TFII-I Regulates Vβ Promoter Activity through an Initiator Element

ABSTRACT

In our effort to understand the transcriptional regulation of naturally occurring TATA-less but initiator (Inr)-containing genes, we have employed the murine T-cell receptor Vβ 5.2 promoter as a model. Here we show by transient-transfection assays that the Inr binding transcription factor TFII-I is required for efficient expression of the Vβ promoter in vivo. Mutations in the Inr element that reduced binding of TFII-I also abolished the Vβ promoter activity by ectopic TFII-I. We further biochemically identified a protease-resistant N-terminal DNA binding fragment of TFII-I, p70. When ectopically expressed, recombinant p70 bound to the Vβ Inr element with a specificity similar to that of wild-type TFII-I. More importantly, p70, which lacks independent activation functions, behaved as a dominant negative mutant that inhibited Inr-specific function of wild-type TFII-I. However, the activation functions of p70 were restored when fused to the heterologous activation domain of the yeast activator protein GAL4. Taken together, these data suggest that TFII-I functions in vivo require an intact Inr element and that the Inr-specific transcriptional functions of TFII-I are solely dictated by its N-terminal DNA binding domain and do not require its own C-terminal activation domain.

ACKNOWLEDGMENTS

N-terminal microsequencing and DNA sequencing were performed at Tufts University protein sequencing facility. We are grateful to Jeff Parvin, Brent Cochran, and Michael Meisterernst for plasmids and reagents. We are particularly thankful to Ranjan Sen for support and critically reading the manuscript. Finally, we thank Bob Roeder for helpful suggestions.

This work was funded by grants from the American Cancer Society (RPG-98-104-01-TBE) to A.L.R.