Cyclophilin A (Cyp A) binds the human immunodeficiency virus type 1 (HIV-1) capsid (CA) protein and contributes to the early events in virus replication in some cells. The retroviral restriction factor TRIM5alpha can inhibit the early, post-entry phase of infection by associating with the incoming viral capsid. Cyp A has been proposed to prevent restriction factor binding in human cells, thus enhancing HIV-1 infectivity, and to potentiate restriction of HIV-1 in monkey cells. Here we show that the positive effects of Cyp A-CA binding on HIV-1 infectivity do not depend on human TRIM5alpha. Disruption of Cyp A binding to CA partially relieved the block to HIV-1 infection imposed by several TRIM5alpha variants, but Cyp A-CA binding was not absolutely required for TRIM5alpha antiviral activity. Inhibition of Cyp A function by cyclosporine significantly decreased the efficiency of TRIM5alpha-mediated restriction only when the restricted virus capsid interacted with Cyp A.