Dipartimento di Bioscienze, Università degli Studi di Milano, Milan, Italy.
Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico, UOC Medicina Generale, Milan, Italy.
Genet Med. 2020 Jan;22(1):35-43. doi: 10.1038/s41436-019-0584-0. Epub 2019 Jul 5.
Existing data do not explain the reason why some individuals homozygous for the hypomorphic FECH allele develop erythropoietic protoporphyria (EPP) while the majority are completely asymptomatic. This study aims to identify novel possible genetic variants contributing to this variable phenotype.
High-throughput resequencing of the FECH gene, qualitative analysis of RNA, and quantitative DNA methylation examination were performed on a cohort of 72 subjects.
A novel deep intronic variant was found in four homozygous carriers developing a clinically overt disease. We demonstrate that this genetic variant leads to the insertion of a pseudo-exon containing a stop codon in the mature FECH transcript by the abolition of an exonic splicing silencer site and the concurrent institution of a new methylated CpG dinucleotide. Moreover, we show that the hypomorphic FECH allele is linked to a single haplotype of about 20 kb in size that encompasses three noncoding variants that were previously associated with expression quantitative trait loci (eQTLs).
This study confirms that intronic variants could explain the variability in the clinical manifestations of EPP. Moreover, it supports the hypothesis that the control of the FECH gene expression can be mediated through a methylation-dependent modulation of the precursor messenger RNA (pre-mRNA) splicing pattern.
现有数据无法解释为什么一些纯合子携带低功能 FECH 等位基因的个体发生红细胞生成性原卟啉症（EPP），而大多数个体则完全无症状。本研究旨在确定可能导致这种表型可变的新的遗传变异。
对 72 例患者进行 FECH 基因高通量重测序、RNA 定性分析和定量 DNA 甲基化检测。
在四个表现为临床显性疾病的纯合子携带者中发现了一种新的深内含子变异。我们证明，这种遗传变异通过消除外显子剪接沉默位点并同时建立新的甲基化 CpG 二核苷酸，导致成熟 FECH 转录本中插入一个包含终止密码子的假外显子。此外，我们表明，低功能 FECH 等位基因与一个大约 20kb 大小的单倍型相关，该单倍型包含三个先前与表达数量性状基因座（eQTLs）相关的非编码变异。
本研究证实，内含子变异可以解释 EPP 临床表现的可变性。此外，它支持这样一种假设，即 FECH 基因表达的控制可以通过对前体信使 RNA（pre-mRNA）剪接模式的甲基化依赖性调节来介导。
