2 EXON-SKIPPING MUTATIONS AS THE MOLECULAR-BASIS OF SUCCINIC SEMIALDEHYDE DEHYDROGENASE-DEFICIENCY (4-HYDROXYBUTYRIC ACIDURIA)

Succinic semialdehyde dehydrogenase (SSADH) deficiency, a rare metabol ic disorder of 4-aminobutyric acid degradation, has been identified in similar to 150 patients. Affected individuals accumulate large quanti ties of 4-hydroxybutyric acid, a compound with a wide range of neuroph armacological activities, in physiological fluids. As a first step in beginning an investigation of the molecular genetics of SSADH deficien cy, we have utilized SSADH cDNA and genomic sequences to identify two point mutations in the SSADH genes derived from four patients. These m utations, identified by standard methods of reverse transcription, PCR , dideoxy-chain termination, and cycle sequencing, alter highly conser ved sequences at intron/exon boundaries and prevent the RNA-splicing a pparatus from properly recognizing the normal splice junction. Each fa mily segregated a mutation in a different splice site, resulting in ex on skipping and, in one case, a frameshift and premature termination a nd, in the other case, an in-frame deletion in the resulting protein. Family members, including parents and siblings of these patients, were shown to be heterozygotes for the splicing abnormality, providing add itional evidence for autosomal recessive inheritance. Our results prov ide the first evidence that 4-hydroxybutyric aciduria, resulting from SSADH deficiency, is the result of genetic defects in the human SSADH gene.