Enzymatic characterization of novel arylsulfatase A variants using human arylsulfatase A-deficient immortalized mesenchymal stromal cells.

Abstract

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.

• Authors: Judith Böhringer, René Santer, Neele Schumacher, Friederike Gieseke, Kerstin Cornils, Maria Pechan, Birgit Kustermann-Kuhn, Rupert Handgretinger, Ludger Schöls, Klaus Harzer, Ingeborg Krägeloh-Mann, Ingo Müller
• Journal: Human mutation (Hum Mutat) Vol. 38 Issue 11 Pg. 1511-1520 (11 2017) ISSN: 1098-1004 [Electronic] United States
• PMID: 28762252 (Publication Type: Journal Article)
• Copyright: © 2017 Wiley Periodicals, Inc.

Chemical References

• Cerebroside-Sulfatase

Topics

• Adolescent
• Alleles
• Cell Line, Transformed
• Cerebroside-Sulfatase (genetics, metabolism)
• Child
• DNA Mutational Analysis
• Enzyme Activation
• Exons
• Female
• Flow Cytometry
• Gene Expression
• Genetic Variation
• Genotype
• Humans
• Immunophenotyping
• Leukodystrophy, Metachromatic (diagnosis, enzymology, genetics)
• Male
• Mesenchymal Stem Cells (enzymology)
• Mutagenesis, Site-Directed
• Mutation
• Open Reading Frames
• Plasmids (genetics)
• Young Adult