Two WXXF-based motifs in NECAPs define the specificity of accessory protein binding to AP-1 and AP-2

Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montreal, QC, Canada.
The EMBO Journal (Impact Factor: 10.43). 11/2004; 23(19):3701-10. DOI: 10.1038/sj.emboj.7600378
Source: PubMed


The adaptor proteins AP-2 and AP-1/GGAs are essential components of clathrin coats at the plasma membrane and trans-Golgi network, respectively. The adaptors recruit accessory proteins to clathrin-coated pits, which is dependent on the adaptor ear domains engaging short peptide motifs in the accessory proteins. Here, we perform an extensive mutational analysis of a novel WXXF-based motif that functions to mediate the binding of an array of accessory proteins to the alpha-adaptin ear domain of AP-2. Using nuclear magnetic resonance and mutational studies, we identified WXXF-based motifs as major ligands for a site on the alpha-ear previously shown to bind the DPW-bearing proteins epsin 1/2. We also defined the determinants that allow for specific binding of the alpha-ear motif to AP-2 as compared to those that allow a highly related WXXF-based motif to bind to the ear domains of AP-1/GGAs. Intriguingly, placement of acidic residues around the WXXF cores is critical for binding specificity. These studies provide a structural basis for the specific recruitment of accessory proteins to appropriate sites of clathrin-coated vesicle formation.

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Available from: Brigitte Ritter, Sep 17, 2014
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    • "These sorting motifs are recognized by a member of the family of clathrin-associated sorting adaptors (CLASPs). The cytosolic tail of Kremen1 does not have typical tyrosine- or dileucine-based sorting motifs, although we did identify a WXXF sequence that might bind to the α-appendage of AP-2 as previously reported for many CLASPs [36], [37]. Mutating either tryptophan to alanine (data not shown) or phenylalanine to alanine of WWXF (Fig. S1A) did not interfere with the trafficking of Kremen1, indicating that other trafficking motifs must be present in the cytosolic tail of Kremen1. "
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    • "Blue circles indicate NPF motifs. Three motifs, shown as triangles, interact with the α-ear domains of the AP2 complex: red triangles indicate WxxF motifs [29], [30], orange triangles indicate DPF motifs [28], and the green triangle indicates an FxDxF motif [28]. The pink diamonds indicate C-terminal PDZ domain-binding motifs [31]. "
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