Targeting signals and subunit interactions in coated vesicle adaptor complexes

Abstract

There are two clathrin-coated vesicle adaptor complexes in the cell, one associated with the plasma membrane and one associated with the TGN. The subunit composition of the plasma membrane adaptor complex is α-adaptin, β- adaptin, AP50, and AP17; while that of the TGN adaptor complex is γ-adaptin, β'-adaptin, AP47, and AP19. To search for adaptor targeting signals, we have constructed chimeras between α-adaptin and γ-adaptin within their NH2- terminal domains. We have identified stretches of sequence in the two proteins between amino acids ~130 and 330-350 that are essential for targeting. Immunoprecipitation reveals that this region determines whether a construct coassembles with AP50 and AP17, or with AP47 and AP19. These observations suggest that these other subunits may play an important role in targeting. In contrast, β- and β'-adaptins are clearly not involved in this event. Chimeras between the α- and γ-adaptin COOH-terminal domains reveal the presence of a second targeting signal. We have further investigated the interactions between the adaptor subunits using the yeast two-hybrid system. Interactions can be detected between the β/β'-adaptins and the α/γ- adaptins, between the β/β'-adaptins and the AP50/AP47 subunits, between α- adaptin and AP17, and between γ-adaptin and AP19. These results indicate that the adaptor subunits act in concert to target the complex to the appropriate membrane.