The results of yeast and mammalian two-hybrid assays previously indicated complex formation between prenylated Rab acceptor 1 (PRA1) and the cytoplasmic domain of gp41 (gp41CD) for both the human and simian immunodeficiency viruses [Evans, D. T., Tilman, K. C. & Desrosiers, R. C. (2002). J Virol 76, 327-337]. The assembly and release of infectious virus particles was studied under conditions of PRA1 overexpression in a transient transfection assay or suppression by RNA interference. Although a clear pattern of co-localization of PRA1 and gp41 was observed, no changes in virion release, infectivity or envelope content were observed as a result of either PRA1 suppression or overexpression. These data show that PRA1 co-localizes with gp41 inside cells and they are consistent with a direct or indirect interaction between these proteins. However, variation in the levels of PRA1 expression did not influence virion production, infectivity or envelope incorporation under the conditions of these assays.