Lee T X, Johnson L F
Department of Molecular Genetics, The Ohio State University, Columbus 43210, USA.
J Cell Biochem. 1998 May 1;69(2):104-16.
The pre-mRNA processing enhancer (PPE) element is an RNA sequence element derived from the intronless HSV-TK gene. Insertion of the element into the highly intron-dependent human beta-globin gene leads to efficient expression in the absence of splicing. We have analyzed the effect of the PPE element on the expression of mouse thymidylate synthase (TS) minigenes. We have previously shown that the expression of intronless TS minigenes is moderately (up to 20-fold) stimulated by the inclusion of introns. Furthermore, S phase-specific expression of TS minigenes in growth-stimulated cells depends on the presence of a spliceable intron as well as the TS promoter. The goal of our study was to determine if the PPE element would overcome the dependence on introns for efficient expression and for S phase-specific expression of transfected TS minigenes. We found that insertion of the PPE element into an intronless TS minigene partially overcame intron dependence. However, the increase in expression was much less than that observed for the intronless beta-globin gene. We also found that intronless TS or HSV-TK genes that contained the PPE element and that were driven by the TS promoter were expressed at a constant level in serum-stimulated cells. However, when an intron was included in these genes, they were expressed in an S phase-specific manner. Thus the PPE element was not able to overcome the dependence on introns for S phase-specific expression of TS minigenes.
前体mRNA加工增强子（PPE）元件是一种源自无内含子单纯疱疹病毒胸苷激酶（HSV-TK）基因的RNA序列元件。将该元件插入高度依赖内含子的人β-珠蛋白基因中，可在无剪接的情况下实现高效表达。我们分析了PPE元件对小鼠胸苷酸合成酶（TS）小基因表达的影响。我们之前已经表明，无内含子TS小基因的表达通过内含子的加入受到适度（高达20倍）的刺激。此外，TS小基因在生长刺激细胞中的S期特异性表达取决于可剪接内含子的存在以及TS启动子。我们研究的目的是确定PPE元件是否能克服对内含子的依赖性，以实现转染的TS小基因的高效表达和S期特异性表达。我们发现，将PPE元件插入无内含子TS小基因中可部分克服对内含子的依赖性。然而，表达的增加远低于无内含子β-珠蛋白基因所观察到的情况。我们还发现，含有PPE元件且由TS启动子驱动的无内含子TS或HSV-TK基因在血清刺激的细胞中以恒定水平表达。然而，当这些基因中包含一个内含子时，它们以S期特异性方式表达。因此，PPE元件无法克服对内含子的依赖性，以实现TS小基因的S期特异性表达。
