HTLV-1 Tax oncoprotein exerts pleiotropic effects on cellular regulatory systems, trans-activation and trans-repression of transcription and promotion of the cell cycle, through interaction with various cellular factors. Here, we report Tax-mediated inhibition of DNA topoisomerase I. The interaction of Tax with topoisomerase I was demonstrated in vitro and also in vivo, and inhibition of DNA relaxation activity of topoisomerase I was clearly shown. These results suggested that Tax has a novel potential to affect various cellular processes, such as transcription and maintenance of genomic stability, in which DNA topoisomerase I is involved.