Karcagi Ildikó, Rauch Tibor, Hiripi László, Rentsendorj Otgonchimeg, Nagy Andrea, Bõsze Zsuzsa, Kiss Ibolya
Institute of Biochemistry, Biological Research Center of the Hungarian Academy of Sciences, P.O. Box 521, H-6701 Szeged, Hungary.
Matrix Biol. 2004 Feb;22(8):605-18. doi: 10.1016/j.matbio.2003.11.009.
Matrilin-1 is a non-collagenous protein, which functions in the organization of the extracellular matrix by forming collagen-dependent and -independent filamentous networks. It is secreted primarily by chondrocytes in a characteristic spatial, temporal and developmental stage-specific pattern during skeletogenesis. As a first step to define the tissue- and site-specific regulatory regions of the chicken matrilin-1 gene in vivo, we generated transgenic mice harboring various promoter and intronic fragments fused to the LacZ reporter gene. Histological analysis of the transgene expression pattern during ontogenic development revealed specific X-gal staining in most primordial elements of endochondral bones of transgenic mouse lines carrying either the long promoter between -2011 and +67 or the intronic fragment with a short promoter between -338 and +1819. The cartilage-specific activity of the latter transgene, however, was accompanied with variable ectopic expression pattern in neural and other tissues depending on the site of integration. The presence of both promoter upstream and intronic elements was necessary for the high level transgene activity in all chondrogenic tissues and for the extraskeletal transgene expression pattern resembling the most to that of the chicken matrilin-1 gene, e.g. expression in the eye, and lack of expression in the diminishing notochord and nucleus pulposus. The activity of the transgenes was restricted to the columnar proliferating and pre-hypertrophic chondrocytes visualized by BrdU incorporation and distribution of phosphorylated Sox9, respectively. DNA elements between -2011 and -338 also mediated ectopic LacZ expression in cells of neural crest origin. These results suggest that an interplay of modularly arranged cartilage- and neural crest-specific DNA elements control the expression of the matrilin-1 gene. The dispersal of cartilage-specific elements in the promoter upstream and intronic regions shows similarity to the transcriptional regulation of the Col11a2 gene.
基质金属蛋白酶-1是一种非胶原蛋白，通过形成依赖胶原蛋白和不依赖胶原蛋白的丝状网络在细胞外基质的组织中发挥作用。在骨骼发生过程中，它主要由软骨细胞以特征性的空间、时间和发育阶段特异性模式分泌。作为在体内定义鸡基质金属蛋白酶-1基因的组织和位点特异性调控区域的第一步，我们构建了携带与LacZ报告基因融合的各种启动子和内含子片段的转基因小鼠。对转基因小鼠个体发育过程中转基因表达模式的组织学分析显示，携带-2011至+67之间的长启动子或-338至+1819之间带有短启动子的内含子片段的转基因小鼠品系的软骨内骨的大多数原始元素中出现了特异性的X-gal染色。然而，后一种转基因的软骨特异性活性伴随着神经和其他组织中可变的异位表达模式，这取决于整合位点。启动子上游和内含子元件的存在对于所有软骨生成组织中的高水平转基因活性以及对于与鸡基质金属蛋白酶-1基因最相似的骨骼外转基因表达模式（例如在眼睛中的表达以及在逐渐退化的脊索和髓核中缺乏表达）是必要的。转基因的活性分别局限于通过BrdU掺入和磷酸化Sox9分布可视化的柱状增殖和前肥大软骨细胞。-2011至-338之间的DNA元件也介导了神经嵴来源细胞中的异位LacZ表达。这些结果表明，模块化排列的软骨和神经嵴特异性DNA元件之间的相互作用控制了基质金属蛋白酶-1基因的表达。启动子上游和内含子区域中软骨特异性元件的分散与Col11a2基因的转录调控相似。
