Actin-binding protein, supervillin, has been identified as an androgen receptor (AR) coregulator. Although actin has been suggested to participate in transcription regulation, the mechanism is not clear. Here we demonstrate signals involved in the cytoskeleton dynamic can modulate the coregulator function of supervillin. Three actin isoforms cooperate with supervillin in additive manner to further enhance AR transactivation. Latrunculin B toxin, an actin chelator, reduces the availability of monomer actin and attenuates supervillin function. Rac, the small G-protein kinase, is well studied in reorganization of cytoskeleton. The overexpression of constitutive-active Rac triggers the membrane ruffling site and reduces the coregulator activity of supervillin. Together, the availability of actin monomer affects supervillin-modulated AR transactivation.