Human foamy virus encodes a nuclear regulatory protein, termed Bel-1, that serves as a potent activator of viral transcription. Mutational analysis has identified a small, discrete activation domain within Bel-1 that is highly active in both higher and lower eukaryotic cells. Here, we demonstrate that the activation domain of Bel-1 is highly dependent on the ADA2 transcriptional adaptor for biological activity in yeast cells, a property previously shown to be a hallmark of the VP16 class of acidic transcriptional activators (S. L. Berger, B. Pina, N. Silverman, G. A. Marcus, J. Agapite, J. L. Regier, S. J. Triezenberg, and L. Guarente, Cell 70:251-265, 1992). Using genetic selection in yeast cells, we have derived a set of point mutants within the Bel-1 activation domain that display a qualitatively similar loss in activation potential when examined in either yeast or human cells. These data indicate that the Bel-1 activation domain functions similarly in both lower and higher eukaryotes and strongly suggest that Bel-1 belongs to the VP16 class of acidic transcription factors.