It has been suggested that the signal transduction initiated by apolipoprotein A-I (apo A-I) activates key proteins involved in cholesterol efflux. ATP binding cassette protein A1 (ABCA1) has been shown to serve as a binding partner for apo A-I, but its participation in apo A-I-induced signaling remains uncertain. We show here that the exposure of human fibroblasts to ABCA1 ligands, such as apolipoproteins and amphipathic helical peptides but not to compounds inducing unspecific cholesterol efflux, such cyclodextrins or liposomes result in the generation of intracellular signals, including activation of small G-protein Cdc42, protein kinases located downstream to Cdc42 (PAK-1, p54JNK), and actin polymerization. The apo A-I-induced signaling was abrogated by glyburide, a nonspecific inhibitor of the ABC transporter family and in fibroblasts from patients with Tangier disease, which do not express functional ABCA1. Conversely, induction of ABCA1 expression by treatment of fibroblasts with combination of liver X receptor (LXR) agonist, T0901317, and retinoid X receptor (RXR) agonist, R0264456, potentiated apo A-I-induced signal transduction. Similar effects were observed in HEK293 cells overexpressing ABCA1-GFP fusion protein but not a mutant form of ABCA1-GFP (K939M) that fails to hydrolyze ATP, or a nonfunctional ABCA1-GFP variant with a truncated C-terminus. We further found that Cdc42 co-immunoprecipitates with ABCA1 in ABCA1-GFP-expressing HEK293 cells exposed to apo A-I but not in cells expressing one of the mutant forms of ABCA1. We conclude that ABCA1 transduces signals from apo A-I by complexing and activating Cdc42 and downstream protein kinases and, therefore, acts as a full apo A-I receptor.