The trans-acting factor of human immunodeficiency virus (HIV), Tat, has a basic amino-acid cluster that is highly conserved among different HIV isolates. We have examined the effects of mutations in the basic region of Tat on its trans-acting activity and cellular localization. Introduction of a stop codon immediately preceding the basic region abolished the activity, while the truncated mutant with the basic region retained some activity. The basic region of Tat was replaceable with that of Rev (another trans-acting factor of HIV) but not with that of adenovirus Ela nor cellular enzyme. The result of immunofluorescence analysis revealed a correlation between the nuclear, especially nucleolar, accumulation and the activities of mutant Tat proteins.