The unique and complex architecture of the Golgi apparatus has been conserved through most of eukaryotic evolution but the role it plays in the functioning of this organelle is far from clear. One reason has been the lack of knowledge concerning those structural proteins responsible for generating and maintaining the central architectural feature of the Golgi apparatus, the stacks of closely-apposed and flattened cisternal membranes with associated transport vesicles. By exploiting the dramatic transformation of the Golgi during mitosis in animal cells, we have identified a complex of two proteins that play a crucial role in Golgi architecture. GRASP65 is involved in stacking cisternae whereas GM130 mediates the docking (or tethering) or transport vesicles with cisternal rims.
Our aim i s to work our precisely how GRASP65 and GM130 carry out their functions and to identify other proteins that help them determine Golgi structure. These proteins will also be used as tools to manipulate Golgi architecture and so help us understand its relationship to Golgi function.
The specific aims of this proposal are: 1. To work out how GRASP65 stacks cisternae and whether additional proteins are required. If GRASP65 only stacks a subset of Golgi cisternae then the stacking mechanism of other cisternae will be studied. 2. To disrupt the architecture of the Golgi in vivo using GRASP65 antibodies and fragments as tools and to determine the effects on a variety of Golgi functions. 3. To elucidate the mechanism of vesicle tethering and the roles played by GM130, Giantin and p115. Biochemical and microscopic approaches will be used to work out the sequence of events that assemble and disassemble tethering complexes and to identify and characterise those GTPases and kinases that regulate this process. 4. To determine whether a template is needed for the reassembly of the Golgi at the end of mitosis and during biogenesis by studying cytoplasts that lack a Golgi apparatus.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
Project #
Application #
Study Section
Cell Development and Function Integrated Review Group (CDF)
Program Officer
Shapiro, Bert I
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Yale University
Anatomy/Cell Biology
Schools of Medicine
New Haven
United States
Zip Code
Franklin, Joseph B; Ullu, Elisabetta (2010) Biochemical analysis of PIFTC3, the Trypanosoma brucei orthologue of nematode DYF-13, reveals interactions with established and putative intraflagellar transport components. Mol Microbiol 78:173-86
de Graffenried, Christopher L; Ho, Helen H; Warren, Graham (2008) Polo-like kinase is required for Golgi and bilobe biogenesis in Trypanosoma brucei. J Cell Biol 181:431-8
Ang, Agnes Lee; Taguchi, Tomohiko; Francis, Stephen et al. (2004) Recycling endosomes can serve as intermediates during transport from the Golgi to the plasma membrane of MDCK cells. J Cell Biol 167:531-43
Shorter, James; Beard, Matthew B; Seemann, Joachim et al. (2002) Sequential tethering of Golgins and catalysis of SNAREpin assembly by the vesicle-tethering protein p115. J Cell Biol 157:45-62
Jokitalo, E; Cabrera-Poch, N; Warren, G et al. (2001) Golgi clusters and vesicles mediate mitotic inheritance independently of the endoplasmic reticulum. J Cell Biol 154:317-30
Pelletier, L; Jokitalo, E; Warren, G (2000) The effect of Golgi depletion on exocytic transport. Nat Cell Biol 2:840-6
Dirac-Svejstrup, A B; Shorter, J; Waters, M G et al. (2000) Phosphorylation of the vesicle-tethering protein p115 by a casein kinase II-like enzyme is required for Golgi reassembly from isolated mitotic fragments. J Cell Biol 150:475-88