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(Received for publication, December 19, 1996, and in revised form, February 14, 1997)
From the Department of Medicine, Cedars-Sinai Research
Institute-UCLA School of Medicine, Los Angeles, California 90048
Using murine AtT20 pituitary cells transfected
with a rat pro-opiomelanocortin (POMC) promoter (
Volume 272, Number 16,
Issue of April 18, 1997
pp. 10551-10557
©1997 by The American Society for Biochemistry and Molecular Biology, Inc.
706/+64) linked to
the luciferase reporter, we showed leukemia inhibitory factor (LIF) to
strongly potentiate corticotropin-releasing hormone (CRH) induction of POMC gene expression. We therefore tested mechanisms for molecular interactions between LIF and CRH. Although LIF and CRH synergized to
induce an 8-fold induction of POMC transcription, CRH alone (but not
LIF) induced cAMP response element-binding protein phosphorylation (5-fold) or an increase of c-fos mRNA levels
(>100-fold), suggesting that these pathways are not implicated in LIF
transcriptional synergistic effects. Using a DNase I footprint assay,
POMC promoter regions protected by AtT20 cell nuclear extracts were
identified (121/109, and 143/134, and 173/160). The
protected 173/160 element fused to a heterologous promoter
conferred LIF-CRH synergy (6.5-fold induction of POMC) and formed a
specific complex with AtT20 cell nuclear extracts. This complex was
supershifted by an anti-phosphoserine antibody, and a serine/threonine
kinase inhibitor also altered both this complex and LIF-CRH
transcriptional synergy on the POMC promoter-luciferase reporter
construct, indicating that these events depend on post-translational
serine phosphorylations. LIF-CRH synergy on POMC transcription is
therefore mediated at least in part by 173/160 sequences conferring
confluent transcriptional activity of both peptides.
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