播散性浅表性光线性汗孔角化症1例致病突变位点的鉴定及功能初探

Identification and Functional Investigation of a Pathogenic Mutation Site in a Case of Disseminated Superficial Actinic Porokeratosis

目的对播散性浅表性光线性汗孔角化症的突变位点进行鉴定并初步探讨其功能。方法对患者的外周血进行采样,收集全基因组DNA,采用PCR结合Sanger测序验证突变位点。构建野生型和突变型5-焦磷酸甲羟戊酸脱羧酶(MVD)相关真核表达载体,通过免疫印迹法检测MVD相关突变对蛋白在细胞中溶解度的影响。结果测序发现,MVD cDNA,c.746T>C是该患者的致病位点,此突变属于播散性浅表性光化性汗孔角化症的热点突变。功能研究发现该突变导致MVD蛋白在细胞中的溶解度改变,具体表现为野生型MVD主要存在于上清中,突变导致MVD蛋白以不可溶的状态存在于细胞沉淀中。结论本研究鉴定了该患者发病的遗传突变,证实了既往已报道过的突变位点,进一步表明MVD为播散性浅表性光线性汗孔角化症的致病基因,突变导致蛋白的溶解度降低可能是该病发病的原因之一。

Objective To identify and conduct a preliminary functional study of mutant loci in a case of disseminated superficial actinic porokeratosis.Methods Peripheral blood samples were collected from the patient for whole-genome DNA extraction.Subsequently,whole-genome exon sequencing was performed.The identified mutant loci were verified using a combination of PCR and Sanger sequencing.Additionally,wild-type and mutant mevalonate 5-pyrophosphate decarboxylase(MVD)-associated eukaryotic expression vectors were constructed,and the impact of MVD-associated mutations on protein solubility in cells was evaluated through immunoblotting.Results Sequencing analysis identified the MVD cDNA,specifically the c.746T>C mutation,as the causative locus in this patient.This mutation represented a hotspot mutation in cases of disseminated superficial actinic porokeratosis.Functional investigations demonstrated that the mutation led to altered solubility of the MVD protein within cells.Specifically,wild-type MVD protein was predominantly found in the supernatant,whereas the mutation resulted in the presence of insoluble MVD protein in the cellular sediment.Conclusion The findings of this study confirm the presence of genetic mutations associated with the pathogenesis of the patient′s condition.Furthermore,the results support previous reports highlighting the involvement of mutant loci in disseminated superficial actinic porokeratosis,with MVD being identified as the causative gene.Additionally,this study suggests that mutations leading to reduced solubility of proteins may play a significant role in the development and progression of the disease.