Biology of Disease

CHROMOSOMES AND THE HUMAN KARYOTYPE

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BOX 15.2 Dynamic mutations and DNA methylation

In the early 1990s types of mutation called dynamic or expansion
mutations were identified that were associated with a number
of genetic disorders that increased in severity, or had an earlier
onset over several generations. These disorders are referred to as
trinucleotide repeat disorders. Repeating combinations of three
nucleotides occur commonly in DNA molecules. All combina-
tions of triplets, such as CGG, CAG, AGG and ACC, are found
but the first two are commonest. Such sequences seem to be a
normal part of the DNA and are thought to have regulatory roles
in gene expression. However, if the number of repeats becomes
too large, clinical problems that result in an identifiable disease
are triggered. Fragile X syndrome is one such disease and is asso-
ciated with an increase in the number of CGG repeats that are
normally found in the FMR1 (fragile X mental retardation 1) gene
at the Xq27.3 fragile site. One consequence of the expansion in
the number of CGG repeats is that methylation of the regulatory
region of the gene occurs and prevents the cell from express-
ingFMR1 and synthesizing fragile X mental retardation protein
(fmrp), an RNA binding protein that is expressed in the brain. The
lack of fmrp leads to fragile X syndrome. Methylation of DNA is
the addition of methyl (–CH 3 ) groups (Chapter 10) to some of its
bases from the donor molecule, S-adenosylmethionine (SAM).
Only a few percent of A and C nucleotides are methylated and
in vertebrate eukaryotic cells only the formation of 5-methyl-
cytosine (Me^5 C) occurs (Figure 15.22). Methylation at particular
CpG sequences may inhibit transcription and have a role in gene
regulation by switching off the expression of that gene, although
the mechanism by which this occurs is unclear. However, the

methyl group is known to project into the DNA molecule and

interfere with the attachment of DNA binding proteins. The

patterns of methylation are inherited, that is they are repeated

from generation to generation. In general, FMR1 is copied by the

DNA polymerase during DNA replication with high fidelity and

this stable version is almost always inherited. However, in some

circumstances amplification occurs that increases the number

of CGG repeats in the daughter chromosome. This is possible

because repeats containing G and C nucleotides can base pair

with themselves to form hairpin structures (Figure 15.23), which

increases the risk of slipped mispairing occurring during DNA

replication leading to an increase in the number of triplet repeats.

The number of CGG repeats in the FMR1 is the major factor that

determines the presence or absence of fragile X syndrome. In

fragile X syndrome, the mutant FMR1 gene is nonmethylated in

asymptomatic males, methylated in the inactive X chromosome

of females and totally methylated in most fragile X males, which

prevents its expression.

H

NH 2

O

CH 3

N

N

DNA

H

H

NH 2

O

N

N

DNA

Cytosine nucleotide

SAM

5-Methylcytosine

nucleotide (m^5 C)

Figure 15.22 Schematic illustrating the methylation of a

cytosine base in DNA to 5-methylcytosine. SAM,

S-adenosylmethionine.

... G C G G C G G C G G C G G C G G C G G C G G...

C

C

G

C

G

C

C

C

G

G

G

G...

G

G

G

G

G

C G

G

... G G

Hairpin

bend

formation

Figure 15.23 A highly schematic

representation of a hairpin bend formed

by a series of repeated CGG trinucleotides.