Methylation of the G-C rich DMPK gene: causation for the severity difference between adultonset and congenital myotonic dystrophy?
Date
2010-05
Authors
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Journal ISSN
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Publisher
University of Delaware
Abstract
Myotonic dystrophy type I is an autosomal dominant genetic disease. The
disease arises from an expansion of a trinucleotide repeat, (CTG)n located in the 3’
untranslated (3’ UTR) region of the dystrophica myotonin protein kinase (DMPK)
gene on chromosome 19q13.2-q13.3. Located proximal to this region is a CpG island,
a region of DNA with a high concentration of CpG sequences that often associate with
gene promoters in humans. Changes in these promoter regions, including the
methylation status, modify expression of genes that they regulate. The purpose of this
study was to test the hypothesis that there are allelic differences in the methylation
status of this CpG island in normal, adult-onset and congenital myotonic dystrophy
patients by using DNA prepared from somatic cell hybrids that carry either the normal
or expanded DMPK allele. In addition, once the methylation status had been
ascertained, it was the goal to determine if hypermethylation of the normal allele
correlated with a decreased expression of the DMPK gene. This correlation could
suggest the reason for severity differences between adult-onset and congenital DM1.
The method of bisulfite-conversion was used to ascertain the methylation status of the
CpG island. This method converts all unmethylated cytosine residues into uracil
residues. The converted DNA of congenital DM1 patients was then amplified by
means of High-Fidelity PCR where the converted uracil residues are amplified as
thymine residues. This amplified DNA is then cloned and sequenced to compare the
methylation status of the CpG island among individuals. After examining the methylation status of this CpG island in one CDM1 patient and a patient presenting
with signs and symptoms associated with DM1, it was determined that the non-mutant
allele of these patients is not methylated. Since the non-mutant allele was determined
to be unmethylated, hypermethylation could not be the cause of its decreased
expression.