Mammalian actin binding protein 1 (mAbp1, also called SH3P7/Hip55) is structurally and functionally related to yeast Abp1 and to cortactin, both of which have been implicated in endocytotic processes. mAbp1 associates through its SH3 domain with dynamin, a large GTPase essential for vesicle fission. To clarify the function of mAbp1, we specifically knocked down its expression in human embryonic kidney 293T cells, using RNA interference (RNAi). Co-transfection of a short interfering RNA (siRNA) together with a plasmid coding for a surface marker, followed by purification of transfected cells, enabled us to obtain a cell population having up to 90% inhibition of mAbp1 expression. In mAbp1-knocked down cells, transferrin (Tf) receptor endocytosis was significantly inhibited and intracellular distribution of the early endosomal compartment was modified. In contrast, in these cells actin and microtubule filaments appeared normal, and formation of lamellipodia induced by active Rac was not inhibited. This study provides definitive evidence that mAbp1 is indispensable for receptor-mediated endocytosis. (C) 2003 Elsevier Science (USA). All rights reserved.