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Abstract：
BACKGROUND: Wolfram syndrome (WS) is a rare autosomal recessive disorder characterized by diabetes insipidus, diabetes mellitus, optic atrophy and deafness. Mutations in Wolfram syndrome 1 (WFS1) gene may cause dysregulated endoplasmic reticulum (ER)-stress and cell apoptosis, contributing to WS symptoms. The aim of this study was to identify the molecular etiology of a case of WS and to explore the functional consequence of the mutant WFS1 gene in vitro.
METHODS: A 27 years-old Chinese man was diagnosed as wolfram syndrome type 1 based on clinical data and laboratory data. DNA sequencing of WFS1 gene and mitochondrial m.3337G > A, m.3243A > G mutations were performed in the patient and his 4 family members. Functional analysis was performed to assessed the in vitro effect of the newly identified mutant. ER stress were evaluated by ER stress response element (ERSE)-luciferase assay. Cell apoptosis were performed by CCK-8, TUNEL staining and flow cytometric analysis.
RESULTS: A novel heterozygous 10-base deletion (c. 2067_2076 del10, p.W690fsX706) was identified in the patient. In vitro studies showed that mutant p.W690fsX706 increased ERSE reporter activity in the presence or absence of thapsigargin instead of wild type WFS1. Knockdown of WFS1 activated the unfolded protein response (UPR) pathway and increased the cell apoptosis, which could not be restored by transfection with WFS1 mutant (p.W690fsX706) comparable to the wild type WFS1.
CONCLUSIONS: A novel heterozygous mutation of WFS1 detected in the patient resulted in loss-of-function of wolframin, thereby inducing dysregulated ER stress signaling and cell apoptosis. These findings increase the spectrum of WFS1 gene mutations and broaden our insights into the roles of mutant WFS1 in the pathogenesis of WS.
METHODS: A 27 years-old Chinese man was diagnosed as wolfram syndrome type 1 based on clinical data and laboratory data. DNA sequencing of WFS1 gene and mitochondrial m.3337G > A, m.3243A > G mutations were performed in the patient and his 4 family members. Functional analysis was performed to assessed the in vitro effect of the newly identified mutant. ER stress were evaluated by ER stress response element (ERSE)-luciferase assay. Cell apoptosis were performed by CCK-8, TUNEL staining and flow cytometric analysis.
RESULTS: A novel heterozygous 10-base deletion (c. 2067_2076 del10, p.W690fsX706) was identified in the patient. In vitro studies showed that mutant p.W690fsX706 increased ERSE reporter activity in the presence or absence of thapsigargin instead of wild type WFS1. Knockdown of WFS1 activated the unfolded protein response (UPR) pathway and increased the cell apoptosis, which could not be restored by transfection with WFS1 mutant (p.W690fsX706) comparable to the wild type WFS1.
CONCLUSIONS: A novel heterozygous mutation of WFS1 detected in the patient resulted in loss-of-function of wolframin, thereby inducing dysregulated ER stress signaling and cell apoptosis. These findings increase the spectrum of WFS1 gene mutations and broaden our insights into the roles of mutant WFS1 in the pathogenesis of WS.
摘要：
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