前脑小脑发育不全6型(PCH6)是一种早发性脑病,有/没有线粒体呼吸复合物缺陷,由线粒体精氨酰-tRNA合成酶(RARS2)的隐性突变引起。高度异质性的临床表型和许多意义不确定的错义变化使诊断变得困难。PCH6的发病机制尚不清楚。
评估患者的面部特征。进行基因测试。结构预测基于来自AlphaFold蛋白质结构数据库的模板。在HEK293T细胞中测试突变体RARS2的表达。检测患者来源的诱导多能干细胞(iPSCs)的人线粒体tRNAArg(hmtRNAArg)稳态水平,线粒体呼吸复合体(MRC)活性,耗氧率(OCR),细胞外酸化率(ECAR),线粒体膜电位(MMP),活性氧(ROS)丰度,和凋亡水平。
三个家系被诊断为由复合杂合RARS2变异引起的PCH6。鉴定了五种RARS2变体:c.3G>C(p。M1?),c.685C>T(p。R229*),c.1060T>A(p。F354I),c.1210A>G(p。M404V),和c.1369G>A(p。G457R)。RARS2c.3G>C破坏的蛋白质表达。RARS2c.685C>T产生了一种截短的蛋白质,缺乏完整的催化核心和反密码子结合结构域。预测RARS2c.1060T>A和c.1369G>A会导致结构异常。患者iPSCs中hmtRNAArg稳态丰度未受影响。线粒体能量代谢正常,包括MRC活动,OCR,ECAR,和MMP,而线粒体相关的细胞特征,包括ROS(P<0.001)和凋亡水平(P<0.001),增加。
本研究报告了五种RARS2变体,其中c.3G>C和c.1060T>A是新颖的。总结PCH6患者的面部特征将有助于诊断。线粒体能量代谢缺陷可能不是关键点,但是线粒体相关的异常细胞生理学,包括细胞凋亡,可能是潜在的发病机制。
Pontocerebellar hypoplasia type 6 (PCH6) is an early-onset encephalopathy with/without mitochondrial respiratory complex defects caused by recessive mutations in mitochondrial arginyl-tRNA synthetase (RARS2). Highly heterogeneous clinical phenotypes and numerous missense variations of uncertain significance make diagnosis difficult. Pathogenesis of PCH6 remains unclear.
Facial characteristics of patients were assessed. Genetic tests were performed. Structure prediction was based on the template from AlphaFold Protein Structure Database. Expression of mutant
RARS2 was tested in HEK293T cells. Patient-derived induced pluripotent stem cells (iPSCs) were detected for human mitochondrial tRNAArg (hmtRNAArg) steady-state level, mitochondrial respiratory complex (MRC) activity, oxygen consumption rate (OCR), extracellular acidification rate (ECAR), mitochondrial membrane potential (MMP), reactive oxygen species (ROS) abundance, and apoptosis level.
The three pedigrees were diagnosed as PCH6 caused by compound heterozygous RARS2 variations. Five
RARS2 variants were identified: c.3G>C(p.M1?), c.685C>T(p.R229∗), c.1060T>A(p.F354I), c.1210A>G(p.M404V), and c.1369G>A(p.G457R).
RARS2 c.3G>C disrupted protein expression. RARS2 c.685C>T created a truncated protein lacking complete catalytic core and anticodon-binding domain.
RARS2 c.1060T>A and c.1369G>A were predicted to cause structural abnormality. The hmtRNAArg steady-state abundance in a patient\'s iPSCs was unaffected. Mitochondrial energy metabolism was normal, including MRC activity, OCR, ECAR, and MMP, while mitochondria-related cellular characteristics, including ROS (P < 0.001) and apoptosis levels (P < 0.001), increased.
This study reports five
RARS2 variations among which c.3G>C and c.1060T>A are novel. Summarized facial features of PCH6 patients will facilitate diagnosis. Defective mitochondrial energy metabolism may not be key points, but mitochondria-related abnormal cellular physiology, including apoptosis, may be an underlying pathogenesis.