPAC-1 cell line was Cilengitide established from a hepatic metastasis in a 26-year-old Caucasian male bearing a pancreatic adenocarcinoma and with CF (homozygote ��F508) (Schoumacher et al. 1990).