Separate cis-acting DNA Elements of the Mouse Pro-cd(I) Collagen Promoter Direct Expression of Reporter Genes to Different Type I Collagen-producing Cells in Transgenic Mice

The genes coding for the two type I collagen chains, which are active selectively in osteoblasts, odontoblasts, fibroblasts, and some mesenchymal cells, constitute good models for studying the mechanisms responsible for the cell-specific activity of genes which are expressed in a small number of discrete cell types. To test whether separate genetic elements could direct the activity of the mouse pro-~l(I) collagen gene to different cell types in which it is expressed, transgenic mice were generated harboring various fragments of the proximal promoter of this gene cloned upstream of the Escherichia coli [3-galactosidase gene. During embryonic development, X-gal staining allows for the precise identification of the different cell types in which the [3-galactosidase gene is active. Transgenic mice harboring 900 bp of the pro-al(I) proximal promoter expressed the transgene at relatively low levels almost exclusively in skin. In mice containing 2.3 kb of this proximal promoter, the transgene was also expressed at high levels in osteoblasts and odontoblasts, but not in other type I collagen-producing cells. Transgenic mice harboring 3.2 kb of the proximal promoter showed an additional high level expression of the transgene in tendon and fascia fibroblasts. The pattern of expression of the lacZ transgene directed by the 0.9and 2.3-kb proa l ( I ) proximal promoters was confirmed by using the firefly luciferase gene as a reporter gene. The pattern of expression of this transgene, which can be detected even when it is active at very low levels, paralleled that of the [3-galactosidase gene. These data strongly suggest a modular arrangement of separate cell-specific c/s-acting elements that can activate the mouse pro-a(I) collagen gene in different type I collagen-producing cells. At least three different types of cell-specific elements would be located in the first 3.2 kb of the promoter: (a) an element that confers low level expression in dermal fibroblasts; (b) a second that mediates high level expression in osteoblasts and odontoblasts; and (c) one responsible for high level expression in tendon and fascia fibroblasts. Our data also imply that other cis-acting cell-specific elements which direct activity of the gene to still other type I collagen-producing cells remain to be identified. D IFFERENT mechanisms can be envisioned to account for the cell-specific activity of a gene that is expressed in a small number of discrete cell types. A first possible mechanism is that a single type of cis-acting element recognized by a unique cell-specific transcription factor be present in the different cell types that are actively expressing this gene. According to another hypothesis, separate cis-acting elements would be present in the regulatory segments of such genes and each of these would be recognized by distinct transcription factors speAddress all correspondence to J6rome Rossert, Dept. of Molecular Genetics, Box 11, 1515 Holcombe Blvd., Houston, TX 77030. Tel.: (713) 7922590. Fax: (713) 794-4295. cific for each of the cell types in which the gene is active. The genes for the type I collagen chains are good models for testing these possibilities. Indeed, these genes are active in several discrete cell types, such as osteoblasts, odontoblasts, and various types of fibroblasts and mesenchymal cells. In case the second hypothesis were true, it would also be important to begin to identify the individual elements as a prelude to the isolation of the various lineagespecific transcription factors which would bind to these elements. Although several laboratories have generated transgenic mice harboring various regulatory segments of the human and rat pro-od(I) collagen genes, the precise sequences responsible for the lineage-specific expression of this gene have not yet been defined. These previous exper© The Rockefeller University Press, 0021-9525/95/06/1421/12 $2.00 The Journal of Cell Biology, Volume 129, Number 5, June 1995 1421-1432 1421 on Jne 7, 2017 D ow nladed fom Published June 1, 1995