Binding of a CTF/NF1-like protein to the mouse colony-stimulating factor-1 gene promoter.
Circulating and tissue-specific monocytes/macrophages, through production of hydrolytic enzymes and growth factors, can dramatically affect the local tissue environment. Colony-stimulating factor-1 (CSF-1) is a key regulator of monocyte/macrophage cell activity. CSF-1 is produced by stromal elements, including fibroblasts, which are found in all tissues. To understand at the molecular level how changes in CSF-1 gene transcription are initiated in fibroblasts, we set out to identify the cis-acting elements and cognate trans-acting factor(s) that bind regulatory regions of the mouse CSF-1 gene. Analysis of heterologous reporter constructs containing the mouse CSF-1 promoter linked to the bacterial chloramphenicol acetyltransferase (CAT) gene in transiently transfected fibroblasts identified a cis-acting element located between base pairs -88 and -43 of the CSF-1 gene. Electrophoretic mobility-shift assays (EMSAs) and DNase I protection assays with nuclear extracts isolated from proliferating fibroblasts revealed distinct protein binding to the region spanning base pairs -90 to -68. Results from methylation interference assays suggest CTF/NF1 or a CTF/NF1-like factor is the cognate trans-acting factor. Mutation of the putative CTF/NF1 binding site in the CSF-1 promoter lead to a modest decrease in promoter activity in transiently transfected fibroblasts and monocytes. Therefore, we have demonstrated that CTF/NF1 or a CTF/NF1-like protein binds to the CSF-1 gene promoter; however, binding of the CTF/NF1-like protein alone does not significantly effect changes in CSF-1 gene promoter activity.
Konicek
B W
BW
Department of Biochemistry, Indiana University School of Medicine, Indianapolis 46202-5121, USA.
Xia
X L
XL
Harrington
M A
MA
D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D022762:CCAAT-Enhancer-Binding Proteins; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis / Q000235:genetics; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005347:Fibroblasts; D016173:Macrophage Colony-Stimulating Factor / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D009000:Monocytes; D009154:Mutation; D051839:NFI Transcription Factors; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D022762:CCAAT-Enhancer-Binding Proteins; C077305:CTF-1 transcription factor; D004268:DNA-Binding Proteins; D051839:NFI Transcription Factors; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D016173:Macrophage Colony-Stimulating Factor; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase