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Vol. 9, Issue 8, 2269-2285, August 1998
§ and
*Molecular and Structural Neurobiology and Gene Therapy
Program, Barrow Neurological Institute, Phoenix, Arizona 85013, Arizona
State University, Tempe, Arizona 85287, and State University of New
York, Buffalo, New York 14214; and
Induction of the fibroblast growth factor-2 (FGF-2) gene
and the consequent accumulation of FGF-2 in the nucleus are operative events in mitotic activation and hypertrophy of human astrocytes. In
the brain, these events are associated with cellular degeneration and
may reflect release of the FGF-2 gene from cell contact inhibition. We
used cultures of human astrocytes to examine whether expression of
FGF-2 is also controlled by soluble growth factors. Treatment of
subconfluent astrocytes with interleukin-1Scripps Research
Institute, La Jolla, California 92037
, epidermal or
platelet-derived growth factors, 18-kDa FGF-2, or serum or direct
stimulation of protein kinase C (PKC) with phorbol 12-myristate
13-acetate or adenylate cyclase with forskolin increased the
levels of 18-, 22-, and 24-kDa FGF-2 isoforms and FGF-2 mRNA.
Transfection of FGF-2 promoter-luciferase constructs identified a
unique 
555/513 bp growth factor-responsive element (GFRE) that
confers high basal promoter activity and activation by growth factors
to a downstream promoter region. It also identified a separate region
(624/556 bp) essential for PKC and cAMP stimulation. DNA-protein
binding assays indicated that novel cis-acting elements
and trans-acting factors mediate activation of the FGF-2
gene. Southwestern analysis identified 40-, 50-, 60-, and 100-kDa
GFRE-binding proteins and 165-, 112-, and 90-kDa proteins that
interacted with the PKC/cAMP-responsive region. The GFRE and the
element essential for PKC and cAMP stimulation overlap with the region
that mediates cell contact inhibition of the FGF-2 promoter. The
results show a two-stage regulation of the FGF-2 gene: 1) an initial
induction by reduced cell contact, and 2) further activation by growth
factors or the PKC-signaling pathway. The hierarchic regulation of the
FGF-2 gene promoter by cell density and growth factors or PKC reflects
a two-stage activation of protein binding to the GFRE and to the
PKC/cAMP-responsive region, respectively.
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