BINDING OF A CTF NF1-LIKE PROTEIN TO THE MOUSE COLONY-STIMULATING FACTOR-I GENE PROMOTER/

Circulating and tissue-specific monocytes/macrophages, through product ion 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 ti ssues. 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 regu latory regions of the mouse CSF-1 gene. Analysis of heterologous repor ter constructs containing the mouse CSF-1 promoter linked to the bacte rial chloramphenicol acetyltransferase (CAT) gene in transiently trans fected fibroblasts identified a cis-acting element located between bas e pairs -88 and -43 of the CSF-1 gene. Electrophoretic mobility-shift assays (EMSAs) and DNase I protection assays with nuclear extracts iso lated from proliferating fibroblasts revealed distinct protein binding to the region spanning base pairs -90 to -68. Results from methylatio n 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 activ ity in transiently transfected fibroblasts and monocytes. Therefore, w e have demonstrated that CTF/NF1 or a CTF/NF1-like protein binds to th e CSF-1 gene promoter; however, binding of the CTF/NF1-like protein al one does not significantly effect changes in CSF-1 gene promoter activ ity.