Abstract
Genomic technologies such as next-generation sequencing (NGS) are revolutionizing molecular diagnostics and clinical medicine. However, these approaches have proven inefficient at identifying pathogenic repeat expansions. Here, we apply a collection of bioinformatics tools that can be utilized to identify either known or novel expanded repeat sequences in NGS data. We performed genetic studies of a cohort of 35 individuals from 22 families with a clinical diagnosis of cerebellar ataxia with neuropathy and bilateral vestibular areflexia syndrome (CANVAS). Analysis of whole-genome sequence (WGS) data with five independent algorithms identified a recessively inherited intronic repeat expansion [(AAGGG)exp] in the gene encoding Replication Factor C1 (RFC1). This motif, not reported in the reference sequence, localized to an Alu element and replaced the reference (AAAAG)11 short tandem repeat. Genetic analyses confirmed the pathogenic expansion in 18 of 22 CANVAS-affected families and identified a core ancestral haplotype, estimated to have arisen in Europe more than twenty-five thousand years ago. WGS of the four RFC1-negative CANVAS-affected families identified plausible variants in three, with genomic re-diagnosis of SCA3, spastic ataxia of the Charlevoix-Saguenay type, and SCA45. This study identified the genetic basis of CANVAS and demonstrated that these improved bioinformatics tools increase the diagnostic utility of WGS to determine the genetic basis of a heterogeneous group of clinically overlapping neurogenetic disorders.
Highlights
Repetitive DNA sequences constitute approximately one third of the genome and are thought to contribute to diversity within and between species.[1]
While variable between case subjects, data leading to the clinical diagnosis included evidence of combined cerebellar and bilateral vestibular impairment, cerebellar atrophy on MRI, neurophysiological evidence of impaired sensory nerve function, and negative genetic testing for pathogenic repeat expansions (REs) at common spinocerebellar ataxia (SCA) loci and FRDA (Friedreich ataxia, FRDA)
Since the first description of the syndrome of cerebellar ataxia with bilateral vestibulopathy in 200446 and proposal of CANVAS as a distinct clinical entity in 2011,11 there has been little progress made in delineating the etiology of the disorder
Summary
Repetitive DNA sequences constitute approximately one third of the genome and are thought to contribute to diversity within and between species.[1] Microsatellites or short tandem repeats (STRs) are mini-repeats of DNA, typically two to five base-pairs in length, which are usually present in a concatamer of between 5 and 50 repeated elements. There are thousands of STRs scattered through the human genome and recent studies have suggested important roles for STRs in the regulation of gene expression.[2,3] STRs display considerable variability in length between individuals, which is presumed to have no detrimental consequences for humans[4,5] unless the repeat number is expanded beyond a gene-specific threshold.[6,7] Pathogenic repeat expansions (REs) have been shown to underlie at.
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