Peden, A. A. orcid.org/0000-0003-0144-7712, Rudge, R. E., Lui, W. W. et al. (1 more author) (2002) Assembly and function of AP-3 complexes in cells expressing mutant subunits. Journal of Cell Biology, 156 (2). pp. 327-336. ISSN 0021-9525
Abstract
The mouse mutants mocha and pearl are deficient in the AP-3 delta and beta3A subunits, respectively. We have used cells from these mice to investigate both the assembly of AP-3 complexes and AP-3 function. In mocha cells, the beta3 and mu3 subunits coassemble into a heterodimer, whereas the sigma3 subunit remains monomeric. In pearl cells, the delta and sigma3 subunits coassemble into a heterodimer, whereas mu3 gets destroyed. The yeast two hybrid system was used to confirm these interactions, and also to demonstrate that the A (ubiquitous) and B (neuronal-specific) isoforms of beta3 and mu3 can interact with each other. Pearl cell lines were generated that express beta3A, beta3B, a beta3Abeta2 chimera, two beta3A deletion mutants, and a beta3A point mutant lacking a functional clathrin binding site. All six constructs assembled into complexes and were recruited onto membranes. However, only beta3A, beta3B, and the point mutant gave full functional rescue, as assayed by LAMP-1 sorting. The beta3Abeta2 chimera and the beta3A short deletion mutant gave partial functional rescue, whereas the beta3A truncation mutant gave no functional rescue. These results indicate that the hinge and/or ear domains of beta3 are important for function, but the clathrin binding site is not needed.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2002 Rockefeller University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial Share-Alike Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. You may not use the material for commercial purposes. |
Keywords: | Adaptor Proteins, Vesicular Transport; Animals; Antigens, CD; Binding Sites; Blotting, Western; Carrier Proteins; Cell Line; Clathrin; Flow Cytometry; Lysosome-Associated Membrane Glycoproteins; Macromolecular Substances; Membrane Glycoproteins; Membrane Proteins; Mice; Mice, Mutant Strains; Microscopy, Fluorescence; Monomeric Clathrin Assembly Proteins; Mutation; Phenotype; Protein Binding; Protein Structure, Quaternary; Protein Structure, Tertiary; Protein Subunits; Proteins; Recombinant Fusion Proteins; Two-Hybrid System Techniques |
Dates: |
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Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Science (Sheffield) > School of Biosciences (Sheffield) > Department of Biomedical Science (Sheffield) |
Depositing User: | Symplectic Sheffield |
Date Deposited: | 21 Dec 2016 15:34 |
Last Modified: | 21 Dec 2016 15:40 |
Published Version: | http://dx.doi.org/10.1083/jcb.200107140 |
Status: | Published |
Publisher: | Rockefeller University Press |
Refereed: | Yes |
Identification Number: | 10.1083/jcb.200107140 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:108293 |