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Abstract：
BACKGROUND: Neurofibromatosis type 1 (NF-1) is caused by mutations in the NF1 gene that encodes neurofibromin, a negative regulator of RAS proto-oncogene. Approximately one-third of the reported pathogenic mutations in NF1 are splicing mutations, but most consequences are unclear. The objective of this study was to identify the pathogenicity of splicing mutation in a Chinese family with NF-1 and determine the effects of the pre-mRNA splicing mutation by in vitro functional analysis.
METHODS: Next-generation sequencing was used to screen candidate mutations. We performed a minigene splicing assay to determine the effect of the splicing mutation on NF1 expression, and three-dimensional structure models of neurofibromin were generated using SWISS-MODEL and PROCHECK methods, respectively.
RESULTS: A pathogenic splicing mutation c.479+1G>C in NF1 was found in the proband characterized by childhood-onset refractory hypertension. In vitro analysis demonstrated that c.479+1G>C mutation caused the skipping of exon 4, leading to a glutamine-to-valine substitution at position 97 in neurofibromin and an open reading frame shift terminating at codon 108. Protein modeling showed that several major domains were missing in the truncated neurofibromin protein.
CONCLUSIONS: The splicing mutation c.479+1G>C identified in a Chinese patient with NF-1 and childhood-onset refractory hypertension caused the skipping of exon 4 and a truncated protein. Our findings offer new evidence for the molecular diagnosis of NF-1.
METHODS: Next-generation sequencing was used to screen candidate mutations. We performed a minigene splicing assay to determine the effect of the splicing mutation on NF1 expression, and three-dimensional structure models of neurofibromin were generated using SWISS-MODEL and PROCHECK methods, respectively.
RESULTS: A pathogenic splicing mutation c.479+1G>C in NF1 was found in the proband characterized by childhood-onset refractory hypertension. In vitro analysis demonstrated that c.479+1G>C mutation caused the skipping of exon 4, leading to a glutamine-to-valine substitution at position 97 in neurofibromin and an open reading frame shift terminating at codon 108. Protein modeling showed that several major domains were missing in the truncated neurofibromin protein.
CONCLUSIONS: The splicing mutation c.479+1G>C identified in a Chinese patient with NF-1 and childhood-onset refractory hypertension caused the skipping of exon 4 and a truncated protein. Our findings offer new evidence for the molecular diagnosis of NF-1.
摘要：
背景：1型神经纤维瘤病（NF-1）是由编码神经纤维蛋白的NF1基因突变引起的，RAS原癌基因的负调节因子。NF1中报道的致病性突变中约有三分之一是剪接突变，但大多数后果尚不清楚。这项研究的目的是鉴定剪接突变在中国NF-1家族中的致病性，并通过体外功能分析确定pre-mRNA剪接突变的影响。
方法：使用下一代测序筛选候选突变。我们进行了小基因剪接分析，以确定剪接突变对NF1表达的影响，并使用SWISS-MODEL和PROCHECK方法生成了神经纤维蛋白的三维结构模型。分别。
结果：在NF1中发现了致病性剪接突变c.4791G>C，其特征是儿童期发作的难治性高血压。体外分析表明,c.479+1G>C突变引起外显子4的跳跃,导致在神经纤维蛋白中的位置97处的谷氨酰胺到缬氨酸的置换和在密码子108处终止的开放阅读框移位。蛋白质建模表明，截短的神经纤维蛋白中缺少几个主要结构域。
结论：在患有NF-1和儿童期发作的难治性高血压的中国患者中鉴定的剪接突变c.4791G>C导致外显子4和截短蛋白的跳跃。我们的发现为NF-1的分子诊断提供了新的证据。
方法：使用下一代测序筛选候选突变。我们进行了小基因剪接分析，以确定剪接突变对NF1表达的影响，并使用SWISS-MODEL和PROCHECK方法生成了神经纤维蛋白的三维结构模型。分别。
结果：在NF1中发现了致病性剪接突变c.4791G>C，其特征是儿童期发作的难治性高血压。体外分析表明,c.479+1G>C突变引起外显子4的跳跃,导致在神经纤维蛋白中的位置97处的谷氨酰胺到缬氨酸的置换和在密码子108处终止的开放阅读框移位。蛋白质建模表明，截短的神经纤维蛋白中缺少几个主要结构域。
结论：在患有NF-1和儿童期发作的难治性高血压的中国患者中鉴定的剪接突变c.4791G>C导致外显子4和截短蛋白的跳跃。我们的发现为NF-1的分子诊断提供了新的证据。
