Interaction of phosphoinositide cycle intermediates with the plasma membrane-associated clathrin assembly protein AP-2

Kenneth A Beck, James H. Keen

Research output: Contribution to journalArticle

105 Citations (Scopus)

Abstract

Several components of the phosphoinositide cycle have been found to interact specifically and at physiological concentrations with the plasma membrane-associated clathrin assembly (adaptor) protein AP-2. These include phosphatidylinositol 4,5-bisphosphate and inositol 1,4,5-trisphosphate, which are present at the plasma membrane, as well as other polyphosphoinositols. ATP and other polyphosphate molecules compete with the polyphosphoinositols, however, they are at least 80-fold less potent. Also, the effect of ATP, unlike the polyphosphoinositols, is blocked by physiological concentrations of Mg2+. Photoaffinity labeling of AP-2 by [α-32P]8-azidoadenosine 5′-triphosphate and its competition by polyphosphoinositols has been used to identify the α subunit of the AP-2 complex as the site of specific interaction with the polyphosphoinositols and to confirm direct ultrafiltration binding experiments. Proteolytic dissection of the labeled AP-2 demonstrated that binding occurred exclusively on the N-terminal portion of the α subunit. Interaction of purified AP-2 with sub-μM concentrations of polyphosphoinositols has inhibitory effects on a novel AP-2 self-association described in the accompanying paper (Beck, K. A., and Keen, J. H., J. Biol. Chem. 266, 4437-4441), and at higher concentrations on the binding of AP-2 to dissociated clathrin trimers as well as AP-2-mediated clathrin coat assembly. Review of the literature shows that several physiological stimuli that are known to result in increased coat pit formation in intact cells correlate with increased phosphoinositide turnover. These in vivo correlations and the in vitro observations reported here suggest that coated membrane and phosphoinositide cycles may be interdependent within cells.

Original languageEnglish (US)
Pages (from-to)4442-4447
Number of pages6
JournalJournal of Biological Chemistry
Volume266
Issue number7
StatePublished - Mar 5 1991
Externally publishedYes

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Vesicular Transport Adaptor Proteins
Cell membranes
Phosphatidylinositols
Clathrin
Membrane Proteins
Cell Membrane
Adenosine Triphosphate
Dissection
Polyphosphates
Inositol 1,4,5-Trisphosphate
Ultrafiltration
Labeling
Membranes
Molecules
Proteins
Experiments

ASJC Scopus subject areas

  • Biochemistry

Cite this

Interaction of phosphoinositide cycle intermediates with the plasma membrane-associated clathrin assembly protein AP-2. / Beck, Kenneth A; Keen, James H.

In: Journal of Biological Chemistry, Vol. 266, No. 7, 05.03.1991, p. 4442-4447.

Research output: Contribution to journalArticle

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