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Binding partners for the COOH-terminal appendage domains of the GGAs and gamma-adaptin.

The adaptor appendage domains are believed to act as binding platforms for coated vesicle accessory proteins. Using glutathione S-transferase pulldowns from pig brain cytosol, we find three proteins that can bind to the appendage domains of both the AP-1 gamma subunit and the GGAs: gamma-synergin and two novel proteins, p56 and p200. p56 elicited better antibodies than p200 and was generally more tractable. Although p56 and gamma-synergin bind to both GGA and gamma appendages in vitro, immunofluorescence labeling of nocodazole-treated cells shows that p56 colocalizes with GGAs on TGN46-positive membranes, whereas gamma-synergin colocalizes with AP-1 primarily on a different membrane compartment. Furthermore, in AP-1-deficient cells, p56 remains membrane-associated whereas gamma-synergin becomes cytosolic. Thus, p56 and gamma-synergin show very strong preferences for GGAs and AP-1, respectively, in vivo. However, the GGA and gamma appendages share the same fold as determined by x-ray crystallography, and mutagenesis reveals that the same amino acids contribute to their binding sites. By overexpressing wild-type GGA and gamma appendage domains in cells, we can drive p56 and gamma-synergin, respectively, into the cytosol, suggesting a possible mechanism for selectively disrupting the two pathways.

Pubmed ID: 12808037


  • Lui WW
  • Collins BM
  • Hirst J
  • Motley A
  • Millar C
  • Schu P
  • Owen DJ
  • Robinson MS


Molecular biology of the cell

Publication Data

June 16, 2003

Associated Grants


Mesh Terms

  • ADP-Ribosylation Factors
  • Adaptor Protein Complex 1
  • Adaptor Protein Complex gamma Subunits
  • Adaptor Proteins, Vesicular Transport
  • Amino Acid Sequence
  • Animals
  • COS Cells
  • Carrier Proteins
  • Genes, Reporter
  • Molecular Sequence Data
  • Mutation
  • Protein Structure, Tertiary