Differential regulation of MAGE-A1 promoter activity by BORIS and Sp1, both interacting with the TATA binding protein

ABSTRACT 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 Alapromoter 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.

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Transcriptional Regulation of the TATA-Binding Protein by Ras Cellular Signaling

Molecular and Cellular Biology ◽

10.1128/mcb.20.14.5000-5009.2000 ◽

2000 ◽

Vol 20 (14) ◽

pp. 5000-5009 ◽

Cited By ~ 27

Author(s):

Sandra A. S. Johnson ◽

Nihar Mandavia ◽

Horng-Dar Wang ◽

Deborah L. Johnson

Keyword(s):

Transcription Factor ◽

Promoter Activity ◽

Binding Protein ◽

Rat Hepatocytes ◽

Tata Binding Protein ◽

Mitogen Activated Protein Kinase ◽

Transcriptional Level ◽

Ras Signaling ◽

Dependent Manner ◽

X Protein

ABSTRACT Our previous studies have demonstrated that the level of the central transcription factor TATA-binding protein (TBP) is increased in cells expressing the hepatitis B virus (HBV) X protein through the activation of the Ras signaling pathway, which serves to enhance both RNA polymerase I and III promoter activities. To understand the mechanism by which TBP is regulated, we have investigated whether enhanced expression is modulated at the transcriptional level. Nuclear run-on assays revealed that the HBV X protein increases the number of active transcription complexes on the TBP gene. In transient-transfection assays with both transformed and primary hepatocytes, the human TBP promoter was shown to be induced by expression of the HBV X protein in a Ras-dependent manner, requiring both Ral guanine nucleotide dissociation stimulator (RalGDS) and Raf signaling. Transient overexpression of TBP did not affect TBP promoter activity. To further delineate the downstream Ras-mediated events contributing to TBP promoter regulation in primary rat hepatocytes, the best-characterized Ras effectors, Raf, phosphoinositide 3-kinase (PI-3 kinase), and RalGDS, were examined. Activation of either Raf or RalGDS, but not that of PI-3 kinase, was sufficient to induce TBP promoter activity. Both Raf- and RalGDS-mediated induction required the activation of mitogen-activated protein kinase kinase (MEK). In addition, another distinct Ras-activated pathway, which does not require MEK activation, appears to induce TBP promoter activity. Analysis of the DNA sequence requirement within the TBP promoter responsible for these regulatory events defined three distinct regions that modulate the abilities of Raf, RalGDS, and the Ras-dependent, MEK-independent pathways to regulate human TBP promoter activity. Together, these results provide new evidence that TBP can be regulated at the transcriptional level and identify three distinct Ras-activated pathways that modulate this central eukaryotic transcription factor.

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