Review on Molecular Diagnostic Techniques in Friedreich’s Ataxia.

Friedreich’s ataxia is a commonly inherited neurodegenerative disease with an autosomal recessive pattern of inheritance, and was described by Nikolaus Friedreich first in 1863. Friedreich’s ataxia is caused due to hyperexpansion of the intronic GAA trinucleotide repeats or mutations in the FXN gene on chromosome 9q13. This gene codes for a mitochondrial protein, frataxin, which is highly conserved in many species and has functions in iron-sulfur cluster biosynthesis. Friedreich’s ataxia mainly results from a deficiency of the frataxin protein, due to mutations in the FXN gene. Formation of sticky DNA, formation of DNA-RNA hybrid and epigenetic changes, including methylation of DNA and histone modifications, are the proposed mechanisms for disruption of FXN gene expression. Most cases of Friedreich’s ataxia are homozygous and caused due to expansion of the GAA trinucleotide repeat in the first intron of the FXN gene, however, some cases can be heterozygous, with GAA expansion in one allele and point mutation or deletion in the FXN gene on the other allele. Therefore, diagnosis of the disease based on only the clinical symptoms becomes difficult. Molecular diagnosis is, therefore, important, in order to detect GAA repeat expansions as well as mutations in the FXN gene. This review represents an overview of the molecular diagnostic studies in Friedreich’s ataxia, including an overview of the disease, as well as the gene and protein involved in the disease and techniques that can be useful in diagnosis of the Friedreich’s ataxia. The described methods include tools that are based on analysis of DNA as well as analysis of mRNA and protein levels. A brief description of mutations found in compound heterozygous Friedreich’s ataxia patients, is also provided.

DNA triplex structures in neurodegenerative disorder, Friedreich’s ataxia.

It is now established that a small fraction of genomic DNA does adopt the non-canonical B-DNA structure or ‘unusual’ DNA structure. The unusual DNA structures like DNA-hairpin, cruciform, Z-DNA, triplex and tetraplex are represented as hotspots of chromosomal breaks, homologous recombination and gross chromosomal rearrangements since they are prone to the structural alterations. Friedreich’s ataxia (FRDA), the autosomal recessive degenerative disorder of nervous and muscles tissue, is caused by the massive expansion of (GAA) repeats that occur in the first intron of Frataxin gene X25 on chromosome 9q13-q21.1. The purine strand of the DNA in the expanded (GAA) repeat region folds back to form the (R∙R*Y) type of triplex, which further inhibits the frataxin gene expression, and this clearly suggests that the shape of DNA is the determining factor in the cellular function. FRDA is the only disease known so far to be associated with DNA triplex. Structural characterization of GAA-containing DNA triplexes using some simple biophysical methods like UV melting, UV absorption, circular dichroic spectroscopy and electrophoretic mobility shift assay are discussed. Further, the clinical aspects and genetic analysis of FRDA patients who carry (GAA) repeat expansions are presented. The potential of some small molecules that do not favour the DNA triplex formation as therapeutics for FRDA are also briefly discussed.