The Actin Cytoskeleton Is Required for Receptor-mediated Endocytosis in Mammalian Cells*
- From the Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037 and the ¶Department of Physiology, University of Texas Southwestern Medical Center, Dallas, Texas 75235
Abstract
Actin filament organization is essential for endocytosis in yeast. In contrast, the actin-depolymerizing agent cytochalasin D has yielded ambiguous results as to a role for actin in receptor-mediated endocytosis in mammalian cells. We have therefore re-examined this issue using highly specific reagents known to sequester actin monomers. Two of these reagents, thymosin β4 and DNase I, potently inhibited the sequestration of transferrin receptors into coated pits as measured in a cell-free system using perforated A431 cells. At low concentrations, thymosin β4 but not DNase I was stimulatory. Importantly, the effects of both reagents were specifically neutralized by the addition of actin monomers. A role for the actin cytoskeleton was also detected in intact cells where latrunculin A, a drug that sequesters actin monomers, inhibited receptor-mediated endocytosis. Biochemical and morphological analyses suggest that these reagents inhibit later events in coated vesicle budding. These results provide new evidence that the actin cytoskeleton is required for receptor-mediated endocytosis in mammalian cells.
Footnotes
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↵* This work was supported by National Institutes of Health Grants GM51112 (to H. L. Y.) and GM42455 and CA58689 (to S. L. S.). This is The Scripps Research Institute Manuscript No. 10577-CB.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‡ Both authors contributed equally to this work.
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↵§ Supported by U. S. Army Medical Research and Material Command Grant DAM17-94-J-4031. Present address: Institut Pasteur, UnitéBiologie des Interactions Cellulaires, 25 rue du Dr Roux, 75015 Paris cédex 15, France.
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↵‖ Established Investigator of the American Heart Association. To whom correspondence should be addressed. Tel.: 619-784-2311; Fax: 619-784-9126; E-mail: slschmid{at}scripps.edu.
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↵1 The abbreviations used are: B-Tfn, biotinylated human diferric transferrin; TfnR, transferrin receptors; Tβ4, thymosin β4; MesNa, β-mercaptoethanesulfonic acid.
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- Received June 13, 1997.
