We find striking similarities in promoter structure and requirements for template commitment on 5S RNA and tRNA genes from silkworms. The promoters are nearly the same size (approximately 160 bp) and include flanking as well as internal sequences. To analyze the factor requirements for 5S RNA transcription complex assembly in a completely homologous system, we have isolated a silkworm fraction that is highly enriched for the 5S RNA-specific transcription factor, TFIIIA. Using this fraction, together with the other silkworm fractions, TFIIIB, TFIIIC, TFIIID and RNA polymerase III, we demonstrate that the requirements for 5S RNA transcription complex assembly are very similar to those previously established for a tRNA(C)(Ala) gene. Specifically, no individual factor fraction is sufficient for commitment of silkworm 5S RNA genes to transcription complex assembly. Rather, combinations of at least three factor fractions are required. Our observation that more than one subset of factors is competent for commitment suggests that silkworm 5S RNA genes further resemble tRNA(C)(Ala) genes in their ability to use multiple pathways for transcription complex formation.