Batarseh Amani, Giatzakis Christoforos, Papadopoulos Vassilios
Department of Biochemistry & Molecular and Cell Biology, Georgetown University Medical Center, Washington, DC 20057, USA.
Biochemistry. 2008 Dec 2;47(48):12886-99. doi: 10.1021/bi8012643.
Translocator protein (TSPO) is an 18-kDa cholesterol-binding protein that is expressed at high levels in steroid synthesizing and several cancer cells where it is involved in steroidogenesis and cell proliferation, respectively. The factors regulating Tspo expression are unknown. We analyzed Tspo transcriptional responses to the tumor promoter, phorbol-12-myristate 13-acetate (PMA), in cells with varying TSPO levels. PMA induced Tspo promoter activity and Tspo mRNA levels in TSPO-poor nonsteroidogenic cells (NIH-3T3 fibroblasts and COS-7 kidney) but not in TSPO-rich steroidogenic cells (MA-10 Leydig) with high basal Tspo transcriptional activity. The stimulatory effect of PMA was mediated by an 805-515-bp region upstream of the transcription start site. Electrophoretic mobility shift assay (EMSA) revealed that PMA induced binding of c-jun and GA-binding protein transcription factor (GABP-alpha) to their respective activator protein 1 (AP1) and v-ets erythroblastosis virus E26 oncogene homologue (Ets) sites in this region. Protein kinase C (PKC)-specific inhibitors blocked PMA induction of Tspo promoter activity with an inhibition profile suggestive of involvement of PKCepsilon. PKCepsilon expression correlated with TSPO content in the three cell lines. In NIH-3T3 cells, PKCepsilon overexpression induced Tspo promoter activity and mRNA levels and enhanced PMA-induced up regulation of c-jun and TSPO. In MA-10 cells, a PKCepsilon-specific translocation inhibitor peptide reduced basal Tspo promoter activity. PKCepsilon siRNA pool reduced PKCepsilon and TSPO levels in MA-10 cells indicating a role for PKCepsilon in regulating TSPO expression. Taken together, these data suggest that elevated TSPO expression in steroidogenic cells may be due to high constitutive expression of PKCepsilon that renders them unresponsive to further induction while PMA activation of PKCepsilon drives inducible TSPO expression in nonsteroidogenic cells, likely through AP1 and Ets.
转位蛋白（TSPO）是一种18 kDa的胆固醇结合蛋白，在类固醇合成细胞和几种癌细胞中高表达，分别参与类固醇生成和细胞增殖。调节Tspo表达的因素尚不清楚。我们分析了不同TSPO水平的细胞中Tspo对肿瘤启动子佛波醇-12-肉豆蔻酸酯13-乙酸酯（PMA）的转录反应。PMA诱导TSPO含量低的非类固醇生成细胞（NIH-3T3成纤维细胞和COS-7肾细胞）中Tspo启动子活性和Tspo mRNA水平，但不诱导基础Tspo转录活性高的TSPO含量高的类固醇生成细胞（MA-10睾丸间质细胞）中的上述指标。PMA的刺激作用由转录起始位点上游805 - 515 bp区域介导。电泳迁移率变动分析（EMSA）显示，PMA诱导c-jun和GA结合蛋白转录因子（GABP-α）与其在该区域各自的激活蛋白1（AP1）和v-ets成红细胞增多症病毒E26癌基因同源物（Ets）位点结合。蛋白激酶C（PKC）特异性抑制剂阻断PMA对Tspo启动子活性的诱导，其抑制谱提示PKCε参与其中。PKCε的表达与三种细胞系中的TSPO含量相关。在NIH-3T3细胞中，PKCε过表达诱导Tspo启动子活性和mRNA水平，并增强PMA诱导的c-jun和TSPO上调。在MA-10细胞中，一种PKCε特异性转位抑制剂肽降低基础Tspo启动子活性。PKCε siRNA池降低MA-10细胞中的PKCε和TSPO水平，表明PKCε在调节TSPO表达中起作用。综上所述，这些数据表明，类固醇生成细胞中TSPO表达升高可能是由于PKCε的高组成性表达，使其对进一步诱导无反应，而PMA激活PKCε可能通过AP1和Ets驱动非类固醇生成细胞中可诱导的TSPO表达。
