Overexpression of human kynurenine-3-monooxygenase protects against 3-hydroxykynurenine-mediated apoptosis through bidirectional nonlinear feedback
Overexpression of human kynurenine-3-monooxygenase protects against 3-hydroxykynurenine-mediated apoptosis through bidirectional nonlinear feedback
复制标题
人犬尿氨酸-3-单加氧酶的过度表达通过双向非线性反馈防止 3-羟基犬尿氨酸介导的细胞凋亡
作者：
Kris Wilson;M. Auer;M. Binnie;Xiaozhong Zheng;N. Pham;J. Iredale;S. Webster;D. Mole
犬尿氨酸3-单加氧酶（KMO）是炎症的关键调节剂。优选的KMO底物犬尿氨酸转化为3-羟基犬尿氨酸（3 HK），该产物通过最终导致细胞凋亡的机制表现出细胞毒性。在这里，我们报告说，过表达的人KMO与原位定位到线粒体创建一个代谢环境，在此期间，细胞表现出增加的耐受性外源性3 HK介导的细胞损伤。使用选择性KMO抑制剂Ro 61 -8048，我们表明，KMO酶的功能是必不可少的细胞保护。用Z-VAD-FMK抑制泛半胱天冬酶证实细胞凋亡是细胞死亡的模式。通过定义KMO上游和下游通路组分的表达，我们观察到其他关键犬尿氨酸通路组分的改变，特别是通过双向非线性反馈的色氨酸-2，3-双加氧酶上调。KMO过表达也增加诱导型一氧化氮合酶（iNOS）的表达。基因表达的这些变化在功能上是相关的，因为途径组分犬尿氨酸酶和喹啉磷酸核糖转移酶的siRNA敲低导致细胞恢复到对3 HK介导的细胞凋亡敏感的状态。总之，KMO过表达，重要的是KMO活性，具有从根本上影响细胞应激抗性的代谢影响。
Kynurenine 3-monooxygenase (KMO) is a critical regulator of inflammation. The preferred KMO substrate, kynurenine, is converted to 3-hydroxykynurenine (3HK), and this product exhibits cytotoxicity through mechanisms that culminate in apoptosis. Here, we report that overexpression of human KMO with orthotopic localisation to mitochondria creates a metabolic environment during which the cell exhibits increased tolerance for exogenous 3HK-mediated cellular injury. Using the selective KMO inhibitor Ro61-8048, we show that KMO enzyme function is essential for cellular protection. Pan-caspase inhibition with Z-VAD-FMK confirmed apoptosis as the mode of cell death. By defining expression of pathway components upstream and downstream of KMO, we observed alterations in other key kynurenine pathway components, particularly tryptophan-2,3-dioxygenase upregulation, through bidirectional nonlinear feedback. KMO overexpression also increased expression of inducible nitric oxide synthase (iNOS). These changes in gene expression are functionally relevant, because siRNA knockdown of the pathway components kynureninase and quinolinate phosphoribosyl transferase caused cells to revert to a state of susceptibility to 3HK-mediated apoptosis. In summary, KMO overexpression, and importantly KMO activity, have metabolic repercussions that fundamentally affect resistance to cell stress.