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Molecular diagnostics

Dr. István Balogh, Dr. János Kappelmayer, Dr. József Tőzsér (2011)

University of Debrecen

Chapter 5. 5. Genetic diseases caused by expandable repeats - dynamic mutations

Chapter 5. 5. Genetic diseases caused by expandable repeats - dynamic mutations

Table of Contents

In addition to the classical monogenic genetic disorders that show Mendelian inheritance, there exists another group of genetic diseases, in which certain, meiotically unstable DNA repeats are the causing factors. A typical phenomenon to be observed in these disorders is anticipation, which means that the phenotype will be increasingly more severe in subsequent generations or it will appear at an increasingly younger age. These repeats form the mutation category called dynamic mutations. The number of repeats must normally reach a threshold value for the disease phenotype to appear. To date, approximately 30 such diseases are known (Figure 5.1.).

Figure 5.1. Figure 5.1. Genetic diseases that are caused by expandable repeats

Figure 5.1. Genetic diseases that are caused by expandable repeats

Most of the cases show triplet expansions, but tetranucleotide, pentanucleotide and even dodecanucleotide expansions are possible as well. These unstable repeats might be present in different functionally important areas of the gene, like in the 5’ untranslated sequence region, in the introns, in the coding region, in the 3’ untranslated sequence region as well as in the promoter. The mechanism that leads to disease largely depends on where the expansion occurred in the gene.

  • The repeat expansion in the 5’ untranslated sequence causes promoter methylation in the case of fragile X syndrome, and blocks the transcription of the gene.

  • The CTG expansion in the case of dystrophia myotonica (myotonic dystrophy) in the 3’ end of the gene induces CUG-binding splicing factors in the mRNA, which will interfere with the correct splicing processes in some independent genes.

  • CAG expansion in the coding region will result in the expression of toxic protein product in Huntington’s disease.

Abbreviations (Figure 5.1.):

EPM1: progressive myoclonic epilepsy 1, FRAXA: fragile X syndrome, FRAXE: fragile X mental retardation associated with FRAXE site, FXTAS: fragile X tremor and ataxia syndrome, SCA: spinocerebellar ataxia, DRPLA: dentatorubral-pallidoluysian atrophy, HD: Huntington’s disease, SBMA: spinal and bulbar muscular atrophy, FRDA: Friedreich ataxia, DM: myotonic dystrophy, BPES: blepharophimosis, CCD: cleidocranial dysplasia, CCHS: congenital central hypoventilation syndrome, HFG: hand-foot-genital syndrome, HPE5: holoprosencephaly 5, ISSX: X-linked infantile spasm syndrome, MRGH: mental retardation with isolated growth hormone deficiency, OPMD: oculopharyngeal muscular dystrophy, SPD: synpolydactyly, HDL2: Huntington’s disease-like 2.

The number of the repeats shows extreme variability. In the FRAXA gene, the normal CGG repeat number is between 6 and 54. This can be expanded from 200 to 1000 in the case of unstable repeats. Myotonic dystrophy is a similar case. In healthy individuals, the repeat CTG number is between 5 and 37, which can be expanded to 50-10000 in patients. Because of the variable scale of the change in the number of repeats, the molecular testing of these disorders is very difficult. Different methodologies have been developed, such as special PCR assays and Southern blot.

Expansions occurring in the coding region of the affected gene are usually shorter than the ones located in the regulatory elements. In the case of Huntington's disease, the number of original (normal) 6-34 CAG repeats can reach 36-100 in unstable cases. Repeat number in one of the spinocerebellar ataxia type (SCA6) shows only a small increase, from 4-17 in normal cases to 33 in affected individuals.

An important question to ask is what kind of molecular mechanisms play a role in the development of unstable repeats. It is thought that the instability might be a consequence of the disturbances in the DNA repair mechanisms and the recombination system. The repeats that are prone to instability form several unusual DNS structures (Figure 5.2.)

Figure 5.2. Figure 5.2. Unusual DNA structures caused by expandable repeats

Figure 5.2. Unusual DNA structures caused by expandable repeats

DNA repeats are capable of forming several unusual structures:

  1. imperfect hairpin structure formed by (CNG)n repeats

  2. (CGG)n repeats that form quadruplex-like structure

  3. Structures that are caused by (CTG)n (CAG)n repeats

  4. H-DNA and sticky DNA that are caused by (GAA)n (TTC)n repeats

  5. Structures that are caused by (ATTCT)n (AGAAT)n repeats

The repeats which do not form structures are much more stable genetically. The unusual DNA structures add a significant component to the instability of the DNA repeats, thus they contribute to a large extent to the development of the disease.