Background

Acromesomelic dysplasia, type Maroteaux (AMDM) is a rare skeletal dysplasia following autosomal recessive mode of inheritance and characterized by abnormal growth plates, short and abnormal bones in the extremities and spine.

Objective

Present study was designed to report the molecular basis of AMDM in enrolled consanguineous family from Pakistan.

Methods

A consanguineous family from Vehari District in Pakistan having multiple siblings suffering from AMDM was enrolled in present study. Whole exome sequencing (WES) approach was adopted to identify causative agent of AMDM. Human full length NPR2 gene and sequence with nonsense mutation was amplified by using Myc-tagged pXN vector and transformed in E. coli DH5α cells to confirm mutation. SDS-PAGE and Western blotting were done to confirm the production of truncated protein. Computational three dimensional structure generation through homology modeling approach was done to compare protein structure between patients and controls.

Results

WES reveled a nonsense mutation (c.613 C>T, p.R205X) in exon 1 of NPR2 gene leading to premature termination codon in mRNA of NPR2 gene resulting in a truncated protein with 204 amino acid residues that was confirmed by SDS-PAGE and Western blotting. Sanger sequencing confirmed that mutation in all subjects and mutation followed Mendalian pattern of inheritance. Multiple sequence alignment by ClustalW revealed that mutated domain of NPR2 is conserved region. Proetin structure comparison revealed a significant structural part of NPR2 was missing in truncated protein as compared to control.

Conclusion

We are reporting that a novel nonsense mutation (c.613 C>T, p.R205X) in exon 1 of NPR2 gene is causing AMDM in a consanguineous Pakistani family.

中文翻译：

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NPR2基因中的一个新的无意义突变，导致Acomesomelic发育异常，在旁遮普省南部（巴基斯坦）近亲家庭中为Maroteaux型。

背景

马洛索氏型不典型增生（AMDM）是继常染色体隐性遗传后的罕见骨骼发育不良，其特征是生长板异常，四肢和脊柱骨短而异常。

目的

本研究旨在报告来自巴基斯坦已入血的近亲家庭中AMDM的分子基础。

方法

本研究纳入了来自巴基斯坦Vehari区的一个近亲家庭，该家庭有多个患有AMDM的兄弟姐妹。采用全外显子组测序（WES）方法确定AMDM的病原体。用Myc标记的pXN载体扩增人无义突变的全长NPR2基因和序列，并在大肠杆菌DH5α细胞中转化以确认突变。进行SDS-PAGE和蛋白质印迹以确认截短蛋白的产生。通过同源建模方法计算了三维结构，以比较患者和对照之间的蛋白质结构。

结果

WES揭示了NPR2基因外显子1的无意义突变（c.613 C> T，p.R205X），导致NPR2基因mRNA的过早终止密码子，导致具有204个氨基酸残基的截短蛋白已通过SDS-PAGE证实和蛋白质印迹。桑格测序证实，所有受试者的突变和突变均遵循Mendalian遗传模式。ClustalW的多序列比对揭示了NPR2的突变域是保守区。蛋白质结构比较显示与对照相比，截短蛋白中NPR2的重要结构部分缺失。

结论

我们报道NPR2基因外显子1中的一个新的无意义突变（c.613 C> T，p.R205X）导致巴基斯坦近亲家庭中的AMDM。