Hester Ian, McKee Sarah, Pelletier Phillip, Thompson Charles, Storbeck Christopher, Mears Alan, Schulz Jörg B, Hakim Antoine A, Sabourin Luc A
University of Ottawa, Department of Cellular and Molecular Medicine, Ottawa, ON, Canada.
Brain Res. 2007 Mar 2;1135(1):1-11. doi: 10.1016/j.brainres.2006.11.083. Epub 2007 Jan 9.
Cortical spreading depression (CSD) induces waves of neuronal depolarization that confer neuroprotection to subsequent ischemic events in the rat brain. To gain insights into the molecular mechanisms elicited by CSD, we used representational difference analysis (RDA) to identify mRNAs induced by potassium depolarization in vivo. Using this approach, we have isolated a cDNA encoding the SIM2-related bHLH-PAS protein Nxf. Our results confirm that Nxf mRNA and protein are rapidly and transiently expressed in cortical neurons following CSD. Reporter assays show that Nxf is a transcriptional activator that associates with the bHLH-PAS sub-class co-factor ARNT2. Adenovirus-mediated expression of epitope-tagged Nxf results in cell death and the direct activation of the Bax gene in cultured cells. However, RNA interference studies show that endogenous Nxf is required for optimal neuroprotection by preconditioning in cultured F-11 cells. Together, our data indicate that Nxf is a novel bHLH-PAS transactivator transiently induced by preconditioning and that its sustained expression is detrimental. The identification of Nxf may represent an important step in our understanding of the molecular mechanisms of brain preconditioning and injury.
皮层扩散性抑制（CSD）会引发神经元去极化波，这种去极化波能为大鼠大脑后续的缺血事件提供神经保护作用。为深入了解CSD引发的分子机制，我们运用代表性差异分析（RDA）来鉴定体内钾离子去极化诱导产生的信使核糖核酸（mRNA）。通过这种方法，我们分离出了一个编码与SIM2相关的碱性螺旋-环-螺旋-芳香烃受体蛋白（bHLH-PAS蛋白）Nxf的互补脱氧核糖核酸（cDNA）。我们的研究结果证实，在CSD后，Nxf信使核糖核酸和蛋白在皮层神经元中迅速且短暂地表达。报告基因检测表明，Nxf是一种转录激活因子，它与bHLH-PAS亚类辅因子芳香烃受体核转运蛋白2（ARNT2）相互作用。腺病毒介导的表位标记Nxf的表达会导致细胞死亡，并在培养细胞中直接激活Bax基因。然而，RNA干扰研究表明，内源性Nxf对于培养的F-11细胞预处理产生的最佳神经保护作用是必需的。综合来看，我们的数据表明，Nxf是一种由预处理短暂诱导产生的新型bHLH-PAS转录激活因子，其持续表达是有害的。Nxf的鉴定可能是我们理解大脑预处理和损伤分子机制的重要一步。
