Transcription initiation at RNA polymerase III promoters requires transcription factor IIIB (TFIIIB), an activity that binds to RNA polymerase III promoters, generally through protein-protein contacts with DNA binding factors, and directly recruits RNA polymerase III. Saccharomyces cerevisiae TFIIIB is a complex of three subunits, TBP, the TFIIB-related factor BRF, and the more loosely associated polypeptide beta("). Although human homologs for two of the TFIIIB subunits, the TATA box-binding protein TBP and the TFIIB-related factor BRF, have been characterized, a human homolog of yeast B(") has not been described. Moreover, human BRF, unlike yeast BRF, is not universally required for RNA polymerase III transcription. In particular, it is not involved in transcription from the small nuclear RNA (snRNA)-type, TATA-containing, RNA polymerase III promoters. Here, we characterize two novel activities, a human homolog of yeast B("), which is required for transcription of both TATA-less and snRNA-type RNA polymerase III promoters, and a factor equally related to human BRF and TFIIB, designated BRFU, which is specifically required for transcription of snRNA-type RNA polymerase III promoters. Together, these results contribute to the definition of the basal RNA polymerase III transcription machinery and show that two types of TFIIIB activities, with specificities for different classes of RNA polymerase III promoters, have evolved in human cells.