Detection of a new mutation in the beta-myosin heavy chain gene in an individual with hypertrophic cardiomyopathy.

Abstract

Familial hypertrophic cardiomyopathy (FHCM) is an autosomal dominant disease affecting primarily the myocardium. The gene responsible for FHCM has been localized to chromosome 14 in some families and several mutations have been described in the #-myosin heavy chain (ftMHC), a candidate gene for the disease. We recently identified a family with HCM in whom we did not detect any of the known mutations in the ftMHC gene (the a/i#MHC hybrid gene and the missense mutation in exons 13 and 9). However, we did observe a novel 9.5-kb BamHI restriction fragment length polymorphism detected by a ,6MHC probe on Southern blots of DNA from the proband of this family. Similarly, a novel 3.8-kb TaqI polymorphism and a novel 4.3-kb HindI1 polymorphism were detected on Southern blots of DNA from the same proband. Polymerase chain reaction (PCR) was used to amplify the segment of the ,MHC that was detected by pSC14 probe. PCR amplification of the distal 3'-end of the i6MHC gene yielded an additional product in the DNA template from the proband which was subsequently cloned and sequenced. The sequence analysis showed a 2.4-kb nucleotide deletion involving one allele of the ,6MHC gene. The deletion includes part of the intron 39, exon 40 including the 3'-untranslated region and the polyadenylation signal, and part of the fl-aMHC intergenic region. This deletion was inherited in Mendelian fashion in an additional three members of this small family of which only the proband has developed clinically diagnosed HCM at a very late onset (age 59 yr), the other three family members are younger and have not developed the disease at the ages of 10, 32, and 33 yr. (J. Clin. Invest. 1992.

A transgenic rabbit model for human hypertrophic cardiomyopathy

“…Southern blot analysis was performed as described previously (35). In brief, 10 µg of genomic DNA was digested with Bam H1 restriction endonuclease, which releases a 1.3-kb fragment of the transgenes.…”

“…In brief, RNA was extracted from left ventricle, left atrium, skeletal muscle, aorta, lungs, liver, and spleen of transgenic and NLM rabbits by guanidinium thiocyanate method (36) to characterize expression pattern of the endogenous β-MyHC mRNA. To detect expression of the endogenous β-MyHC mRNA in different tissues, RT-PCR was performed on total RNA extracts using RT primer 5′-CTGCTGCAGCTTCTCGTTGGTG-3′ and PCR primers (forward: 5′-AACAGAAGCAGCGGGAGGAGC-3′ and reverse: the same as the RT primer) as described previously (35,36). The expected size of the RT-PCR product was 368 bp.…”

Dominant-negative effect of a mutant cardiac troponin T on cardiac structure and function in transgenic mice.

“…Southern blotting was performed as described previously (26). In brief, 10-g aliquots of DNA were digested overnight with 40 U of HindIII and NotI, to release the entire transgene.…”

“…Indeed, identification of deletion mutations in the ␤-MyHC or the MyBP-C genes has put forth the "null-allele" hypothesis (26,41). Certain deletion mutations, responsible for human HCM, code for truncated proteins that, even if synthesized, are not expected to be incorporated into myofibrils (26,41).…”

Genetic Bases of Hypertrophic Cardiomyopathy

“…In their near-totality, mutations that affect the gene of the cardiac β myosin heavy chain are point mutations of the type described as "missense mutations" in which the substitution of a single base of the DNA in a given exon results in the exchange of the encoded amino acid 15 . Deletion of the nontranslated 3' region of the gene, inducing the loss of at least 5 amino acids in the rod of the protein has also been reported 43 . Deletions in codons 10 and 930, which do not result in a change in the DNA reading frane, have been subsequently described 42 .…”