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Molecular study on the infantile form of Pompe disease in Chinese in Taiwan
Abstract:
Glycogen-storage disease type II, Pompe disease, is caused by the deficiency of acid alpha-D-glucosidase in lysosome. Previously we found that acid alpha-D-glucosidase did exist in the skin fibroblasts and there was also no difference of mRNA in quantity and size of Chinese infantile type Pompe disease patients in Taiwan. However, functional assay of the acid alpha-D-glucosidase of these patients showed its enzyme function to be defective. In the present study, first we identified a substitution site in four Chinese infantile patients with Pompe disease which is a cytidine to adenosine (C1935-->A) transversion at 5' end of exon 14 causing substitution of glutamic acid for aspartic acid at position 645 of the acid alpha-D-glucosidase. This substitution was introduced in wild-type cDNA and expressed in COS-1 cells. The Asp-645-->Glu substitution resulted in significant reduction of acid alpha-D-glucosidase activity. Second, according to the screening data in 25 Chinese Pompe disease patients using digestion of RT-PCR amplified specific fragment with Aat II, the restriction fragment length analysis showed that patients presented the 861 bp band and the normal individuals presented the 728 bp and 133 bp polymorphic bands. We found that the frequency of mutant allele is 0.8 in infantile patients with Chinese Pompe disease and 0 in normal individuals. These results therefore indicate that Asp-645-->Glu mutation results in infantile form of Pompe disease as the major cause in Chinese patients in Taiwan.
Insights
A specific mutation, Asp-645-->Glu, in acid alpha-D-glucosidase is identified as the primary cause of infantile Pompe disease in Chinese patients in Taiwan. This genetic defect significantly impairs enzyme function, leading to the disease.
Area of Science:
- Genetics
- Biochemistry
- Rare Diseases
Background:
- Pompe disease (Glycogen-storage disease type II) stems from acid alpha-D-glucosidase deficiency.
- Previous studies found normal mRNA levels but defective enzyme function in Chinese infantile Pompe disease patients.
Purpose of the Study:
- To identify the genetic cause of infantile Pompe disease in Chinese patients in Taiwan.
- To characterize the identified mutation and its impact on enzyme activity.
Main Methods:
- Genetic sequencing to identify mutations in affected patients.
- Site-directed mutagenesis and expression in COS-1 cells to assess enzyme function.
- Restriction fragment length polymorphism (RFLP) analysis using RT-PCR and Aat II digestion for mutation screening.
Main Results:
- A C1935-->A transversion was identified, leading to an Asp-645-->Glu substitution in acid alpha-D-glucosidase.
- The Asp-645-->Glu mutation significantly reduced enzyme activity.
- The mutant allele frequency was 0.8 in infantile Pompe disease patients versus 0 in normal individuals.
Conclusions:
- The Asp-645-->Glu mutation is a major cause of infantile Pompe disease in the Chinese population in Taiwan.
- This specific mutation severely impairs acid alpha-D-glucosidase function, leading to the disease phenotype.