Binding of a CTF/NF1-Like Protein to the Mouse Colony-Stimulating Factor-1 Gene Promoter

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Abstract
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.