Division of Pulmonary, Allergy and Critical Care Medicine, University of Pittsburgh Asthma Institute, University of Pittsburgh Medical Center/University of Pittsburgh School of Medicine, Pittsburgh, PA 15213, USA.
Proc Natl Acad Sci U S A. 2011 Aug 23;108(34):14246-51. doi: 10.1073/pnas.1018075108. Epub 2011 Aug 9.
Epithelial 15-lipoxygenase 1 (15LO1) and activated ERK are increased in asthma despite modest elevations in IL-13. MAPK kinase (MEK)/ERK activation is regulated by interactions of Raf-1 with phosphatidylethanolamine-binding protein 1 (PEBP1). Epithelial 15LO1 generates intracellular 15-hydroxyeicosatetraenoic acid (15HETE) conjugated to phosphatidylethanolamine (PE) (15HETE-PE). We hypothesized that (i) 15LO1 and its product 15HETE-PE serve as signaling molecules interacting with PEBP1 to activate Raf-1/MEK/ERK and that (ii) this 15LO1-15HETE-PE-regulated ERK activation amplifies IL-4Rα downstream pathways. Our results demonstrate that high epithelial 15LO1 levels correlate with ERK phosphorylation ex vivo. In vitro, IL-13 induces 15LO1, which preferentially binds to PEBP1, causing PEBP1 to dissociate from Raf-1 and activate ERK. Exogenous 15HETE-PE similarly induces dissociation of PEBP1 from Raf-1 independently of IL-13/15LO1. siRNA knockdown of 15LO1 decreases the dissociation of Raf-1 from PEBP1, and the resulting lower ERK activation leads to lower downstream IL-4Rα-related gene expression. Identical protein-protein interactions are observed in endobronchial biopsies and fresh epithelial cells from asthmatics ex vivo. Colocalization of Raf-1 to PEBP1 is low in asthmatic tissue and cells compared with normals, whereas there is striking colocalization of 15LO1 with PEBP1 in asthma. Low 15LO1 levels in normals limit its colocalization with PEBP1. The results confirm a previously unknown signaling role for 15LO1 and its PE-conjugated eicosanoid product in human airway epithelial cells. This pathway enhances critical inflammatory pathways integral to asthma pathogenesis.
上皮细胞 15-脂氧合酶 1（15LO1）和激活的 ERK 在哮喘中增加，尽管白细胞介素-13（IL-13）略有升高。MAPK 激酶（MEK）/ERK 激活受 Raf-1 与磷脂乙醇胺结合蛋白 1（PEBP1）相互作用的调节。上皮细胞 15LO1 生成与磷脂乙醇胺（PE）结合的细胞内 15-羟二十碳四烯酸（15HETE）（15HETE-PE）。我们假设（i）15LO1 及其产物 15HETE-PE 作为信号分子与 PEBP1 相互作用以激活 Raf-1/MEK/ERK，并且（ii）这种 15LO1-15HETE-PE 调节的 ERK 激活放大了 IL-4Rα 下游途径。我们的结果表明，高上皮细胞 15LO1 水平与 ERK 磷酸化在体外相关。在体外，IL-13 诱导 15LO1，其优先与 PEBP1 结合，导致 PEBP1 与 Raf-1 分离并激活 ERK。外源性 15HETE-PE 也独立于 IL-13/15LO1 诱导 PEBP1 与 Raf-1 分离。15LO1 的 siRNA 敲低降低了 Raf-1 与 PEBP1 的解离，由此导致的较低 ERK 激活导致较低的下游 IL-4Rα 相关基因表达。在哮喘患者的支气管活检和新鲜上皮细胞中观察到相同的蛋白质-蛋白质相互作用。与正常组织相比，哮喘组织和细胞中 Raf-1 与 PEBP1 的共定位较低，而哮喘中 15LO1 与 PEBP1 的共定位则很明显。正常情况下 15LO1 水平较低限制了其与 PEBP1 的共定位。结果证实了 15LO1 及其 PE 结合的类二十烷酸产物在人呼吸道上皮细胞中以前未知的信号作用。该途径增强了哮喘发病机制中关键的炎症途径。
