This project is aimed at understanding cellular and molecular mechanisms that govern colony stimulating factor-1 (CSF-1) gene expression. Our hypothesis is that a basal set of trans-acting factors is bound to the CSF-1 gene transcription, by growth-arrest (decrease) or stimulation of growth-arrested fibroblasts (re-initiate) is mediated by changes in the basal set of factors bound or by the addition of stimulus specific factors. CSF-1-CAT reporter constructs in transient transfection assays will be used to identify genomic sequences that affect transcriptional activity; DNase I protection, electrophoretic mobility shift and methylating interference assays will be used to identify putative cis- acting elements. CSF-1-CAT constructs containing mutated cis-acting elements will be used in transient transection assays to determine if the mutation affects transcriptional activity. As cis-acting elements that affect CSF-1-CAT reporter construct activity are identified, cognate trans-acting factors will be identified, by antibody or by molecular cloning. We will extend our hypothesis to include other cells types to determine if mechanisms used to control CSF-1 gene expression in fibroblasts are unique or represent common non-tissue specific regulatory mechanisms. We will determine if cis-acting element(s) that affect CSF- 1-CAT reporter construct activity in fibroblasts affect CSF-1-CAT activity in monocytes, endothelial cells and sertoli cells. We want to include in our hypothesis how a signal at the plasma membrane in growth- arrested fibroblasts results in activation of CSF-1 gene transcription. We propose to use immunoprecipitates of the IL-1 receptors in fibroblasts. Identification of bound protein(s) will be done by sequence analysis of cloned cDNA inserts or bound protein or alternatively by immunoreactivity. Identification of trans-acting factors that affect CSF-1 gene transcription and signaling pathways that modulate trans- acting factor binding activity will identify targets for therapeutic intervention in situations, such as cancer, atherosclerosis or infertility, where control over CSF-1 gene expression may be desirable.
| Song, A; Xia, X L; Siddiqui, R A et al. (1998) PLA2-independent induction of CSF-1 gene expression in growth-arrested fibroblasts. Int J Hematol 67:249-56 |
