15(S)-lipoxygenase-2 mediates arachidonic acid-stimulated adhesion of human breast carcinoma cells through the activation of TAK1, MKK6, and p38 MAPK
15(S)-lipoxygenase-2 mediates arachidonic acid-stimulated adhesion of human breast carcinoma cells through the activation of TAK1, MKK6, and p38 MAPK
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DOI：
10.1074/jbc.m500418200
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发表时间：
2005-09-09
影响因子：
4.8
通讯作者：
Roberts, JD
中科院分区：
文献类型：
--
作者：
Nony, PA;Kennett, SB;Roberts, JD
饮食顺式多不饱和脂肪酸，花生四烯酸，刺激转移性人乳腺癌细胞（MDA-MB-435）的细胞外基质的粘附，但脂肪酸改变这些细胞的行为的分子机制尚不清楚。暴露于花生四烯酸激活多种信号通路。激活p38丝裂原活化蛋白激酶（p38 MAPK）是增加细胞粘附IV型胶原所必需的，这种激活对脂氧合酶抑制剂敏感，表明需要花生四烯酸代谢。本研究的目的是鉴定一种或多种负责p38 MAPK激活的花生四烯酸的关键代谢物，并阐明导致p38 MAPK激活的上游激酶。高效液相色谱分析表明，MDA-MB-435细胞主要代谢外源花生四烯酸为15（S）-羟基二十碳四烯酸（15（S）-HETE）。用对15（S）-脂氧合酶-1（LOX-1）和15（S）-脂氧合酶-2（LOX-2）特异的抗体进行的免疫印迹分析证实了在这些肿瘤细胞中15-LOX-2而非15-LOX-1的表达。脂氧合酶抑制剂去甲二氢愈创木酸可减弱花生四烯酸诱导的15（S）HETE的产生，并抑制p38 MAPK的磷酸化。相反，LOX-2的过表达使细胞对花生四烯酸的加入敏感，导致p38 MAPK的活化增加。向MDA-MB-435细胞中添加外源性15（S）-HETE刺激细胞与IV型胶原的粘附并激活p38 MAPK途径，包括上游激酶转化生长因子-β（1）-活化蛋白激酶-1（TAK 1）和MAPK激酶6。用显性阴性形式的TAK 1转染这些细胞阻断花生四烯酸刺激的p38 MAPK磷酸化。这些数据表明，15（S）HETE的15（S）-LOX-2生成激活特异性生长因子受体相关的信号传导途径，从而引发信号转导事件，导致细胞与细胞外基质的粘附增加。
The dietary cis-polyunsaturated fatty acid, arachidonic acid, stimulates adhesion of metastatic human breast carcinoma cells (MDA-MB-435) to the extracellular matrix, but the molecular mechanisms by which fatty acids modify the behavior of these cells are unclear. Exposure to arachidonic acid activates multiple signaling pathways. Activation of p38 mitogen-activated protein kinase (p38 MAPK) is required for increased cell adhesion to type IV collagen, and this activation is sensitive to inhibitors of lipoxygenases, suggesting a requirement for arachidonic acid metabolism. The goals of the current study were to identify the one or more key metabolites of arachidonic acid that are responsible for activation of p38 MAPK and to elucidate the upstream kinases that lead to p38 MAPK activation. High performance liquid chromatographic analysis revealed that MDA-MB-435 cells metabolize exogenous arachidonic acid predominantly to 15(S)-hydroxyeicosatetraenoic acid (15(S)-HETE). Immunoblot analysis with antibodies specific to 15(S)-lipoxygenase-1 (LOX-1) and 15(S)-lipoxygenase-2 (LOX-2) demonstrated the expression of 15-LOX-2, but not 15-LOX-1, in these tumor cells. A LOX inhibitor, nordihydroguaiaretic acid, attenuated production of 15(S)HETE and inhibited the phosphorylation of p38 MAPK following exposure to arachidonic acid. In contrast, overexpression of LOX-2 sensitized the cells to the addition of arachidonic acid, leading to increased activation of p38 MAPK. Addition of exogenous 15(S)-HETE to MDA-MB-435 cells stimulated cell adhesion to type IV collagen and activated the p38 MAPK pathway, including the upstream kinases transforming growth factor-beta(1)-activated protein kinase-1 (TAK1) and MAPK kinase 6. Transfection of these cells with a dominant negative form of TAK1 blocked arachidonic acid-stimulated p38 MAPK phosphorylation. These data demonstrate that 15(S)-LOX-2 generation of 15(S)HETE activates specific growth factor receptor-related signaling pathways, thereby initiating signal transduction events leading to increased cell adhesion to the extracellular matrix.