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J Biol Chem, Vol. 275, Issue 19, 14524-14531, May 12, 2000

Involvement of Thyrotroph Embryonic Factor in Calcium-mediated Regulation of Gene Expression^*

Darcy A. Krueger, Elizabeth A. Warner^§, and Diane R. Dowd^¶

From the  E. A. Doisy Department of Biochemistry and Molecular Biology, St. Louis University Health Sciences Center, St. Louis, Missouri 63104 and the ^¶ Department of Pharmacology, Case Western Reserve University, Cleveland, Ohio 44106

In the present study, we used an expression cloning strategy to identify transcription factors that bind specifically to a limited region of the inducible cAMP early repressor (ICER) promoter and regulate transcription. Murine thyrotroph embryonic factor (mTEF) was isolated and was shown to bind to a site located at nucleotides -117 to -108 from the transcriptional start site. Transient expression of reporter constructs containing either a consensus TEFRE or the icerTEF binding site demonstrated that TEF-dependent transcription correlated with relative binding affinities, i.e. the consensus TEFRE bound TEF more tightly and was more responsive to TEF than the icerTEFRE. Because the icerTEFRE overlapped a cAMP response element, the responsiveness of these sequences to either cAMP or Ca²⁺ was tested. Although TEF expression had no effect on the cAMP-regulated transcriptional response of the ICER promoter, TEF did confer calcium responsiveness to these sequences. Calcium also modestly increased the TEF-mediated transcription from a consensus TEFRE. Additional studies using Ca²⁺-activated kinases indicate that Ca²⁺/TEF/TEFRE-regulated transcription may be mediated through Ca²⁺/calmodulin-dependent kinase (CaMK) IV. Moreover, studies with the icerTEFRE in a CaMK IV-deficient cell line demonstrated that these cells were transcriptionally unresponsive to thapsigargin; however, responsiveness was restored by co-expression of the active CaMK IV. These studies are the first to demonstrate that TEF is a calcium-responsive transcription factor, and they suggest that there are two classes of TEF-regulated genes. One class, represented by a consensus TEFRE, is regulated by TEF in the resting cell; the second class, represented by icerTEFRE, is regulated by TEF in the calcium-activated cell.

The nucleotide sequence(s) reported in this paper has been submitted to the GenBank^TM/EMBL Data Bank with accession number(s) AF194420.

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