Kankowski Svenja, Förstera Benjamin, Winkelmann Aline, Knauff Pina, Wanker Erich E, You Xintian A, Semtner Marcus, Hetsch Florian, Meier Jochen C
Division Cell Physiology, Zoological Institute, Technische Universität Braunschweig, Braunschweig, Germany.
Institute for Stroke and Dementia Research, Klinikum der Universität München, Ludwig Maximilians University of Munich, Munich, Germany.
Front Mol Neurosci. 2018 Jan 11;10:439. doi: 10.3389/fnmol.2017.00439. eCollection 2017.
C-to-U RNA editing of glycine receptors (GlyR) can play an important role in disease progression of temporal lobe epilepsy (TLE) as it may contribute in a neuron type-specific way to neuropsychiatric symptoms of the disease. It is therefore necessary to develop tools that allow identification of neuron types that express RNA-edited GlyR protein. In this study, we identify NH as agonist of C-to-U RNA edited GlyRs. Furthermore, we generated a new molecular C-to-U RNA editing sensor tool that detects Apobec-1- dependent RNA editing in HEPG2 cells and rat primary hippocampal neurons. Using this sensor combined with NH application, we were able to identify C-to-U RNA editing-competent neurons and expression of C-to-U RNA-edited GlyR protein in neurons. Bioinformatic analysis of 1,000 Genome Project Phase 3 allele frequencies coding for human Apobec-1 80M and 80I variants showed differences between populations, and the results revealed a preference of the 80I variant to generate RNA-edited GlyR protein. Finally, we established a new PCR-based restriction fragment length polymorphism (RFLP) approach to profile mRNA expression with regard to the genetic dimorphism of patients with intractable temporal lobe epilepsy (iTLE) and found that the patients fall into two groups. Patients with expression of the Apobec-1 80I variant mostly suffered from simple or complex partial seizures, whereas patients with 80M expression exhibited secondarily generalized seizure activity. Thus, our method allows the characterization of Apobec-1 80M and 80l variants in the brain and provides a new way to epidemiologically and semiologically classify iTLE according to the two different alleles. Together, these results demonstrate Apobec-1-dependent expression of RNA-edited GlyR protein in neurons and identify the 80I/M-coding alleles as new genetic risk factors for iTLE patients.
甘氨酸受体（GlyR）的C-to-U RNA编辑在颞叶癫痫（TLE）的疾病进展中可能发挥重要作用，因为它可能以神经元类型特异性的方式导致该疾病的神经精神症状。因此，有必要开发能够识别表达RNA编辑的GlyR蛋白的神经元类型的工具。在本研究中，我们鉴定出NH作为C-to-U RNA编辑的GlyRs的激动剂。此外，我们生成了一种新的分子C-to-U RNA编辑传感器工具，该工具可检测HEPG2细胞和大鼠原代海马神经元中载脂蛋白B mRNA编辑酶1（Apobec-1）依赖性RNA编辑。使用该传感器并结合NH应用，我们能够识别具有C-to-U RNA编辑能力的神经元以及神经元中C-to-U RNA编辑的GlyR蛋白的表达。对千人基因组计划第三阶段编码人类Apobec-1 80M和80I变体的等位基因频率进行的生物信息学分析显示了不同人群之间的差异，结果表明80I变体更倾向于生成RNA编辑的GlyR蛋白。最后，我们建立了一种基于聚合酶链反应（PCR）的限制性片段长度多态性（RFLP）方法，以分析难治性颞叶癫痫（iTLE）患者基因二态性的mRNA表达情况，发现患者分为两组。表达Apobec-1 80I变体的患者大多患有简单或复杂部分性发作，而表达80M的患者表现为继发性全身性发作活动。因此，我们的方法能够对大脑中的Apobec-1 80M和80I变体进行表征，并提供了一种根据两种不同等位基因对iTLE进行流行病学和症状学分类的新方法。总之，这些结果证明了神经元中Apobec-1依赖性RNA编辑的GlyR蛋白表达，并将80I/M编码等位基因鉴定为难治性颞叶癫痫患者的新遗传风险因素。
