Unbalanced GLA mRNAs ratio quantified by real-time PCR in Fabry patients' fibroblasts results in Fabry disease

被引:30
作者
Filoni, Camilla [2 ]
Caciotti, Anna [2 ]
Carraresi, Laura [2 ]
Donati, Maria Alice [2 ]
Mignani, Renzo [3 ]
Parini, Rossella [4 ]
Filocamo, Mirella [5 ]
Soliani, Fausto [6 ]
Simi, Lisa [7 ]
Guerrini, Renzo [2 ]
Zammarchi, Enrico [1 ]
Morrone, Amelia [2 ]
机构
[1] Univ Florence, Dept Pediat, I-50132 Florence, Italy
[2] Univ Florence, Metab & Muscular Unit, Clin Pediat Neurol, AOU Meyer, I-50132 Florence, Italy
[3] Osped Infermi, Nephrol Unit, Rimini, Italy
[4] San Gerardo Hosp, Metab Unit, Monza, Italy
[5] IRCCS Gaslini, Genoa, Italy
[6] Arciospedale S Maria Osped Nuova, Nephrol Unit, Reggio Emilia, Italy
[7] Univ Florence, Dept Clin Physiopathol, I-50132 Florence, Italy
关键词
Fabry disease; alpha-galactosidase; cryptic exon;
D O I
10.1038/ejhg.2008.109
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Total or partial deficiency of the human lysosomal hydrolase alpha-galactosidase A is responsible for Fabry disease, the X-linked inborn error of glycosphingolipid metabolism. Together with the predominant alpha-galactosidase A gene mRNA product encoding the lysosomal enzyme, a weakly regulated alternatively spliced alpha-galactosidase A transcript is expressed in normal tissues, but its overexpression, due to the intronic g.9331G>A mutation, leads to the cardiac variant. We report the molecular characterization of five Fabry patients including two siblings. Sequencing analysis of the alpha-galactosidase A gene coding region and intron/exon boundaries identified the new c.124A>G (p.M42V) genetic lesion as well as a known deletion in three patients, whereas in the two remaining patients, no mutations were identified. To evaluate possible alpha-galactosidase A gene transcription alterations, both predominant and alternatively spliced mRNAs were quantified by absolute real-time PCR on total RNA preparations from the patients' fibroblasts. An impressive reduction in the predominant alpha-galactosidase A transcript was detected in the last patients (Pt 4 and Pt 5). However, the alternatively spliced mRNA was dramatically overexpressed in one of them, carrying a new intronic lesion (g.9273C>T). These findings strongly suggest a correlation between this new intronic mutation and the unbalanced alpha-galactosidase A mRNAs ratio, which could therefore be responsible for the reduced enzyme activity causing Fabry disease. The real-time assay developed here to investigate the two alpha-galactosidase A mRNAs might play a crucial role in revealing possible genetic lesions and in confirming the pathogenetic mechanisms underlying Fabry disease. European Journal of Human Genetics (2008) 16, 1311-1317; doi:10.1038/ejhg.2008.109; published online 18 June 2008
引用
收藏
页码:1311 / 1317
页数:7
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