Vertebrate TBP-like protein (TLP/TRF2/TLF) stimulates TATA-less terminal deoxynucleotidyl transferase promoters in a transient reporter assay, and TFIIA-binding capacity of TLP is required for this function

Abstract

The TBP-like protein (TLP/TRF2/TLF), which belongs to the TBP family of proteins, is present in all metazoan organisms. Although the human TLP has been reported to interfere with transcription from TATA-containing promoters, the transcription activation potential of TLP in higher animals is obscure. We previously demonstrated that artificially promoter-recruited TLP behaves like an unconventional transcriptional activator. In this study, we investigated the effects of TLP on TATA-less promoters of mouse and human terminal deoxynucleotidyl transferase (TdT) genes by transient reporter assays. As expected, TLP repressed both basal and activator-augmented transcription from the TATA-containing adenovirus major late promoter (MLP) and E1B promoter. On the other hand, however, TLP significantly stimulated both basal and activated transcription from TdT promoters. We investigated the strength of the promoters in chicken DT40 cells that lack the TLP gene. The MLP showed higher activity but the TdT promoter showed lower activity in TLP-null cells than in the wild-type cells. Moreover, ectopic expression of mouse TLP in the TLP-null cells considerably stimulated the TdT promoter. Insertion of a TATA element upstream from the TdT core promoter resulted in a loss of TLP-mediated activation. The mouse TLP was demonstrated to bind specifically to TFIIA with greater strength than TBP. We constructed mutated TLPs having amino acid substitutions that impair TFIIA binding. A representative TLP mutant lacking TFIIA-binding ability could not stimulate transcription from the TdT promoter, whereas that mutation suppressed TLP-mediated transcription repression of TATA promoters. The results of the present study suggest that the vertebrate TLP potentiates exogenous TATA-less promoters and that TFIIA plays an important role in the TLP function.

TATA-binding Protein (TBP)-like Protein Is Required for p53-dependent Transcriptional Activation of Upstream Promoter of p21 Gene

“…The upstream and downstream promoters are TATA-less and TATA-containing promoters, respectively. The p21 promoter(s) in p21luc1 was not activated by TLP but rather inhibited as observed previously for several TATA-containing promoters (28,34). Because p21luc1 harbors two tan- dem-orienting promoters, the effect of TLP on the downstream promoter alone was not elucidated from this construct.…”

“…By using TLP knock-out chicken DT40 cells, we demonstrated that TLP inhibits G 2 -M transition (32). Although TLP has no obvious sequence-specific DNA binding activity, many lines of evidence indicate that TLP has a transcription-activating ability (33,34). Through cell-based transient promoter assays, we have demonstrated that TLP activates several promoters that lack a TATA box, whereas TATA-containing promoters are repressed for an unknown reason (28,34,35).…”

“…Although TLP has no obvious sequence-specific DNA binding activity, many lines of evidence indicate that TLP has a transcription-activating ability (33,34). Through cell-based transient promoter assays, we have demonstrated that TLP activates several promoters that lack a TATA box, whereas TATA-containing promoters are repressed for an unknown reason (28,34,35). It has also been shown that TLP regulates cellular genes including cyclin G2 (32), TAp63 (18), wee1 (32,36), proliferating cell nuclear antigen (32,37), and NF1 (38), which are related to cell cycle regulation, apoptosis induction, tumor suppression, and DNA repair, implying that TLP works for cell integrity and growth control.…”

“…A critical factor of this phenomenon is probably the presence of a TATA box in the promoter. It is known that a TATA-containing promoter is repressed by overexpressed TLP in a promoter-reporter assay condition (28,34,35). Because TATA promoters in chromatin are not necessarily inhibited by TLP, a non-physiological chromatin structure may be related to this phenomenon.…”

Taf7l cooperates with Trf2 to regulate spermiogenesis

“…We postulate that a testis-specific transcription preinitiation complex containing both TAF7L and TRF2 is formed and targeted to a subset of "TATA-less" promoters to regulate postmeiotic Trf2-dependent gene expression (43)(44)(45) (Fig. S9).…”