W474C AMINO-ACID SUBSTITUTION AFFECTS EARLY PROCESSING OF THE ALPHA-SUBUNIT OF BETA-HEXOSAMINIDASE-A AND IS ASSOCIATED WITH SUBACUTE G(M2) GANGLIOSIDOSIS

Mutations in the HEXA gene, encoding the alpha-subunit of beta-hexosam inidase A (Hex A), that abolish Hex A enzyme activity cause Tay-Sachs disease (TSD), the fatal infantile form of G(M2) gangliosidosis, Type 1. Less severe, subacute (juvenile onset) and chronic (adult-onset) va riants are characterized by a broad spectrum of clinical manifestation s and are associated with residual levels of Hex A enzyme activity. We identified a 1422 G-->C (amino acid W474C) substitution in the first position of exon 13 of HEXA of a non-Jewish proband who manifested a s ubacute variant of G(M2) gangliosidosis. On the second maternally inhe rited allele, we identified the common infantile disease causing 4-bp insertion, +TATC 1278, in exon 11. Pulse chase analysis using proband fibroblasts revealed that the W474C-containing alpha-subunit precursor was normally synthesized, but not phosphorylated or secreted, and the mature lysosomal alpha-subunit was not detected. When the W474C-conta ining alpha-subunit was transiently co-expressed with the beta-subunit to produce Hex A (alpha beta) in COS-7 cells, the mature alpha-subuni t was present, but its level was much lower than that from normal alph a-subunit transfections, although higher than in those cells transfect ed with an alpha-subunit associated with infantile TSD. Furthermore, t he precursor level of the W474C alpha-subunit was found to accumulate in comparison to the normal alpha-subunit precursor levels. We conclud e that the 1422 G-->C mutation is the cause of Hex A enzyme deficiency in the proband. The resulting W474C substitution clearly interferes w ith alpha-subunit processing but because the base substitution falls a t the first position of exon 13, aberrant splicing may also contribute to Hex A deficiency in this proband. (C) 1998 Wiley-Liss, Inc.