Involvement of transcription factor c/ebp [beta] in stimulation of
pepck gene expression during exercise.
Nizielski, Steven E., Carmen Arizmendi, Ann R. Shteyngarts, Craig J.
Farrell, and Jacob E. Friedman.
Departments of Nutrition and Biochemistry, Case Western Reserve
University School of Medicine, Cleveland, OH 44106-4935 and
Department of Biochemistry and Molecular Biology, University of
Salamanca, Salamanca, Spain E-37007
APStracts 2:0339R, 1995.
Prolonged exercise increases gluconeogenesis and activates
transcription of the hepatic phosphoenolpyruvate carboxykinase
(PEPCK) gene. The mechanisms that regulate the transcriptional
control of gene expression depend upon the interaction of nuclear
proteins with distinct DNA sequences. To determine the involvement of
the liver-enriched transcription factor CCAAT/enhancer binding
protein [beta] (C/EBP[beta]) in the induction of PEPCK gene
transcription during prolonged exercise or cAMP treatment, we
examined C/EBP[beta] mRNA and nuclear protein concentrations, as well
as C/EBP[beta] binding to the PEPCK promoter at the CRE (-87/-74) and
P3I (-248/-230) binding sites. The requirement of these DNA elements
for exercise-induced stimulation of PEPCK gene expression was
established in transgenic mice carrying -460/+73 of the PEPCK
promoter with a mutation in either CRE or P3I binding domain, linked
to a bovine growth hormone (bGH) reporter gene. In mice carrying the
intact promoter, prolonged exercise increased the concentration of
liver bGH mRNA by 510% compared to an increase of only 270% in mice
with a mutation in either the CRE or P3I site. Exercise or cAMP
injection induced a 7.5 and 13 fold increase in nuclear C/EBP[beta]
protein, respectively. In electrophoretic mobility shift assays
(EMSA), the total quantity of nuclear proteins bound to either
oligomer was not altered by treatment. However, addition of
C/EBP[beta] antisera in the EMSA in a supershift assay indicated that
liver nuclear extracts from exercised or cAMP treated mice
demonstrated significantly greater DNA binding due to C/EBP[beta]
(CRE: Control 44.4 +/-2.3%, Exercise 56.7 +/-2.2%, cAMP 54.5 +/-3.6%
of total binding, P<0.001; P3I: Control 35.8 +/-2.5%, Exercise
64.9 +/-1.9%, cAMP 57.3 +/-2.5% of total binding, P<0.001).
Taken together, these results suggest that exercise and cAMP
treatment induce a transient increase in C/EBP[beta] that may
contribute to the molecular mechanism for signaling PEPCK gene
transcription and increasing gluconeogenesis during exercise.
Received 13 June 1995; accepted in final form 7 November 1995
APS Manuscript Number R361-5.
Article publication pending Am. J. Physiol. (Regulatory Integrative
Comp. Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 12 December 95