© 1999 American Heart Association, Inc.
Original Contributions |
On the Role of c-Jun in the Induction of PAI-1 Gene Expression by Phorbol Ester, Serum, and IL-1
in HepG2 Cells
From the Gaubius Laboratory, TNO-PG, Leiden, the Netherlands.
Correspondence to Dr T. Kooistra, Gaubius Laboratory, TNO-PG, PO Box 2215, 2301 CE Leiden, the Netherlands.
Abstract—We have characterized
the regulation of plasminogen activator
inhibitor-1 (PAI-1) gene expression by phorbol
12-myristate 13-acetate (PMA), serum, and interleukin-1
(IL-1) in the human hepatoma cell line HepG2. PMA, serum, and
IL-1 induced a rapid and transient 28-fold (PMA), 9-fold (serum),
and 23-fold (IL-1) increase in PAI-1 mRNA, peaking after 
4 hours.
These inductions of PAI-1 mRNA accumulation were reduced by
pretreatment of the HepG2 cells with the protein tyrosine kinase
inhibitor genistein. Conversely, stimulation of tyrosine
phosphorylation by sodium orthovanadate, an
inhibitor of protein tyrosine phosphatases, caused an
increase in PAI-1 mRNA levels. The effects of PMA, serum, and IL-1
on PAI-1 mRNA expression have been compared with their ability to
modulate the expression of a chloramphenicol acetyltransferase (CAT)
reporter plasmid, which was under control of the -489 to +75 region of
the PAI-1 promoter, and stably transfected into HepG2 cells. This
region of the PAI-1 promoter was previously found to contain a
tetradecanoyl phorbol acetate–response element (TRE; between -58 and
-50) necessary for PMA responsiveness and with a high affinity for
c-Jun homodimers. Whereas incubation of these transfected HepG2 cells
with PMA and serum showed an induction profile of CAT mRNA similar to
that of PAI-1 mRNA, hardly any induction of CAT mRNA was found with
IL-1. In line with these findings, IL-1 poorly induced c-Jun
homodimer binding to the PAI-1 TRE in gel mobility-shift assays.
Pretreatment of HepG2 cells with the protein kinase C
inhibitor Ro 31-8220 or the mitogen-activated
protein kinase kinase (MAPKK)1,2 activity blocker PD98059
selectively suppressed the induction of PAI-1 (and CAT) expression by
PMA, but not that by IL-1. In contrast, the protein tyrosine kinase
inhibitor herbimycin A blocked PAI-1 mRNA induction by IL-1
only. We propose 2 separate PAI-1 inductory pathways for PMA and
IL-1 in HepG2, both involving protein tyrosine kinase activation;
the serum-induced signaling pathway may (partially) overlap with the
PMA-activated protein kinase C/mitogen-activated
protein kinase kinase pathway, leading to c-Jun homodimer binding to
the PAI-1 TRE.
Key Words: plasminogen activator inhibitor-1 • protein kinase C • interleukin-1 • gene transcription • c-Jun
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