Cloning and characterization of the proximal murine Phex promoter
Cloning and characterization of the proximal murine Phex promoter
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DOI：
10.1210/en.142.9.3987
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发表时间：
2001-09-01
期刊：
影响因子：
4.8
通讯作者：
Quarles, LD
中科院分区：
文献类型：
--
作者：
Liu, SG;Guo, R;Quarles, LD
Phex是一种调节系统磷酸盐稳态的内肽酶。我们通过克隆小鼠Phex基因5 '侧翼区2736 bp的启动子并进行功能分析，研究了Phex基因的转录。我们确定了一个转录起始位点，一个共识TATA盒，和多个潜在的顺式作用调节元件。为了确定启动子是否指导细胞类型限制的Phex表达，我们将全长和5 '缺失的Phex荧光素酶报告基因构建体转染到各种细胞系中。与非Phex表达COS-7细胞相比，Phex表达C5.18软骨细胞显示出转染的Phex启动子构建体的最高活性，而启动子活性在ROS 17/2.8成骨细胞中是中等的，并且在MC 3 T3-E1成骨细胞中是成熟阶段依赖性的。对ROS 17/2.8细胞中Phex启动子的连续5 ' -缺失突变体的分析揭示了双峰活性，表明可能存在正和负顺式作用区。软骨形成因子SOX 9显著刺激Phex启动子活性，而Cbfa 1、PTH和1，25（OH）（2）D-3没有影响。我们的发现与Phex在骨和软骨中的主要表达一致。需要进一步的研究来确认Phex启动子中以细胞限制性方式发挥功能的调控区域。
Phex is an endopetidase that regulates systemic phosphate homeostasis. We investigated Phex gene transcription by cloning and performing functional analysis of the 2736 bp of the 5 ' flanking region of the mouse Phex gene containing its promoter. We identified a transcription start site, a consensus TATA-box, and multiple potential cis-acting regulator elements. To determine whether the promoter directs cell-type restricted Phex expression, we transfected full-length and 5 ' -deleted Phex luciferase reporter constructs into various cell lines. Phex-expressing C5.18 chondrocytes displayed the highest activity of the transfected Phex promoter constructs compared with non-Phex-expressing COS-7 cells, whereas promoter activity was intermediate in ROS 17/2.8 osteoblasts and maturation stage-dependent in MC3T3-E1 osteoblasts. Analysis of sequential 5 ' -deletion mutants of the Phex promoter in ROS 17/2.8 cells revealed bimodal activity, suggesting that both positive and negative cis-acting regions may be present. The chondrogenic factor SOX9 markedly stimulated Phex promoter activity, whereas Cbfa1, PTH, and 1,25(OH)(2)D-3 had no effect. Our findings are consistent with the predominant expression of Phex in bone and cartilage. Additional studies will be needed to confirm the regulatory regions in the Phex promoter that function in a cell-restricted manner.