The effects of colony-stimulating factor 1 (CSF-1), the primary regulator of mononuclear phagocyte production, are thought to be mediated by the CSF-1 receptor (CSF-1R), encoded by the c-fms proto-oncogene. To investigate the in vivo specificity of CSF-1 for the CSF-1R, the mouse IntroductionColony-stimulating factor 1 (CSF-1) regulates the survival, proliferation, and differentiation of mononuclear phagocytic cells and is the primary regulator of mononuclear phagocyte production in vivo. 1,2 However, CSF-1 also regulates cells of the female reproductive tract and plays an important role in fertility. 3,4 The effects of CSF-1 are mediated by a high-affinity receptor tyrosine kinase (CSF-1R) [5][6][7][8] encoded by the c-fms proto-oncogene. 9 The CSF-1R is expressed on primitive multipotent hematopoietic cells, 10,11 mononuclear phagocyte progenitor cells, 12 monoblasts, promonocytes, monocytes, 5,6 tissue macrophages, 6,13-15 osteoclasts, 16 B cells, 17,18 smooth muscle cells, 19 and neurons. 20,21 CSF-1R messenger RNA (mRNA) is expressed in Langerhans cells, 22 in the female reproductive tract, in oocytes and embryonic cells of the inner cell mass and trophectoderm, 23 in decidual cells, [24][25][26] and in cells of the trophoblast. 24,25 The expression of the CSF-1R on primitive hematopoietic cells that are unable to proliferate in vitro in response to CSF-1 alone 10,11 but are able to proliferate and differentiate if stimulated with combinations of CSF-1 and other hematopoietic growth factors 10,11,27 suggests that CSF-1R is involved in the regulation of more primitive hematopoietic cells than those that form macrophage colonies in vitro in response to CSF-1 alone.Mice homozygous for the mutation osteopetrotic 28 possess an inactivating mutation in the coding region of the CSF-1 gene and are devoid of detectable CSF-1. 29,30 These Csf1 op /Csf1 op mice are osteopetrotic because of an early and marked deficiency of osteoclasts 28 that spontaneously recovers with age, 31,32 probably because of the action of vascular endothelial growth factor. 33 However, the phenotype of these mice is pleiotropic. 3 They are toothless; have low body weight, low growth rate, and skeletal abnormalities; and are deficient in tissue macrophages. 2,28,30,34,35 They have defects in both male and female fertility, neural development, the dermis, and synovial membranes. 3 The pleiotropic phenotype of the Csf1 op /Csf1 op mouse may be due to a reduction in trophic and/or scavenger functions of the tissue macrophages regulated by CSF-1, secondary to the reduction of their concentration in tissues, 2 because outside the female reproductive tract the CSF-1R is primarily expressed in mononuclear phagocytes. 1,3 However, it is possible that some of these effects may also be due to loss of function of other cells such as neuronal cells and muscle precursors, which have also been reported to express the CSF-1R. 20,36 To address the questions of whether CSF-1 activates other receptors besides the CSF-1R and, conversely, whether the CSF-1R me...

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Granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin 3 (IL-3) are pleiotropic hemopoietic growth factors whose genes are closely linked and induced in T lymphocytes in a cyclosporin A (CsA)-sensitive fashion. Since we found that the human GM-CSF and IL-3 proximal promoters were not sufficient to account for the observed regulation of these genes, we mapped DNase I hypersensitive sites across the GM-CSF/IL-3 locus in the Jurkat human T-celi line to identify additional regulatory elements.We located an inducible DNase I hypersensitive site, 3 kb upstream of the GM-CSF gene, that functioned as a strong CsA-sensitive enhancer of both the GM-CSF and IL-3 promoters. Binding studies employing Jurkat cell nuclear extracts indicated that four sites within the enhancer associate with the inducible transcription factor AP1. Three of these AP1 elements lie within sequences that also associate with factors resembling the CsA-sensitive, T cell-specific transcription factor NFAT. We provide additional evidence suggesting that an APl-like factor represents one of the components of NFAT. We propose that the intergenic enhancer described here is required for the correctly regulated activation of both GM-CSF and IL-3 gene expression in T cells and that it mediates the CsA sensitivity of the GM-CSF/IL-3 locus.

The promoter of the human granulocyte-macrophage colony-stimulating factor gene is regulated by an inducible upstream enhancer. The enhancer encompasses three previously defined binding sites for the transcription factor NFAT (GM170, GM330, and GM550) and a novel NFAT site defined here as the GM420 element. While there was considerable redundancy within the enhancer, the GM330, GM420, and GM550 motifs each functioned efficiently in isolation as enhancer elements and bound NFATp and AP-1 in a highly cooperative fashion. These three NFAT sites closely resembled the distal interleukin-2 NFAT site, and methylation interference assays further defined GGA(N) 9 TCA as a minimum consensus sequence for this family of NFAT sites. By contrast, the GM170 site, which also had conserved GGA and TCA motifs but in which these motifs were separated by 15 bases, supported strong independent but no cooperative binding of AP-1 and NFATp, and this site functioned poorly as an enhancer element. While both the GM330 and GM420 elements were closely associated with the inducible DNase I-hypersensitive site within the enhancer, the GM420 element was the only NFAT site located within a 160-bp HincII-BalI fragment defined by deletion analysis as the essential core of the enhancer. The GM420 element was unusual, however, in containing a high-affinity NFATp/c-binding sequence (TGGAAAGA) immediately upstream of the sequence TGACATCA which more closely resembled a cyclic AMP response-like element than an AP-1 site. We suggest that the cooperative binding of NFATp/c and AP-1 requires a particular spacing of sites and that their cooperativity and induction via independent pathways ensure very tight regulation of the granulocyte-macrophage colony-stimulating factor enhancer.