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

摘要

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