The TATA-binding protein TBP has been recently recognized as a general class III transcription factor. Using the gel shift assay to monitor initiation complex assembly on a yeast tRNA gene, we show that TBP is required for the TFIIIC-dependent assembly of TFIIIB. TFIIIB depleted of TBP by a simple chromatographic step does not bind stably to the TFIIIC-tDNA complex. Addition of yeast or human recombinant TBP allows the formation of a TFIIIB-TBP-TFIIIC-tDNA complex. The presence of TBP in the complex was inferred from the effect of anti-TBP antibodies and from the different migration properties of TFIIIB-TBP-tDNA complexes formed with yeast or human TBP.