Cyclophilins are a family of proteins that bind cyclosporin A (CsA) and possess peptidyl-prolyl cis-trans isomerase activity. In addition, they are secreted by activated cells and act in a cytokine-like manner, presumably via signaling through a cell surface cyclophilin receptor. More recently, host-derived cyclophilin A (CyPA) has been shown to be incorporated into HIV-1 virions and its incorporation essential for viral infectivity. Here we present evidence supporting a role for viral-associated CyPA in the early events of HIV-1 infection. We report that HIV-1 infection of primary peripheral blood mononuclear cells can be inhibited by: (i) an excess of exogenously added CyPA; (ii) a CsA analogue unable to enter the cells; (iii) neutralizing antibodies to CyPA. Taken together with our observations that recombinant CyPA-induced mobilization of calcium in immortalized, as well as primary, CD4+ T lymphocytes, and that incubation of T cells with iodinated CyPA, followed by chemical cross-linking, resulted in the formation of a high molecular mass complex on the cell surface, these results suggest that HIV-1-associated CyPA mediates an early event in viral infection via interaction with a cellular receptor. This interaction may present a target for anti-HIV therapies and vaccines.