Adeleke-Stainback P, Chen E, Collier P, Yuan Z A, Piddington R, Decker S, Rosenbloom J, Gibson C W
Craniofacial Anomalies Research Center, University of Pennsylvania School of Dental Medicine, Philadelphia 19104, USA.
Connect Tissue Res. 1995;32(1-4):115-8. doi: 10.3109/03008209509013712.
The amelogenin proteins, which are crucial for normal enamel mineral formation, are secreted by ameloblasts during development of tooth enamel. In order to better understand the mechanisms involved in regulation of expression of the amelogenin genes, the bovine X-chromosomal amelogenin gene was cloned and a 3.5 KB fragment upstream of exon 1 was inserted into a beta galactosidase (beta gal) expression vector for production of transgenic mice. When tissues from these mice were treated with Xgal, a substrate for beta gal, only ameloblasts and some of the adjacent stratum intermedium cells contained blue stain. To obtain further information concerning regulation of expression, the 3.5 KB amelogenin gene fragment was evaluated in transfection experiments. Nonoverlapping 1.9 and 1.5 KB fragments of the upstream region were subcloned separately into a vector that contains the SV40 promoter and the CAT reporter gene. Each amelogenin gene fragment was able to suppress CAT activity driven by the heterologous SV40 promoter in transfected HeLa cells. We theorize that each of these gene fragments contains regulatory elements important for the tissue-specific and developmentally-regulated pattern of expression of the X-chromosomal amelogenin gene.
釉原蛋白对于正常牙釉质矿化形成至关重要，在牙釉质发育过程中由成釉细胞分泌。为了更好地理解釉原蛋白基因表达调控所涉及的机制，克隆了牛X染色体上的釉原蛋白基因，并将外显子1上游的一个3.5KB片段插入β-半乳糖苷酶（β-gal）表达载体中，用于生产转基因小鼠。当用β-gal的底物Xgal处理这些小鼠的组织时，只有成釉细胞和一些相邻的中间层细胞含有蓝色染色。为了获得有关表达调控的更多信息，在转染实验中对3.5KB的釉原蛋白基因片段进行了评估。将上游区域的非重叠1.9KB和1.5KB片段分别亚克隆到一个含有SV40启动子和CAT报告基因的载体中。每个釉原蛋白基因片段都能够在转染的HeLa细胞中抑制由异源SV40启动子驱动的CAT活性。我们推测这些基因片段中的每一个都包含对X染色体上釉原蛋白基因的组织特异性和发育调控表达模式很重要的调控元件。
