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(Received for publication, June 27, 1997)
From the Division of Endocrinology, Diabetes, and Metabolism,
Department of Medicine, University of Colorado Health Sciences Center,
Denver, Colorado 80262
The molecular determinants governing
cell-specific expression of the thyrotropin (TSH)
Volume 272, Number 39,
Issue of September 26, 1997
pp. 24339-24347
©1997 by The American Society for Biochemistry and Molecular Biology, Inc.
-Subunit Promoter
-subunit gene in
pituitary thyrotropes are not well understood. The P1 region of the
mouse TSH promoter (
133 to 88) region interacts with Pit-1 and
an additional 50-kDa factor at an adjacent site that resembles a
consensus GATA binding site. Northern and Western blot assays
demonstrated the presence of GATA-2 transcripts and protein in TtT-97
thyrotropic tumors. In electrophoretic mobility shift assays, a
comigrating complex was observed with both TtT-97 nuclear extracts and
GATA-2 expressed in COS cells. The complex demonstrated binding
specificity to the P1 region DNA probe and could be disrupted by a
GATA-2 antibody. When both Pit-1 and GATA-2 were combined, a slower
migrating complex, indicative of a ternary protein-DNA interaction was
observed. Cotransfection of both Pit-1 and GATA-2 into CV-1 cells
synergistically stimulated mouse TSH promoter activity 8.5-fold,
while each factor alone had a minimal effect. Mutations that abrogated
this functional stimulatory effect mapped to the P1 region. Finally, we
show that GATA-2 directly interacts with Pit-1 in solution. In summary, these data demonstrate functional synergy and physical interaction between homeobox and zinc finger factors and provide insights into the
transcriptional mechanisms of thyrotrope-specific gene expression.
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