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Volume 272, Number 6, Issue of February 7, 1997 pp. 3852-3859
©1997 by The American Society for Biochemistry and Molecular Biology, Inc.

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Transcription of the Acanthamoeba TATA-binding Protein Gene
A SINGLE TRANSCRIPTION FACTOR ACTS BOTH AS AN ACTIVATOR AND A REPRESSOR

(Received for publication, September 12, 1996, and in revised form, November 12, 1996)

From the Department of Microbiology and Molecular Genetics, Cell and Molecular Biology Program, Markey Center for Molecular Genetics, University of Vermont, Burlington, Vermont 05405

Transcription of the Acanthamoeba TATA-binding protein (TBP) gene is regulated by TBP promoter-binding factor (TPBF), a previously described transactivator that binds as a tetramer to the TBP Promoter Element (TPE) and stimulates transcription up to 10-fold in vitro. Here we report that TPBF also functions as a transcription repressor by binding to a negative cis-element, located between the TATA box and the transcription initiation site. The negative element, referred to as the nTPE, is structurally similar to the TPE, and its disruption increases the transcription potency of the TBP promoter. TPBF binds to the nTPE, as demonstrated by mobility shift assays. However, the binding affinity of TPBF for the nTPE is about 10-fold lower than for the TPE. When placed upstream of the TATA box, the nTPE has very little effect on transcription. However, it inhibits transcription when placed at several positions downstream of the TATA box. Mechanistic studies with the TBP promoter suggest that binding of TPBF to the nTPE not only prevents TBP from binding to the TATA box but also displaces bound TBP, thereby inhibiting further assembly of the preinitiation complex. These results suggest a mechanism in which the cellular TPBF concentration controls the level of TBP gene transcription and show that a single factor can be stimulatory, inhibitory, or neutral depending on the sequence and the context of its binding site.

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