The γ/σ1 and α/σ2 hemicomplexes of clathrin adaptors AP-1 and AP-2 Harbor the dileucine recognition site

Balraj Doray, Intaek Lee, Jane Knisely, Guojun Bu, Stuart Kornfeld

Research output: Contribution to journalArticlepeer-review

120 Scopus citations

Abstract

The clathrin adaptors AP-1 and AP-2 bind cargo proteins via two types of motifs: tyrosine-based Yxxφ and dileucine-based [DE]XXXL[LI]. Although it is well established that Yxxφ motifs bind to the μ subunits of AP-1 or AP-2, dileucine motifs have been reported to bind to either the μ or β subunits of these adaptors as well as the γ/σ1 hemicomplex of AP-1. To clarify this controversy, the various subunits of AP-1 and AP-2 were expressed individually and in hemicomplex form in insect cells, and they were used in glutathione S-transferase pull-down assays to determine their binding properties. We report that the γ/σ1 or α/σ2 hemicomplexes bound the dileucine-based motifs of several proteins quite strongly, whereas binding by the β1/μ1 and β2/μ2 hemicomplexes, and the individual β or subunits, was extremely weak or undetectable. The σ1 and olα hemicomplexes displayed substantial differences in their preference for particular dileucine-based motifs. Most strikingly, an aspartate at position -4 compromised binding to the γ/σ1 hemicomplex, whereas minimally affecting binding to α/σ2. There was an excellent correlation between binding to the α/σ2 hemicomplex and in vivo internalization mediated by the dileucine-based sorting signals. These findings provide new insights into the trafficking mechanisms of D/EXXXL[LI]-mediated sorting signals.

Original languageEnglish (US)
Pages (from-to)1887-1896
Number of pages10
JournalMolecular biology of the cell
Volume18
Issue number5
DOIs
StatePublished - May 2007

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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