两个散发性颅骨锁骨发育不良综合征家系的致病机制探讨  

Pathogenic mechanism of 2 families with sporadic cleidocranial dysplasia syndrome

作　　者：陈红 刘倩 CHEN Hong;LIU Qian(School of Stomatology,Lanzhou University,Orthodontics Department of Gansu Provincial Hospital,China,730000;State Key Laboratory of Oral&Maxillofacial Reconstruction and Regeneration,National Clinical Research Center for Oral Diseases,Shaanxi Clinical Research Center for Oral Diseases,Department of Orthodontics,The Third Hospital Affiliated of Air Force Military Medical University,China)

机构地区：[1]兰州大学口腔医学院,甘肃省人民医院口腔正畸科,730000 [2]口颌系统重建与再生全国重点实验室,国家口腔疾病临床医学研究中心,陕西省口腔疾病临床医学研究中心,空军军医大学第三附属医院

出　　处：《实用口腔医学杂志》2024年第5期666-672,共7页Journal of Practical Stomatology

基　　金：甘肃省自然科学基金(编号:22JR5RA698);甘肃省卫生行业项目(编号:GSWSKY2018-11)。

摘　　要：目的:检测散发性颅骨锁骨发育不良综合征(CCD)家系的RUNX 2基因突变,探讨CCD的致病机制。方法:对2个散发性CCD家系的进行病史采集,外周血基因组DNA的提取,利用聚合酶链式反应,DNA直接测序检测突变,运用Swiss-Model软件预测,SWISS-Pdb、RasMol浏览器分析突变体蛋白构像,构建pEGFP-N1-RUNX 2真核载体后转染293-T细胞,观察荧光蛋白表达情况。结果:发现两例患者均表现出明显的“三联征”临床表型,并并在家系Ⅰ的先证者中检测到一个特定的基因突变,即c.243-260 del18(p.81-86del6),这导致了六个氨基酸的缺失。这一突变引起了蛋白质二级和三级结构的显著变化,包括部分螺旋和折叠结构的丧失。在家系Ⅱ的先证者中,则检测到了一个终止密码子突变c.1200C>A(p.stop400),这导致了转录过程的提前终止,进而使得蛋白质的部分氨基酸序列缺失,并且蛋白质出现了截短和显著的结构变化,失去了所有折叠结构以及大部分的转角结构。进一步的亚细胞定位实验显示,带有c.243-260del18(p.81-86del6)和c.1200C>A(p.stop400)突变的绿色荧光蛋白表达模式与野生型一致。结论:RUNX 2基因突变是CCD的致病基础,其中p.stop400是该研究检测到的新的基因突变位点。Objective:To investigate the pathogenic mechanism of sporadic cleidocranial dysplasia(CCD)by detecting RUNX 2 gene mutation in 2 CCD families.Methods:Collection of medical history and extraction of genomic DNA from peripheral blood were performed in 2 sporadic CCD families.Mutation was detected using PCR and direct DNA sequencing.Swiss-Model software,SWISS-Pdb and RasMol browser were used to predict and analyze the mutant protein conformation,respectively.After construction of the eukaryotic vector pEGFP-N1-RUNX 2,293-T cells were transfected and the fluorescent protein expression was observed.Results:The base deletion mutation c.243-260del18(p.81-86del6)was identified in the proband of familyⅠ,which led to the deletion of 6 amino acids and changes to the secondary and tertiary conformation of the protein.Notably,parts of the spiral and folded structures were lost.The stop codon mutation c.1200C>A(p.stop400)was detected in the proband of familyⅡ,which caused premature termination of transcription,deletion of some amino acids,truncation of the protein,and marked structural changes,including the loss of all folded structures and most corners.Analysis of subcellular localization using green fluorescent protein showed that the c.243-260del18(p.81-86del6)and c.1200C>A(p.stop400)mutants have the same localization as the wild type,indicating that p.81-86del6 and p.stop400 have no effect on the nuclear localization of RUNX 2 protein.Conclusion:RUNX 2 gene mutation is the pathogenic basis of CCD,and p.stop400 is a newly detected gene mutation site for CCD.

关 键 词：颅骨锁骨发育不良综合征 RUNX 2基因 基因突变 蛋白结构 

分 类 号：R78[医药卫生—口腔医学]