Pah基因深内含子突变小鼠的表型分析
Phenotypic characterization of a phenylketonuria mouse model with a deep intronic mutation inPahgene
刘淑婷 1周飞1
作者信息
- 1. 苏州大学苏州医学院剑桥-苏大基因组资源中心，江苏省苏州市 215123
- 折叠
摘要
目的：探讨人源苯丙氨酸羟化酶Pah c.1199＋502A>T深内含子突变对mRNA剪接及蛋白表达的影响，并系统评估该突变导致的小鼠表型特征。方法：利用RT-PCR结合Sanger测序解析转录本剪接模式，通过实时荧光定量PCR及蛋白免疫印迹检测mRNA及蛋白表达水平，并结合血清氨基酸代谢分析及行为学实验，多维度评估小鼠代谢状况与神经行为学表型。结果：分子机制表明c.1199+502A>T突变通过激活隐蔽剪接位点导致25-nt假外显子异常插入，引起mRNA剪接缺陷及蛋白表达显著下调；表型分析显示突变小鼠呈现毛色减退、发育迟缓及血清苯丙氨酸浓度显著升高的典型病理特征；行为学实验证实突变小鼠存在活动量减少及显著的焦虑样表型。结论：该深内含子突变通过诱导假外显子剪接导致蛋白功能缺失，引发严重的代谢紊乱与行为异常；该模型高度还原了人类致病特征，为精准治疗研究提供了可靠的体内评估平台。
Abstract
Objective: To investigate the impact of the human phenylalanine hydroxylase Pah c.1199+502A>T deep intronic mutation on mRNA splicing and protein expression, and to systematically characterize the resulting pathological phenotypes in a mutant mouse model. Methods: Splicing patterns were analyzed through RT-PCR combined with Sanger sequencing; mRNA and protein expression levels were assessed through real-time quantitative PCR (RT-qPCR) and Western blotting. Amino acid metabolism and neurobehavioral phenotypes in mice were evaluated through serum amino acid profiling and behavioral assays. Results:The c.1199+502A>T mutation activates cryptic splice sites, leading to the aberrant insertion of a 25-nt pseudo-exon, which subsequently causes splicing defects and a significant down-regulation of protein expression. Phenotypic analysis demonstrates that mutant mice exhibit typical pathological features, including hypopigmentation, growth retardation, and markedly elevated serum phenylalanine concentrations. Behavioral experiments confirm diminished spontaneous activity and a prominent anxiety-like phenotype. Conclusion:This deep intronic mutation disrupts protein function by inducing pseudo-exon inclusion, leading to severe metabolic derangement and behavioral abnormalities. This model faithfully recapitulates human pathogenic characteristics, providing a robust in vivo platform for research into precision therapeutic interventions.关键词
疾病模型/表型分析/苯丙酮尿症/深内含子突变。Key words
Disease model/Phenotypic analysis/Phenylketonuria/Deep intronic mutation.引用本文复制引用
刘淑婷,周飞.Pah基因深内含子突变小鼠的表型分析[EB/OL].(2026-04-29)[2026-04-30].http://www.paper.edu.cn/releasepaper/content/202604-213.学科分类
生物科学研究方法、生物科学研究技术/遗传学/分子生物学
评论