A 5' SPLICE-REGION MUTATION AND A DINUCLEOTIDE DELETION IN THE LYSOSOMAL ACID LIPASE GENE IN 2 PATIENTS WITH CHOLESTERYL ESTER STORAGE DISEASE

Cholesteryl eater storage disease (CESD) results from inherited defici encies of the lysosomal hydrolase, acid lipase (LAL; E.C. 3.1.1.13). T o establish the molecular defects in LAL deficiency, two unrelated pro bands with severely reduced LAL activity were examined. DNA amplificat ion by reverse-transcription polymerase chain reaction and subsequent sequence analysis of LAL cDNA identified two mutant alleles. Patient 1 , presenting with hepatosplenomegaly, mildly elevated liver function t ests, and hyperlipidemia, was homozygous for a deletion of nucleotides 823 to 894 of the LAL cDNA. This 72-bp deletion maintained the readin g frame and resulted in a loss of 24 amino acids from the LAL protein. Analysis of genomic DNA revealed that the 72 bp corresponded to an ex on of the LAL gene. A single G to A point mutation at the last exon po sition was observed in the genomic DNA of patient 1, indicating a spli cing defect with consecutive exon skipping underlying the 72-bp deleti on. Patient 2 was a compound heterozygote for the 72-bp deletion and a dinucleotide deletion at positions 967 and 968. This deletion resulte d in a shifted reading frame carboxyterminal of codon 296, and 43 rand om amino acids followed the frame shift. A premature stop at codon 339 truncated the mutant LAL protein by 34 amino acids. Allele-specific h ybridization confirmed that patient 1 was homozygous for the 72-bp del etion mutation, and that patient 2 was a compound heterozygote for the 72-bp deletion and the 2-bp deletion.