* Department of Physiology, University College London, Gower Street, London WC1E 6BT, UK‡ Department of Pathology, Centre Medical Universitaire, Rue Michel-Servet 1, CH-1211 Geneva, Switzerland § Max Planck Institute für Medizinische Forschung, Jahnstrasse 29, 69120 Heidelberg, Germany† Present address: Vollum Institute, 3181 Sam Jackson Park Road, Portland, Oregon 97201-3098, USA¶ e-mail: almersw@ ohsu. edu ctin polymerization is thought to provide the motive force for crawling cells by driving the spread of lamellipodia1. It may also be involved when membrane ruffles engulf external fluid during macropinocytosis2. Here we show that, in cells transfected with a fusion protein consisting of green fluorescent protein (GFP) and β-actin, pinosomes (vesicles used for fluid uptake) ignite a burst of actin polymerization when they are pinched off from the plasma membrane. Pinosomes then move into the cytosol at the tips of short-lived actin ‘comet tails’ that are similar to those that propel Listeria3 and other microorganisms4, 5 through infected cells. Like Listeria, pinosomes appear to carry the machinery required for propulsive actin polymerization. The direction of pinosome movement indicates that they may acquire this machinery from the crests of membrane ruffles. We suggest that actin polymerization moves the leading edge of ruffles. Endocytic vesicles may also use actin polymerization to move into the cytosol after being pinched off from the plasma membrane.

We transfected rat basophilic leukaemia (RBL) cells with GFP-labelled β-actin6 and allowed them to ruffle and undergo macropinocytosis in a mildly hyperosmolar medium7. Ruffles faintly aglow with GFP–actin engulfed fluid and formed vesicular structures near the plasma membrane (not shown). The structures fluoresced (Fig. 1a, arrowhead) as they ignited a burst of actin polymerization that caused a streak of actin to move through the cell, much like actin ‘comet tails’. The tips of the actin tails were cup-shaped as if to accommodate vesicles, and appeared to push spherical objects that were visible as dark shadows. Large tails sometimes appeared hollow (not shown).