Type I collagen is the major component of many extracellular matrices, and its accumulation characterizes most fibrotic processes. It is synthesized by a small number of discrete cell types, including fibroblasts, osteoblasts and odontoblasts. Analysis of transgenic mice harbouring different segments of the promoters of the mouse pro-alpha1 (I) and pro-alpha2 (I) genes has led to the conclusion that this tissue-specific expression is controlled by different cis-acting elements which are responsible for the expression of type I collagen genes in different type I collagen-producing cells. Transacting factors which bind to these different tissue-specific elements are still unknown, but they probably act by modifying the chromatin structure. In fibroblastic cells, various soluble molecules can modulate the transcription of type I collagen genes. Analysis of the pro-alpha1 (I) and pro-alpha2 (I) proximal promoters has led to the identification of different cis-acting elements which can modulate the expression of reporter genes, in transfection experiments. Among these cis-acting elements, a sequence located between -378 and -183 bp in the human pro-alpha2 (I) promoter appears to mediate the transcriptional effects of transforming growth factor-beta. It binds a large multimeric complex which contains Sp1, as well as AP1 and other DNA-binding proteins which have not yet been identified.

Mice specifically overexpressing TIMP-1 in osteoblasts have been generated to investigate the role of MMPs in bone in vivo. These mice displayed increased trabecular bone volume and decreased bone turnover. This model provides evidence of the role played by the MMPs in bone remodeling and balance.Introduction: Although it has been suggested that the matrix metalloproteinases (MMPs) may play a role in initiating the bone resorption process in vitro, there is no evidence that they play any role in in vivo bone maintenance. Materials and Methods:We used an artificial promoter specifically driving cells of the osteoblastic lineage to overexpress the tissue inhibitor of MMPs (TIMP-1) cDNA in mice. Densitometric analysis, using DXA and pQCT, and static and dynamic histomorphometry were used to evaluate the bone phenotype both in male and female transgenic mice. We evaluated osteoblastic differentiation using a primary osteoblast culture and osteoclast activity using an ex vivo organ culture. Results and Conclusion:We showed that at 1 and 2.5 months of age, only the female mice exhibited a bone phenotype. These mice displayed specific increases in the BMD and bone volume of trabecular bone. This increase was accompanied by decreased trabecular separation, suggesting a decrease in bone resorption. Using an ex vivo resorption assay, we demonstrated that parathyroid hormone (PTH)-stimulated bone resorption was reduced in these mice. Evaluation of the bone histomorphometric dynamic parameters showed that the mineralizing surfaces and bone formation rate were both reduced. There was no change in the mineralization lag time or number of osteocyte lacunae. Using primary osteoblast culture and molecular analysis, we showed that the differentiation and function of osteoblasts from transgenic mice were normal, but that the ex vivo formation of mineralized nodules was delayed. This model is the first to show that in vivo MMPs play a role in bone remodeling and bone balance. Moreover, our data suggest that MMP activity could be involved in the hormonal regulation of bone resorption by osteoblasts.

The transcription of type I collagen genes is tightly regulated, but few cis-acting elements have been identified that can modulate the levels of expression of these genes. Generation of transgenic mice harboring various segments of the mouse pro-␣1(I) collagen promoter led us to suspect that a repressor element was located between ؊10.5 and ؊17 kilobase pairs. Stable and transient transfection experiments in ROS17/2.8 osteoblastic cells confirmed the existence of such a repressor element at about ؊14 kilobase pairs and showed that it consisted in an almost perfect three-time repeat of a 41-base pair sequence. This element, which we named COIN-1, contains three E2-boxes, and a point mutation in at least two of them completely abolished its repressor effect. In gel shift assays, COIN-1 bound a DNAbinding protein named ␦EF1/ZEB-1, and mutations that abolished the repressor effect of COIN-1 also suppressed the binding of ␦EF1. We also showed that the repressor effect of COIN-1 was not mediated by chromatin compaction. Furthermore, overexpression of ␦EF1 in ROS17/ 2.8 osteoblastic cells enhanced the inhibitory effect of COIN-1 in a dose-dependent manner and repressed the expression of the pro-␣1(I) collagen gene. Thus, ␦EF1 appears to repress the expression of the mouse pro-␣1(I) collagen gene, through its binding to COIN-1.Type I collagen is a fibrillar collagen composed of two ␣1 chains and one ␣2 chain coiled around each others in a triple helix. It is the most abundant protein of mammalian bodies, and a major component of most extracellular matrices. In the extracellular space, type I collagen molecules self-assemble into highly organized fibrils and then fibers, which largely contribute to the high tensile strength of the structural framework supporting body structures (reviewed in Ref.

The genes encoding the two type I collagen chains are selectively activated in few cell types, including fibroblasts and osteoblasts. By generating transgenic mice, we have previously shown that the activity of the mouse pro-␣1(I) promoter was controlled by separate cell-specific cis-acting elements. In particular, a sequence located between ؊3.2 and ؊2.3 kb was needed to induce expression of the reporter gene at high levels in tendon fibroblasts. In the present work, by using the same transgenic approach, we have identified two short elements in this sequence, named tendon-specific element (TSE) 1 and TSE2, that were necessary to direct reporter gene expression selectively in tendon fibroblasts. Gel shift assays showed that TSE1 and TSE2 bound proteins specifically present in nuclear extracts from tendon fibroblasts and that the sequence of TSE2 binding a tendon-specific protein corresponded to an E-box. Analysis of transgenic mice further indicated that TSE1 and TSE2 needed to cooperate not only with each other but also with other cis-acting elements of the proximal promoter to activate reporter gene expression in tendon fibroblasts. Similarly, it pointed out that the so-called osteoblast-specific element had to interact with downstream sequences to drive reporter gene expression in osteoblasts of transgenic mice. Thus, expression of the mouse pro-␣1(I) collagen gene in tendon fibroblasts appears to be the result of a unique combination of different cis-acting elements, including TSE1 and TSE2.Type I collagen is a fibrillar collagen composed of two ␣1 chains and one ␣2 chain coiled around each other in a triple helix. It is a major protein of mammalian bodies and an essential component of most extracellular matrices. In the extracellular space, type I collagen molecules self-assemble into highly organized fibrils and then into fibers, which largely contribute to the high tensile strength of the framework supporting body structures (reviewed in Ref. 1). Evidence for a role of type I collagen in providing mechanical strength to tissues comes from pathophysiological analyses of genetic diseases resulting from mutations in one of the genes encoding type I collagen, such as osteogenesis imperfecta and Ehlers-Danlos syndromes type VIIA and VIIB (2). The hallmark of osteogenesis imperfecta is brittle bones, but the disease can also involve other tissues rich in type I collagen, such as ligaments, tendons, fascia, sclerae, and teeth (3). Ehlers-Danlos syndromes are characterized by skin hyperextensibility, vascular fragility, and increased ligament elasticity responsible for joint hypermobility (4).Type I collagen is synthesized by a discrete subset of cells of mesenchymal origin, which contrasts with its wide distribution throughout the body. These cells are mostly fibroblasts, osteoblasts, and odontoblasts. Studies using transgenic mice harboring various fragments of the mouse and rat pro-␣1(I) collagen promoters have shown that a modular arrangement of separate cell-specific cis-acting elements was responsible for ...