A 2-BASE DELETION IN EXON-6 OF THE 3-HYDROXY-3-METHYLGLUTARYL COENZYME-A LYASE (HL) GENE PRODUCING THE SKIPPING OF EXON-5 AND EXON-6 DETERMINES 3-HYDROXY-3-METHYLGLUTARIC ACIDURIA

A novel two-base deletion in the 3-hydroxy-3-methylglutaryl coenzyme A lyase (HL) gene was found in a Spanish patient with homozygous 3-hydr oxy-3-methylglutaric aciduria. Amplification by RT-PCR of the mRNAs sh owed that the gene was transcribed into three different mRNAs. One sho wed the complete deletion of exons 5 and 6 located between nucleotides 348 and 561 of the HL cDNA. The second transcript showed deletion of exon 6 only, and the third contained a two-base deletion CT in exon 6, corresponding to nucletotides 504 and 505 of the HL cDNA. These aberr ant mRNAs are predicted to encode three abnormal HMG-CoA lyase protein s; tile first (from skipped exons 5 and 6) lacks 7] amino acids, which represents 24% of the mature protein; the second, (from the skipping of exon 6, producing a frameshift) contains only 192 amino acids, the last 26 of which are missense amino acids preceding a stop codon; the third contains only 175 amino acids, the last 7 of which are missense. Northern blot analysis showed that the HL mRNA levels of the pa tient were 4% of the control. PCR quantitative analysis indicated that the mRNA lacking exons 5 and 6 was the most abundant, representing 88% of the total. The other two mRNAs represented 8% and 4%, respectively. In the genomic DNA only one CT deletion was found at positions +7 and +8 at beginning of exon 6. No mutations were observed in the splice dono r, splice acceptor, or pyrimidine-rich sequences of the intronic regio ns flanking exons 5 and 6. All three aberrant mRNAs resulted only from tile deletion of nucleotides CT. We suggest that this deletion may af fect the interaction be tween the small nuclear ribonucleoproteins (sn RNPs) and exon 6, and that, as a result, the abnormal splicing of tile pre-mRNA produces two different aberrant transcripts.