Transcriptional regulation of fibroblast growth factor-2 expression in human astrocytes: implications for cell plasticity.
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-1beta, 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.
Molecular and Structural Neurobiology and Gene Therapy Program, Barrow Neurological Institute, Phoenix, Arizona 85013, USA.
J
J
Moffett
E
E
Kratz
J
J
Myers
E K
EK
Stachowiak
R Z
RZ
Florkiewicz
M K
MK
Stachowiak
D001253:Astrocytes / Q000166:cytology / Q000378:metabolism* / Q000502:physiology; D001483:Base Sequence; D001665:Binding Sites; D001921:Brain / Q000166:cytology / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000494:pharmacology; D016222:Fibroblast Growth Factor 2 / Q000096:biosynthesis / Q000235:genetics* / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D008156:Luciferases / Q000096:biosynthesis; D008938:Mitosis; D008969:Molecular Sequence Data; D009473:Neuronal Plasticity / Q000502:physiology*; D009838:Oligodeoxyribonucleotides / Q000494:pharmacology; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection
D004268:DNA-Binding Proteins; D007375:Interleukin-1; D009838:Oligodeoxyribonucleotides; D016376:Oligonucleotides, Antisense; D011993:Recombinant Fusion Proteins; D016222:Fibroblast Growth Factor 2; D005576:Colforsin; D004815:Epidermal Growth Factor; D008156:Luciferases; D013755:Tetradecanoylphorbol Acetate