Wang Z Y, Masaharu N, Qiu Q Q, Takimoto Y, Deuel T F
Department of Medicine, Jewish Hospital of St Louis, MO 63110.
Nucleic Acids Res. 1994 Feb 11;22(3):457-64. doi: 10.1093/nar/22.3.457.
The platelet-derived growth factor (PDGF) A-chain gene is expressed in a tissue- and developmental stage-specific manner. Here we identify an S1 nuclease sensitive region within the first intron that functions as a negative regulatory element in HeLa but not in human glioblastoma (A172) cells in transient transfection assays. A 147 bp DNA fragment that contains this element functions in a position and orientation independent manner to negatively regulate both the PDGF A-chain promoter and the heterologous herpes simplex virus thymidine kinase (TK) promoter. The cell-type specific effect of this 147 bp DNA fragment is seen when it is located downstream but not upstream of the reporter gene driven by either the PDGF A-chain or TK promoters. The negative regulatory element has been localized to a 24 bp DNA sequence within the S1 sensitive site that retains negative regulatory activity and recognizes a nuclear protein in HeLa but not in A172 cells. Furthermore, the 24 bp element functions as a cell type-specific negative element independent of its position. These results suggest that a functional silencer within the first intron exhibits a non-B-form DNA structure under superhelical stress in vitro and may contribute to the cell type-specific transcriptional regulation of PDGF A-chain gene in vivo.
血小板衍生生长因子（PDGF）A链基因以组织和发育阶段特异性的方式表达。在此，我们在第一个内含子中鉴定出一个S1核酸酶敏感区域，在瞬时转染实验中，该区域在HeLa细胞中作为负调控元件发挥作用，但在人胶质母细胞瘤（A172）细胞中则不然。包含该元件的一个147 bp DNA片段以位置和方向独立的方式发挥作用，对PDGF A链启动子和异源单纯疱疹病毒胸苷激酶（TK）启动子均进行负调控。当这个147 bp DNA片段位于由PDGF A链或TK启动子驱动的报告基因的下游而非上游时，可观察到其对细胞类型的特异性影响。负调控元件已定位到S1敏感位点内的一个24 bp DNA序列，该序列保留负调控活性，并在HeLa细胞而非A172细胞中识别一种核蛋白。此外，24 bp元件作为一种细胞类型特异性负元件发挥作用，与其位置无关。这些结果表明，第一个内含子内的一个功能性沉默子在体外超螺旋应力下呈现非B型DNA结构，可能在体内对PDGF A链基因的细胞类型特异性转录调控起作用。
