Cell-Free Analysis of ER to Golgi Apparatus Vesicular Traffic

Abstract

Membrane traffic between the endoplasmic reticulum (ER) and the Golgi apparatus has been analyzed using a cell-free system in which transfer is via 50 to 70 nm transition vesicle intermediates. The latter have been isolated and characterized. Vesicle formation is temperature-and ATP-dependent and inhibited by N-ethylmaleimide. Transfer to Golgi apparatus is specific for cis Golgi apparatus membranes as acceptor. Heterologous transfer has been demonstrated among animal and plant systems. Nuclei with intact or semi-intact membranes will replace ER as the donor. The ability to form transition vesicles is restricted to a subpopulation of the ER, called the transitional or cis Golgi apparatus-associated ER. The cell-free systems employ a fraction enriched in this ER subpopulation. Both lipids and proteins are transferred. Transferred proteins and lipids are processed at the Golgi apparatus suggestive of membrane fusion of the transition vesicle donor with membranes of the Golgi apparatus acceptors. An ATPase with unique properties has been purified from transitional ER of liver. Other proteins under study and potentially involved in the transfer process include a 55 kDa GTP-binding protein of the transitional ER and a putative docking protein of the cis Golgi.