BIOCHEMICAL REQUIREMENTS FOR THE TARGETING AND FUSION OF ER-DERIVED TRANSPORT VESICLES WITH PURIFIED YEAST GOLGI MEMBRANES

In order for secretion to progress, ER-derived transport vesicles must target to, and fuse with the cis-Golgi compartment. These processes h ave been reconstituted using highly enriched membrane fractions and pa rtially purified soluble components. The functionally active yeast Gol gi membranes that have been purified are highly enriched in the cis-Go lgi marker enzymes alpha 1,6 mannosyltransferase and GDP-ase. Fusion o f transport vesicles with these membranes requires both GTP and ATP hy drolysis, and depends on cytosolic and peripheral membrane proteins. A t least two protein fractions from yeast cytosol are required for the reconstitution of ER-derived vesicle fusion. Soluble fractions prepare d from temperature-sensitive mutants revealed requirements for the Ypt 1p, Sec19p, Sly1p, Sec7p, and Uso1 proteins. A model for the sequentia l involvement of these components in the targeting and fusion reaction is proposed.