Characterisation of Glycosylation Mutants in Drosophila melanogaster

Glycosylation is a fundamental process in cellular life, conferring structural and functional properties to proteins and lipids. In order for glycosylation to take place properly, a series of carefully regulated steps must be carried out in specific organelles of the cell. The Golgi apparatus is one of the key organelles where this post-translational modification is carefully controlled by a series of steps in a process known as vesicle trafficking. This multi-layered mechanism has many key phases by which vesicles loaded with cargo are transported from one membrane compartment to another. Glycosylation enzymes make up this cargo and the distribution of these proteins throughout the cisternal compartments of the Golgi is crucial for proper glycosylation to take place. One of the steps of vesicle trafficking is tethering, a process regulated by various proteins including the Conserved Oligomeric Golgi (COG) complex. Defects in subunits of the COG complex leads to perturbations in glycosylation which consequently can result in alterations of cellular functional homeostasis. In this project we utilise Drosophila melanogaster (Dm) as a model organism to try and understand more about the role of COG in vesicle trafficking and glycosylation both in vitro and in vivo. To achieve these aims, using COG mutants, we have carried out N- and O-linked glycan profiling and flight test analysis in an effort to connect COG defects in vivo to altered glycan patterns in vitro. Also using a yeast-two-hybrid (Y2H) approach we looked at COG-Rab interactions to observe how well conserved mammalian-invertebrate trafficking interactions are. What we found are not only functional implications of COG in determining Drosophila glycan synthesis and flight ability, but also evidence of evolutionary conservation during interactions associated with membrane trafficking events.