A TATA Element Is Required for tRNA Promoter Activity and Confers TATA-binding Protein Responsiveness in DrosophilaSchneider-2 Cells

Abstract

In contrast to yeast and mammalian systems, which depend principally on internal promoter elements for tRNA gene transcription, insect systems require additional upstream sequences. To understand the function of the upstream sequences, we have asked whether the Bombyx mori tRNA C Ala and tRNA SG Ala genes, which are absolutely dependent on these sequences in vitro, also require them for transcription in vivo. We introduced wild-type and mutant versions of the Bombyx tRNA Ala genes into Drosophila Schneider-2 cells and found that the tRNA C Ala gene is efficiently transcribed and that its transcription depends strongly on the distal segment of its upstream promoter. In contrast, the tRNA SG Ala gene is inefficiently transcribed, and this inefficiency results from lack of a specific sequence within the distal tRNA C Ala upstream promoter. This sequence, 5-TTT-ATAT-3, is sufficient to increase the activity of the tRNA SG Ala promoter to that of the tRNA C Ala promoter. Moreover, promoters containing the 5-TTTATAT-3 element are stimulated by increased levels of cellular TATA-binding protein. Together these results indicate that, in insect cells, a TATA-like element is specifically required to form functional TATA-binding protein-containing complexes that promote efficient transcription of tRNA genes.

A Composite Upstream Sequence Motif Potentiates tRNA Gene Transcription in Yeast

“…the 2 35 TTTTTC element in C. elegans), even though TATA-like sequences might be present and play important roles in special cases. 5,6 Also, completely new blocks of sequence conservation, located near position 2 20, appear in insect and mammals, as exemplified by the nearly identical A-rich motif found at position 2 22 upstream of mouse and human tDNAs. Because of its striking evolutionary conservation, such a motif is likely to play an important role in mammalian tRNA gene transcription.…”

“…The differential in vitro activity of silkgland-specific and constitutive tRNA Ala genes in the silkworm, Bombyx mori, has long been known to be due to sequence elements upstream of the TSS, 4 and it has later been shown to rely on the interaction of the TBP component of TFIIIB with a TATA-like element upstream of the constitutive gene. 5,6 Recent analyses have further revealed the presence of TATA-like elements upstream of many Saccharomyces cerevisiae tDNAs, and of the majority of Arabidopsis thaliana and Schizosaccharomyces pombe tDNAs. 7,8 The presence of upstream TATA elements is thus not restricted to particular class III genes, such as SNR6 (coding for the U6 snRNA), that have long been known to support TFIIIC-independent TFIIIB recruitment in vitro.…”

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

“…It is unlikely, however, that increased TBP concentration is sufficient to rescue tRNA SG Ala transcription in the silk gland, since tRNA SG Ala transcription complexes contain normal amounts of TBP but are nonetheless functionally impaired. Indeed, the tRNA SG Ala promoter does not respond in vivo to elevated levels of TBP that are sufficient to stimulate the tRNA C Ala promoter (49). Moreover, the presence of a TBP binding site in the wrong part of the tRNA SG Ala promoter could (33).…”

A Novel Upstream RNA Polymerase III Promoter Element Becomes Essential When the Chromatin Structure of the Yeast U6 RNA Gene Is Altered

“…For example, like TFIID, TFIIIB contains the TATA-binding protein as a subunit. Although most Pol III transcription units lack a consensus TATA box in the TFIIIB-binding region, A/T-rich sequences 15 to 30 base pairs upstream of the start site are known to contribute to the efficiency of tRNA and 5S rRNA gene transcription in vitro (29, 38) and in vivo (23,40). In addition, an increasing number of Pol II transcription units have been found to contain TFIID-binding promoter elements 20 to 30 base pairs downstream of the transcription start site in a position analogous to that of the Pol III A block element (3, 33).…”