Spatiotemporal dynamics of COPII vesicle formation and morphogenesis of exit sites in the endoplasmic reticulum

• Applicant: Professor Dr. Matthias Weiss
• Subject Area: Biophysics
• Term: from 2008 to 2012
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 68851344

The formation of transport vesicles at the endoplasmic reticulum (ER) is a generic phenomenon in eucaryotic cells. Due to the transient, sequential, and energy-dependent binding of peripheral membrane proteins (COPII) and the subsequent assembly of COPII complexes the ER membrane is deformed and a cargo-loaded bud/vesicle emerges. As of yet, the spatiotemporal aspects underlying the assembly of COPII-coated buds are as poorly explored as the concomitant emergence of ER exit sites (ERES) of which ~100 copies are typically observed in mammalian cells. The project aims at elucidating the dynamic aspects of COPII vesicle formation and ERES morphogenesis. To this end, fluorescently tagged proteins/lipids will be monitored with advanced light microscopy (e.g. fluorescence correlation spectroscopy) on the (almost) single molecule level in vitro and in vivo. Here, we will use giant unilamellar vesicles of dedicated lipid composition and purified COPII proteins; we will compare our insights to observations in transiently/stably transfected cell lines. Special attention will be given to the scaffolding role of lipids and the influence of non-equilibrium events. The obtained quantitative data on the binding kinetics of COPII proteins, the diffusion and interactions of lipids/proteins under various conditions, and the overall change in membrane morphology will be used to formulate and computationally implement a predictive mesoscopic model that unifies the experimental findings and yields a deeper understanding of the spatiotemporal aspects of COPII vesicle and ERES formation.

• DFG Programme: Research Grants