A role for a wortmannin-sensitive phosphatidylinositol-4-kinase in the endocytosis of muscarinic cholinergic receptors.

Abstract

A role for phosphoinositides in the endocytosis of muscarinic cholinergic receptors (mAChRs) has been investigated via inhibition of the activity of phosphatidylinositol-4-kinase (PI4K). Pretreatment of SH-SY5Y neuroblastoma cells with micromolar concentrations of wortmannin (WT), LY-294002, or phenylarsine oxide (PAO), three chemically distinct agents known to inhibit PI4K, resulted in both an inhibition of agonist-induced endocytosis of mAChRs and a selective reduction in the 32P-labeling of phosphatidylinositol-4-phosphate. PAO-mediated inhibition of both receptor endocytosis and phosphoinositide synthesis could be fully reversed by inclusion of the bifunctional thiol 2, 3-dimercaptopropanol. The requirement for phosphoinositide synthesis in mAChR endocytosis was independent of a role for these lipids in the maintenance of the cytoskeleton because disruption of the latter with cytochalasin D, ML-7, or colchicine failed to inhibit receptor internalization. Determination of PI4K activity in subcellular fractions of SH-SY5Y cells indicated that enzyme activity in fractions enriched in endocytic vesicles and cytosol was preferentially inhibited by WT, LY-294002, and PAO, a profile consistent with the subcellular distribution of the 110-kDa beta isoform of PI4K, as determined by Western blot analysis. Activity of PI4Kbeta present in immunoprecipitated cell lysates was inhibited >75% by inclusion of each of the three inhibitors. These results indicate that ongoing synthesis of phosphoinositides is necessary for mAChR endocytosis and that the activity of a WT-sensitive form of PI4K, such as PI4Kbeta, is required.

13 Figures and Tables

Statistics

050'00'02'04'06'08'10'12'14'16
Citations per Year

195 Citations

Semantic Scholar estimates that this publication has 195 citations based on the available data.

See our FAQ for additional information.

Cite this paper

@article{Sorensen1998ARF, title={A role for a wortmannin-sensitive phosphatidylinositol-4-kinase in the endocytosis of muscarinic cholinergic receptors.}, author={Scott D Sorensen and Daniel A Linseman and Edward L. McEwen and Anne M. Heacock and Stephen K . Fisher}, journal={Molecular pharmacology}, year={1998}, volume={53 5}, pages={827-36} }