Abstract：
OBJECTIVE: The RELN gene encodes the secreted glycoprotein Reelin and has important functions in both developing and adult brains. In this study, we aimed to explore the association between the RELN and genetic generalized epilepsy (GGE).
METHODS: We performed whole-exome sequencing on a cohort of 92 patients with GGE. Based on amino acid sequence alignments, allele frequency, pedigree validation and computational modeling, the RELN variants were identified and clinical features of cases were summarized. Cell-based Reelin secretion assays were examined by Western blotting. Alterations of mutant Reelin transport through the secretion pathway were detected by immunofluorescence staining.
RESULTS: Three novel pathogenic RELN variants (3.26%; c.2260C>T/p.R754W, c.2914C>G/p.P972A and c.3029G>A/p.R1010H) were identified. All probands showed adolescence-onset generalized seizures characterized by generalized epileptiform discharges with normal EEG backgrounds, no or mild cognitive impairment, and responded well to anti-seizure medications. All these variants were located in the central regions from 1B to 2A consecutive repeats, and protein modeling demonstrated structural alterations in Reelin. Moreover, we found that these heterozygous missense variants significantly decreased the secretion of mutant proteins in HEK-293T cells, and this impairment was due to the altered transport of mutant Reelin in the secretion pathway.
CONCLUSIONS: These results suggest that RELN is potentially associated with GGE. The phenotype of GGE caused by RELN variants is relatively mild, and the pathogenic mechanism may involve a loss-of-function.
METHODS: We performed whole-exome sequencing on a cohort of 92 patients with GGE. Based on amino acid sequence alignments, allele frequency, pedigree validation and computational modeling, the RELN variants were identified and clinical features of cases were summarized. Cell-based Reelin secretion assays were examined by Western blotting. Alterations of mutant Reelin transport through the secretion pathway were detected by immunofluorescence staining.
RESULTS: Three novel pathogenic RELN variants (3.26%; c.2260C>T/p.R754W, c.2914C>G/p.P972A and c.3029G>A/p.R1010H) were identified. All probands showed adolescence-onset generalized seizures characterized by generalized epileptiform discharges with normal EEG backgrounds, no or mild cognitive impairment, and responded well to anti-seizure medications. All these variants were located in the central regions from 1B to 2A consecutive repeats, and protein modeling demonstrated structural alterations in Reelin. Moreover, we found that these heterozygous missense variants significantly decreased the secretion of mutant proteins in HEK-293T cells, and this impairment was due to the altered transport of mutant Reelin in the secretion pathway.
CONCLUSIONS: These results suggest that RELN is potentially associated with GGE. The phenotype of GGE caused by RELN variants is relatively mild, and the pathogenic mechanism may involve a loss-of-function.
摘要：
目的：RELN基因编码分泌的糖蛋白Reelin，在发育和成年大脑中都具有重要功能。在这项研究中,我们旨在探讨RELN与遗传性全身性癫痫(GGE)之间的关联.
方法：我们对92例GGE患者进行了全外显子组测序。基于氨基酸序列比对，等位基因频率，谱系验证和计算建模，确定了RELN变异体,并总结了病例的临床特征.通过Western印迹检查基于细胞的Reelin分泌测定。通过免疫荧光染色检测突变体Reelin通过分泌途径转运的改变。
结果：三种新的致病性RELN变体（3.26％；c.2260C>T/p。R754W,c.2914C>G/p。P972A和c.3029G>A/p。R1010H)被鉴定。所有先证者均显示青春期发作的全身性癫痫发作，其特征是具有正常EEG背景的全身性癫痫样放电，无或轻度认知障碍，对抗癫痫药物反应良好。所有这些变体都位于从1B到2A连续重复的中心区域，和蛋白质模型证明了Reelin的结构改变。此外，我们发现这些杂合错义变体显著降低了HEK-293T细胞中突变蛋白的分泌,这种损害是由于突变Reelin在分泌途径中的转运改变。
结论：这些结果表明RELN可能与GGE相关。由RELN变体引起的GGE的表型相对温和，致病机制可能涉及功能丧失。
方法：我们对92例GGE患者进行了全外显子组测序。基于氨基酸序列比对，等位基因频率，谱系验证和计算建模，确定了RELN变异体,并总结了病例的临床特征.通过Western印迹检查基于细胞的Reelin分泌测定。通过免疫荧光染色检测突变体Reelin通过分泌途径转运的改变。
结果：三种新的致病性RELN变体（3.26％；c.2260C>T/p。R754W,c.2914C>G/p。P972A和c.3029G>A/p。R1010H)被鉴定。所有先证者均显示青春期发作的全身性癫痫发作，其特征是具有正常EEG背景的全身性癫痫样放电，无或轻度认知障碍，对抗癫痫药物反应良好。所有这些变体都位于从1B到2A连续重复的中心区域，和蛋白质模型证明了Reelin的结构改变。此外，我们发现这些杂合错义变体显著降低了HEK-293T细胞中突变蛋白的分泌,这种损害是由于突变Reelin在分泌途径中的转运改变。
结论：这些结果表明RELN可能与GGE相关。由RELN变体引起的GGE的表型相对温和，致病机制可能涉及功能丧失。
