Green C J, Lichtlen P, Huynh N T, Yanovsky M, Laderoute K R, Schaffner W, Murphy B J
The Pharmaceutical Discovery Division, SRI International, Menlo Park, California 94025, USA.
Cancer Res. 2001 Mar 15;61(6):2696-703.
Placenta growth factor (PlGF) is a mitogen for endothelial cells that can potentiate the growth and permeabilizing effects on endothelium of vascular endothelial growth factor. Here we report that hypoxia induces the expression of both PlGF mRNA and protein in immortalized/transformed mouse embryonic fibroblasts (mEFs) and in NIH 3T3 cells. Importantly, the magnitude of the induction of PlGF expression by hypoxia is enhanced by the presence of oncogenic Ras. To investigate the transcriptional component of hypoxia-inducible PlGF expression, we cloned and sequenced a 1350-bp fragment of the 5'-flanking region of the mouse gene. Analysis of the promoter region indicated the presence of putative consensus sequences for known hypoxia-responsive regulatory sites, including metal response elements and Sp1-like sites. In the present study, we show that the induction of PlGF expression by hypoxia is dependent on the presence of the metal response element-binding transcription factor 1 (MTF-1). Thus, in mEFs with targeted deletions of both MTF-1 alleles, hypoxia-induced increases of PIGF mRNA and protein levels were greatly attenuated compared with those in wild-type mEFs. Moreover, transient transfection of a PlGF promoter reporter gene into NIH 3T3 cells resulted in hypoxia-responsive transcriptional activation of the reporter. Finally, ectopic expression of MTF-1 resulted in increased basal transcriptional activity of a PlGF promoter reporter. Together, these findings demonstrate that the PlGF gene is responsive to hypoxia and that this response is mediated by MTF-1. It remains to be determined whether this activation is the result of direct and/or indirect transcriptional activation by MTF-1. The stimulatory effect of oncogenic Ras on the induction of PlGF expression in hypoxic cells suggests that PlGF could be an important proangiogenic factor in the tumor microenvironment.
胎盘生长因子（PlGF）是一种内皮细胞有丝分裂原，可增强血管内皮生长因子对内皮的生长和通透作用。在此我们报告，缺氧可诱导永生化/转化的小鼠胚胎成纤维细胞（mEFs）和NIH 3T3细胞中PlGF mRNA和蛋白的表达。重要的是，致癌性Ras的存在增强了缺氧诱导的PlGF表达程度。为了研究缺氧诱导的PlGF表达的转录成分，我们克隆并测序了小鼠基因5'侧翼区的一个1350 bp片段。对启动子区域的分析表明存在已知缺氧反应调节位点的推定共有序列，包括金属反应元件和Sp1样位点。在本研究中，我们表明缺氧诱导的PlGF表达依赖于金属反应元件结合转录因子1（MTF-1）的存在。因此，在两个MTF-1等位基因均有靶向缺失的mEFs中，与野生型mEFs相比，缺氧诱导的PIGF mRNA和蛋白水平的增加大大减弱。此外，将PlGF启动子报告基因瞬时转染到NIH 3T3细胞中导致报告基因的缺氧反应性转录激活。最后，MTF-1的异位表达导致PlGF启动子报告基因的基础转录活性增加。总之，这些发现表明PlGF基因对缺氧有反应，且这种反应由MTF-1介导。这种激活是否是MTF-1直接和/或间接转录激活的结果仍有待确定。致癌性Ras对缺氧细胞中PlGF表达诱导的刺激作用表明PlGF可能是肿瘤微环境中一种重要的促血管生成因子。
