Reversal of Fortune

The proper targeting and fusion of transport vesicles with the correct membrane is a critical event in the determination of the identity of different compartments within the cell. Work over the last decade has made tremendous progress toward determining a general mechanism by which this occurs. The cornerstones of such a mechanism will have to include two families of proteins: Rab GTPases and SNARE proteins. SNARE proteins are thought to have a central role in catalyzing the fusion of the vesicle with the target membrane (Weber et al., 1998), while Rab GTPases appear to work upstream of this in mediating the initial docking or tethering of the vesicle to the target membrane (Cao et al., 1998; Waters and Pfeffer, 1999). To understand the mechanism by which these two classes of proteins collaborate in this process, it is important to know the arrangements of these proteins with respect to the ves-icle and target membrane. Work over the last decade has given us insights as to the membrane surfaces with which these proteins are localized. It is from this localization that the terms vesicle or v-SNARE and target membrane or t-SNARE have their origin. Likewise Rab proteins, such as Sec4, Ypt1, and Rab3 have all been found associated with transport vesicles: Sec4 on post-Golgi vesicles (Goud et al. Other than these clues given by their presence on vesicle or target membrane compartments, no direct evidence of their site of action was known. This is especially important considering that in many cases the SNARE and Rab proteins are present at significant levels on both the target and vesicle membranes. The article by Cao and Barlowe in this issue, provides the first comprehensive test of the site of action of SNAREs and Rab proteins in the fusion of transport vesicles with a target membrane (Cao and Barlowe, 2000). The results are quite surprising. The authors make use of a two-stage in vitro system for examining the docking and fusion of ER-derived vesicles with the Golgi apparatus. In this assay vesicles are produced by incubation of donor membrane containing 35 S-labeled cargo with purified COPII coat subunits to initiate the production of ER-to-Golgi vesicles. These vesicles, containing radiolabeled cargo, can then be readily separated from the donor membrane by centrifugation. The second stage of the assay is carried out by incubating the isolated vesicles with an acceptor compartment containing the target Golgi membranes. Fusion is …