RAS/ERK modulates TGFβ-regulated PTEN expression in human pancreatic adenocarcinoma cells
RAS/ERK modulates TGFβ-regulated PTEN expression in human pancreatic adenocarcinoma cells
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DOI：
10.1093/carcin/bgm159
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发表时间：
2007-11-01
期刊：
影响因子：
4.7
通讯作者：
Carethers, John M.
中科院分区：
文献类型：
--
作者：
Chow, Jimmy Y. C.;Quach, Khai T.;Carethers, John M.
10号染色体上缺失的磷酸酶和张力蛋白同源基因(PTEN)在胰腺癌中很少发生突变，但其受转化生长因子(TGF)-β的调节可能介导了生长抑制和其他致癌作用。在这里，我们研究了在Smad4缺失的情况下，转化生长因子β的作用以及致癌K-RAS/ERK对PTEN表达的影响。我们利用了两个Smad4缺失的胰腺细胞系Capan-1(K-Ras突变体)和BxPc-3(WT-K-RAS)，这两个细胞株都表达转化生长因子β受体。用转化生长因子β1处理细胞，分离成胞浆/胞核部分，用磷酸化Smad2、4磷酸化三磷酸腺苷依赖的酪氨酸激酶(Akt)、Akt和PTEN抗体进行免疫印迹。逆转录-聚合酶链式反应检测PTEN基因表达水平。MEK1抑制剂PD98059被用来阻断致癌K-RAS/ERK的下游作用，这是一个显性-阴性(DN)K-RAS结构。转化生长因子β可增加胞浆和胞核中的磷酸化Smad2。PD98059处理进一步增加了两种胰腺细胞系细胞核中磷酸化Smad2的表达，而dN-K-RAS进一步改善了K-RAS突变型CAPAN细胞中SMAD的易位。在48h时，转化生长因子β可显著抑制伴随Akt活化的PTEN蛋白水平，这种抑制作用在6h时表现明显，PD98059和dN-K-RAS可逆转转化生长因子β诱导的PTEN抑制作用。转化生长因子β诱导的PTEN表达与细胞增殖呈负相关。因此，胰腺癌中致癌的K-RAS/ERK以Smad4非依赖的方式促进转化生长因子β诱导的肿瘤抑制基因PTEN的转录下调，并可能构成胰腺癌从生长抑制到生长促进的信号转换机制。
Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is rarely mutated in pancreatic cancers, but its regulation by transforming growth factor (TGF)-beta might mediate growth suppression and other oncogenic actions. Here, we examined the role of TGF beta and the effects of oncogenic K-RAS/ERK upon PTEN expression in the absence of SMAD4. We utilized two SMAD4-null pancreatic cell lines, CAPAN-1 (K-RAS mutant) and BxPc-3 (WT-K-RAS), both of which express TGF beta surface receptors. Cells were treated with TGF beta 1 and separated into cytosolic/nuclear fractions for western blotting with phospho-SMAD2, SMAD 2, 4 phospho-ATP-dependent tyrosine kinases (Akt), Akt and PTEN antibodies. PTEN mRNA levels were assessed by reverse transcriptase-polymerase chain reaction. The MEK1 inhibitor, PD98059, was used to block the downstream action of oncogenic K-RAS/ERK, as was a dominant-negative (DN) K-RAS construct. TGF beta increased phospho-SMAD2 in both cytosolic and nuclear fractions. PD98059 treatment further increased phospho-SMAD2 in the nucleus of both pancreatic cell lines, and DN-K-RAS further improved SMAD translocation in K-RAS mutant CAPAN cells. TGF beta treatment significantly suppressed PTEN protein levels concomitant with activation of Akt by 48 h through transcriptional reduction of PTEN mRNA that was evident by 6 h. TGF beta-induced PTEN suppression was reversed by PD98059 and DN-K-RAS compared with treatments without TGF beta. TGF beta-induced PTEN expression was inversely related to cellular proliferation. Thus, oncogenic K-RAS/ERK in pancreatic adenocarcinoma facilitates TGF beta-induced transcriptional down-regulation of the tumor suppressor PTEN in a SMAD4-independent manner and could constitute a signaling switch mechanism from growth suppression to growth promotion in pancreatic cancers.