TATA-Binding Protein–TATA Interaction Is a Key Determinant of Differential Transcription of Silkworm Constitutive and Silk Gland-Specific tRNA Ala Genes

Abstract

ABSTRACT We have investigated the contribution of specific TATA-binding protein (TBP)–TATA interactions to the promoter activity of a constitutively expressed silkworm tRNA C Ala gene and have also asked whether the lack of similar interactions accounts for the low promoter activity of a silk gland-specific tRNA SG Ala gene. We compared TBP binding, TFIIIB-promoter complex stability (measured by heparin resistance), and in vitro transcriptional activity in a series of mutant tRNA C Ala promoters and found that specific TBP-TATA contacts are important for TFIIIB-promoter interaction and for transcriptional activity. Although the wild-type tRNA C Ala promoter contains two functional TBP binding sequences that overlap, the tRNA SG Ala promoter lacks any TBP binding site in the corresponding region. This feature appears to account for the inefficiency of the tRNA SG Ala promoter since provision of either of the wild-type TATA sequences derived from the tRNA C Ala promoter confers robust transcriptional activity. Transcriptional impairment of the wild-type tRNA SG Ala gene is not due to reduced incorporation of TBP into transcription complexes since both the tRNA C Ala and tRNA SG Ala promoters form transcription complexes that contain the same amount of TBP. Thus, the deleterious consequences of the lack of appropriate TBP-TATA contacts in the tRNA SG Ala promoter must come from failure to incorporate some other essential transcription factor(s) or to stabilize the complete complex in an active conformation.