Functional domain mapping of the clathrin-associated adaptor medium chains mu1 and mu2.
The clathrin-associated adaptors AP-1 and AP-2 are heterotetrameric complexes involved in the recognition of sorting signals present within the cytosolic domain of integral membrane proteins. The medium chains of these complexes, mu1 and mu2, have been implicated in two types of interaction: assembly with the beta1 and beta2 chains of the corresponding complexes and recognition of tyrosine-based sorting signals. In this study, we report the results of a structure-function analysis of the mu1 and mu2 chains aimed at identifying regions of the molecules that are responsible for each of the two interactions. Analyses using the yeast two-hybrid system and proteolytic digestion experiments suggest that mu1 and mu2 have a bipartite structure, with the amino-terminal one-third (residues 1-145 of mu1 and mu2) being involved in assembly with the beta chains and the carboxyl-terminal two-thirds (residues 147-423 of mu1 and 164-435 of mu2) binding tyrosine-based sorting signals. These observations support a model in which the amino-terminal one-third of mu2 is embedded within the core of the AP-2 complex, while the carboxyl-terminal two-thirds of the protein are exposed to the medium, placing this region in a position to interact with tyrosine-based sorting signals.
Aguilar
R C
RC
Cell Biology and Metabolism Branch, NICHD, National Institutes of Health, Bethesda, Maryland 20892-5430, USA.
Ohno
H
H
Roche
K W
KW
Bonifacino
J S
JS
D033961:Adaptor Protein Complex 1; D033962:Adaptor Protein Complex 2; D033963:Adaptor Protein Complex 3; D033983:Adaptor Protein Complex mu Subunits; D033942:Adaptor Proteins, Vesicular Transport; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000906:Antibodies; D002966:Clathrin / Q000378:metabolism*; D019976:Cloning, Organism; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000096:biosynthesis; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism*; D010750:Phosphoproteins / Q000737:chemistry* / Q000378:metabolism*; D055503:Protein Multimerization; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion; D014157:Transcription Factors; D014158:Transcription, Genetic; D020798:Two-Hybrid System Techniques; D014443:Tyrosine; D001616:beta-Galactosidase / Q000096:biosynthesis
C438890:APM1 protein, S cerevisiae; D033961:Adaptor Protein Complex 1; D033962:Adaptor Protein Complex 2; D033963:Adaptor Protein Complex 3; D033983:Adaptor Protein Complex mu Subunits; D033942:Adaptor Proteins, Vesicular Transport; D000906:Antibodies; D002966:Clathrin; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D046911:Macromolecular Substances; D009419:Nerve Tissue Proteins; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; C499066:adaptor protein complex 1, mu 1 subunit; C499065:adaptor protein complex 1, mu 2 subunit; C438885:adaptor protein complex 2, mu 1 subunit; C438886:adaptor protein complex 2, mu 2 subunit; D014443:Tyrosine; D001616:beta-Galactosidase