Organization of glycosylation enzymes in the plant Golgi apparatus

The endomembrane system is an internal membrane system that controls secretion and uptake of biomolecules in all eukaryotic cells. At the heart of the endomembrane system is the Golgi apparatus, which receives cargo from the endoplasmic reticulum and sorts and delivers the cargo to various organelles within a cell. Within the secretory pathway the plant Golgi apparatus is the major biosynthetic organelle harbouring a large number of enzymatic activities for modification of proteins, lipids and for cell wall polysaccharide biosynthesis. Morphologically, the plant Golgi apparatus is a polarized structure consisting of flattened cisternae organized into stacks, which compartmentalize the components for the biosynthesis and trafficking/sorting of proteins, carbohydrates and lipids. Correct function of the Golgi apparatus is therefore essential for plant viability and consequently for the whole ecosystem on earth. Despite this importance little is known about cellular mechanisms that control the organization of the Golgi and its biosynthetic machinery. An unsolved fundamental question of cell biology is how the functional integrity of the Golgi is maintained while facing a constant flow of secretory products. Cargo is transported through the Golgi and modified within different cisternae, while Golgi-resident enzymes have to be retained or retrieved and organized along the different cisternae in order to maintain their biosynthetic function. In this project I will address this exciting question and try to unravel the molecular mechanisms involved in the sub-Golgi distribution of glycosylation enzymes in plants.

A novel molecular mechanism for the organisation of glycosyltransferases within the Golgi apparatus of plants was revealed in this project. The Golgi apparatus is a multifaceted, multitasking organelle that is pivotal to the life of the cell. The compartmentation of biosynthetic activities in different cisternae of a polarized Golgi stack is a major function of the Golgi. The specialized Golgi architecture provides optimal conditions for the asymmetric distribution of enzymes and biosynthetic reactions and the accuracy of these processing steps is achieved by the strict intra-Golgi localization of sequentially acting glycosidases and glycosyltransferases. In this project, we examined the contribution of individual protein regions from the glycosyltransferase GnTI - a key enzyme of the N-glycan processing pathway - to Golgi localization, protein complex formation and the ability to restore N-glycan processing in the model plant Arabidopsis thaliana. Our results show that a conserved amino acid signature within the transmembrane domain of GnTI accounts for its steady-state distribution in a distinct area of the Golgi. Importantly, disturbance of the amino acid signature alters the Golgi localization and subsequently leads to severe defects in N-glycan processing of proteins. Our increased understanding of these fundamental cellular processes is essential for the biotechnological use of plants as manufacturing hosts for valuable products like recombinant biopharmaceuticals.