Targeting signals and subunit interactions in coated vesicle adaptor complexes.
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 alpha-adaptin, beta-adaptin, AP50, and AP17; while that of the TGN adaptor complex is gamma-adaptin, beta'-adaptin, AP47, and AP19. To search for adaptor targeting signals, we have constructed chimeras between alpha-adaptin and gamma-adaptin within their NH2-terminal domains. We have identified stretches of sequence in the two proteins between amino acids approximately 130 and 330-350 that are essential for targeting. Immunoprecipitation reveals that this region determines whether a construct coassemblies 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, beta- and beta'-adaptins are clearly not involved in this event. Chimeras between the alpha- and gamma-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 beta/beta'-adaptins and the alpha/gamma-adaptins, between the beta/beta'-adaptins and the AP50/AP47 subunits, between alpha-adaptin and AP17, and between gamma-adaptin and AP19. These results indicate that the adaptor subunits act in concert to target the complex to the appropriate membrane.
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L J
LJ
Department of Clinical Biochemistry, University of Cambridge, Addenbrooke's Hospital, England.
Robinson
M S
MS
D033961:Adaptor Protein Complex 1; D033962:Adaptor Protein Complex 2; D033965:Adaptor Protein Complex alpha Subunits; D033966:Adaptor Protein Complex beta Subunits; D033982:Adaptor Protein Complex gamma Subunits; D033983:Adaptor Protein Complex mu Subunits; D034002:Adaptor Protein Complex sigma Subunits; D033942:Adaptor Proteins, Vesicular Transport; D000818:Animals; D002462:Cell Membrane / Q000502:physiology; D018699:Coated Vesicles / Q000737:chemistry* / Q000502:physiology*; D005347:Fibroblasts / Q000166:cytology; D005455:Fluorescent Antibody Technique; D006056:Golgi Apparatus / Q000502:physiology; D008565:Membrane Proteins / Q000032:analysis; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000502:physiology / Q000648:ultrastructure
D033961:Adaptor Protein Complex 1; D033962:Adaptor Protein Complex 2; D033965:Adaptor Protein Complex alpha Subunits; D033966:Adaptor Protein Complex beta Subunits; D033982:Adaptor Protein Complex gamma Subunits; D033983:Adaptor Protein Complex mu Subunits; D034002:Adaptor Protein Complex sigma Subunits; D033942:Adaptor Proteins, Vesicular Transport; C510067:Ap2s1 protein, rat; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; C499071:adaptor protein complex 1, sigma 1 subunit; C438885:adaptor protein complex 2, mu 1 subunit; C438886:adaptor protein complex 2, mu 2 subunit