No abstract

Three novel Tay--Sachs Disease (TSD) mutations have been identified in two unrelated, non-Jewish compound heterozygous patients. A G772C transversion mutation causing an Asp258His substitution is shared by both patients. The mutant enzyme had been characterized, on the basis of previous kinetic studies (1) as a B1, or alpha-subunit active site mutation. This is the first B1 mutation not found in codon 178 (exon 5). A C508T transition causing an Arg170Trp substitution also occurred in one of the patients. The third mutation is a two base deletion occurring in exon 8 involving the loss of either nts 927-928 or 929-930 in codon 310. The deletion creates an inframe termination codon 35 bases downstream. The Arg170Trp mutation was also detected in a third unrelated TSD patient. In both families this allele was traced to French Canadian ancestors originating in the Estrie region of the province of Quebec. This mutation is the third TSD allele unique to the French Canadian population and the ancestral origins of the carrier parents are distant from the center of diffusion of the more common 7.6 kb deletion mutation which is in the eastern part of the province.

Abstract. The major excreted protein (MEP) of transformed mouse fibroblasts is the lysosomal protease, cathepsin L. MEP is also secreted by untransformed mouse cells in response to growth factors and tumor promoters, and is thought to play a role in cell growth and transformation. To determine the relationship between MEP synthesis and MEP secretion, we have examined these events in PDGF-treated NIH 3T3 cells. PDGF enhanced MEP synthesis and caused the diversion of MEP from the lysosomal delivery pathway to a secretory pathway. These two effects were found to be regulated independently at various times after growth factor addition. Short PDGF treatments (0.5 or 1 h) resulted in quantitative secretion of MEP although synthesis was near the control level. High levels of both synthesis and secretion occurred between 2 and 14 h of PDGF treatment. Between 18 and 30 h, the amount of secreted MEP returned to the low control level even though synthesis remained elevated. The secretion was specific for MEP; other lysosomal enzymes were not found in the media from PDGFtreated cells. PDGF-induced secretion of MEP was inhibited 84% by cycloheximide, suggesting that protein synthesis is required to elicit this effect. PDGF also caused a time-dependent increase in mannose 6-phosphate (Man-6-P) receptor-mediated endocytosis. These data support a model in which PDGF alters the distribution of Man-6-P receptors such that the Golgi concentration of receptors becomes limiting, thereby causing the selective secretion of the low affinity ligand, MEP.

Tay-Sachs disease (TSD) is an autosomal recessive genetic disorder resulting from mutation of the HEXA gene encoding the alpha-subunit of the lysosomal enzyme, beta-N-acetylhexosaminidase A (Hex A). We have discovered that a Tay-Sachs mutation, IVS-9 + 1 G-->A, first detected by Akli et al. (Genomics 11:124-134, 1991), is a common disease allele in non-Jewish Caucasians (10/58 alleles examined). A PCR-based diagnostic test, which detects an NlaIII site generated by the mutation, revealed a frequency among enzyme-defined carriers of 9/64 (14%). Most of those carrying the allele trace their origins to the United Kingdom, Ireland, or Western Europe. It was not identified among 12 Black American TSD alleles or in any of 18 Ashkenazi Jewish, enzyme-defined carriers who did not carry any of the mutations common to this population. No normally spliced RNA was detected in PCR products generated from reverse transcription of RNA carrying the IVS-9 mutation. Instead, the low levels of mRNA from this allele were comprised of aberrant species resulting from the use of either of two cryptic donor sites, one truncating exon 9 and the other within IVS-9, spliced to exon 10. Numerous additional splice products were detected, most involving skipping of one or more surrounding exons. Together with a recently identified allele responsible for Hex A pseudodeficiency (Triggs-Raine et al. Am J Hum Genet, 1992), these two alleles accounted for almost 50% (29/64) of TSD or carrier alleles ascertained by enzyme screening tests in non-Jewish Caucasians.