Amoui Mehran, Baylink David J, Tillman John B, Lau K-H William
Musculoskeletal Disease Center, Jerry L. Pettis Memorial Veterans Affairs Medical Center, Loma Linda University, Loma Linda, California 92357, USA.
J Biol Chem. 2003 Nov 7;278(45):44273-80. doi: 10.1074/jbc.M303933200. Epub 2003 Aug 29.
An osteoclastic protein-tyrosine phosphatase (PTP-oc), essential for osteoclast activity, shows sequence identity with the intracellular domain of GLEPP1, a renal receptor-like transmembrane PTP. PTP-oc has been assumed to be a truncated variant of GLEPP1, resulting from alternative splicing. However, the 5'-untranslated region sequence of PTP-oc mRNA contains 217 bp from an intron of GLEPP1. There are no splicing acceptor sites at the PTP-oc transcription site. The intronic sequence flanking the 5' end of the PTP-oc transcription start site contains potential promoter elements essential for transcriptional initiation. To test the hypothesis that the PTP-oc gene has an alternative, tissue-specific, intronic promoter, the promoter activity of a 1.3-kb PCR fragment covering the 5'-flanking region of the PTP-oc gene was measured. The putative PTP-oc promoter fragment showed strong promoter activity in U937 cells. Mutation of the putative TATA box within the PTP-oc promoter abolished 60-90% of its promoter activity. The PTP-oc promoter fragment showed strong promoter activity in cells that express PTP-oc (U937 cells and RAW264.7 cells) but not in cells that do not express the enzyme (skin fibroblasts, TE85 cells, and HEK293 cells). These findings strongly support the conclusion that the 1.3-kb intronic fragment contains the tissue-specific, PTP-oc proximal promoter. Deletion and functional analyses indicate that the proximal 5' sequence flanking the TATA box of the PTP-oc contains potential repressor elements. The removal of the putative repressor elements led to the apparent loss of tissue specificity. In summary, we conclude that an intronic promoter within the GLEPP1 gene drives the expression of the PTP-oc in a cell type-specific manner. This GLEPP1/PTP-oc gene system is one of the very few systems in which two important tissue-specific enzymes are derived from the same gene by the use of alternative intronic promoters.
破骨细胞蛋白酪氨酸磷酸酶（PTP-oc）对破骨细胞活性至关重要，它与肾受体样跨膜蛋白酪氨酸磷酸酶GLEPP1的胞内结构域具有序列同源性。PTP-oc被认为是GLEPP1的截短变体，由可变剪接产生。然而，PTP-oc mRNA的5'-非翻译区序列包含来自GLEPP1一个内含子的217 bp。在PTP-oc转录位点没有剪接受体位点。PTP-oc转录起始位点5'端侧翼的内含子序列包含转录起始所必需的潜在启动子元件。为了验证PTP-oc基因有一个可变的、组织特异性的内含子启动子这一假说，测定了覆盖PTP-oc基因5'-侧翼区的1.3 kb PCR片段的启动子活性。推测的PTP-oc启动子片段在U937细胞中显示出强启动子活性。PTP-oc启动子内推测的TATA框突变使其启动子活性丧失60 - 90%。PTP-oc启动子片段在表达PTP-oc的细胞（U937细胞和RAW264.7细胞）中显示出强启动子活性，但在不表达该酶的细胞（皮肤成纤维细胞、TE85细胞和HEK293细胞）中则没有。这些发现有力地支持了以下结论：1.3 kb内含子片段包含组织特异性的PTP-oc近端启动子。缺失和功能分析表明，PTP-oc的TATA框侧翼的近端5'序列包含潜在的抑制元件。去除推测的抑制元件导致组织特异性明显丧失。总之，我们得出结论，GLEPP1基因内的一个内含子启动子以细胞类型特异性方式驱动PTP-oc的表达。这种GLEPP1/PTP-oc基因系统是极少数通过使用可变内含子启动子从同一基因衍生出两种重要组织特异性酶的系统之一。
