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Enzymatic characterization of novel arylsulfatase A variants using human arylsulfatase A‐deficient immortalized mesenchymal stromal cells
Author(s) -
Böhringer Judith,
Santer René,
Schumacher Neele,
Gieseke Friederike,
Cornils Kerstin,
Pechan Maria,
KustermannKuhn Birgit,
Handgretinger Rupert,
Schöls Ludger,
Harzer Klaus,
KrägelohMann Ingeborg,
Müller Ingo
Publication year - 2017
Publication title -
human mutation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 162
eISSN - 1098-1004
pISSN - 1059-7794
DOI - 10.1002/humu.23306
Subject(s) - arylsulfatase a , metachromatic leukodystrophy , biology , lysosomal storage disease , mutation , gene , microbiology and biotechnology , genetics , enzyme , biochemistry
Metachromatic leukodystrophy (MLD) is an autosomal‐recessive lysosomal storage disease caused by mutations in the ARSA gene leading to arylsulfatase A (ARSA) deficiency and causing sulfatide accumulation. Main symptoms of the disease are progressive demyelination, neurological dysfunction, and reduced life expectancy. To date, more than 200 different ARSA variants have been reported in MLD patients. Here, we report the biochemical characterization of seven novel pathogenic variants (c.98T > C, c.195delC, c.229G > C, c.545C > G, c.674A > G, c.852T > A, and c.1274A > G), which were found when sequencing a cohort of 31 German MLD families. For that purpose, the ARSA cDNAs carrying the respective mutations inserted by site‐directed mutagenesis were cloned into a MigR1 (MSCV, IRES, GFP, retrovirus‐1) vector. The constructs were overexpressed using retroviral gene transfer in immortalized, human multipotent mesenchymal stromal cells prepared from a patient deficient in ARSA activity (late infantile MLD). In this novel ARSA −/− cell system, the seven ARSA mutants showed ARSA activity of less than 10% when compared with wild type, which is evidence for the pathogenicity of all seven variants. In conclusion, the system of ARSA −/− ‐immortalized MSC turned out to be a helpful novel tool for the biochemical characterization of ARSA variants.