A novel role for clathrin in mediating assembly of the Golgi apparatus

Abstract

The Golgi apparatus is the central processing and sorting station for lipids and proteins in the secretory pathway and it consists of a network of stacked, specialized cisternae localized to the perinuclear region of the cell.;The structural integrity of the Golgi apparatus is maintained by a proteinaceous matrix, cytoskeletal components and inositol phospholipids. In mammalian cells the organelle disassembles reversibly at the onset of mitosis and irreversibly during apoptosis. Pharmacological agents cause the Golgi complex to fragment into vesicles, tubules and ministacks by disrupting specific components of its architecture. Our laboratory has shown previously that alcohol treatment induces Golgi fragmentation and upon its removal, the Golgi apparatus rapidly regains its characteristic morphology. Interestingly, Golgi reassembly was accompanied by recruitment of abundant clathrin coated vesicles (CCVs) to the reforming Golgi cisternae. Moreover, TGN adaptors and SNARES co-localized with clathrin positive structures at the reforming Golgi complex. These results suggested that clathrin and/or CCVs may provide a scaffold for reassembly of Golgi cisternae into an intact structure by virtue of differential cargo and adaptor protein segregation.;A main goal of this thesis project was to test the hypothesis that clathrin is required for Golgi reformation and to dissect its role in organelle reassembly. Knockdown of endogenous clathrin heavy chain (CHC) showed that clathrin was not essential for maintaining Golgi morphology or fragmentation. Strikingly, CHC was required for Golgi reformation, independent of the mechanism used to disassemble the organelle. In addition, I identified a novel short-lived, putative assembly intermediate which has a "tight" compact morphology. Kinetic analysis suggests that clathrin might act at two stages, initially regulating the rate of transition from vesicles to the tight intermediate and subsequently to mediate assembly of Golgi cisternae.;We propose that an extension of these studies may provide a model for Golgi biogenesis at the completion of mitosis. Both clathrin heavy and light chains as well as the golgin p115 segregate with the mitotic spindle and the centrosome throughout mitosis and we suggest that these structures may act as a nucleating site for Golgi vesicle fusion in the pericentrosomal area of the cell.