Alternative Splicing in the α-Galactosidase A Gene: Increased Exon Inclusion Results in the Fabry Cardiac Phenotype

Fabry disease is an inborn error of glycosphingolipid catabolism, resulting from deficient activity of lysosomal α-galactosidase A (α-Gal A). A rare alternative splicing that introduces a 57-nucleotide (nt) intronic sequence to the α-Gal A transcript from intron 4 of the gene has been identified. In addition, a novel midintronic base substitution that results in substantially increased alternative splicing has been identified in a patient with Fabry disease who has the cardiac variant phenotype. The sequence of the patient's intron 4 contains a single G→A transversion at genomic nt 9331 (IVS4+919G→A), located at the −4 position of the 3′ end of the intronic insertion (nts 9278–9334 in the genomic sequence). Minigene constructs containing the entire intron 4 sequence with G, A, C, or T at nt 9331 within an α-Gal A complementary DNA expression vector were prepared and expressed in COS-1 cells. Whereas transfection of the G or T minigenes transcribed predominantly normal-sized transcripts, the transfection of the A or C minigenes produced a large amount of the alternatively spliced transcript. These results suggest that the G→A mutation, within an A/C-rich domain, results in increased recognition of the alternative splicing by an A/C-rich enhancer–type exonic splicing enhancer. The intronic mutation was not observed in 100 unrelated unaffected men but was present in 6 unrelated patients with cardiac Fabry disease. Reverse-transcriptase polymerase chain reaction of total RNA of various normal human tissues revealed that the alternatively spliced transcript was present in all of the samples, and especially at a higher ratio in the lung and muscle. The normal transcript was present in the patients' lymphoblasts and resulted in ∼10% residual enzyme activity, leading to a cardiac phenotype of Fabry disease