scholarly journals Approaches to Sequence the HTT CAG Repeat Expansion and Quantify Repeat Length Variation

2021 ◽  
Vol 10 (1) ◽  
pp. 53-74
Author(s):  
Marc Ciosi ◽  
Sarah A. Cumming ◽  
Afroditi Chatzi ◽  
Eloise Larson ◽  
William Tottey ◽  
...  
Keyword(s):  
Neurodegenerative Disorder ◽  
Disease Onset ◽  
Somatic Mosaicism ◽  
Genotyping By Sequencing ◽  
Repeat Length ◽  
Cag Repeat ◽  
Length Variation ◽  
Allele Size ◽  
Pcr Products ◽  
Alternative Approaches

Background: Huntington’s disease (HD) is an autosomal dominant neurodegenerative disorder caused by the expansion of the HTT CAG repeat. Affected individuals inherit ≥36 repeats and longer alleles cause earlier onset, greater disease severity and faster disease progression. The HTT CAG repeat is genetically unstable in the soma in a process that preferentially generates somatic expansions, the proportion of which is associated with disease onset, severity and progression. Somatic mosaicism of the HTT CAG repeat has traditionally been assessed by semi-quantitative PCR-electrophoresis approaches that have limitations (e.g., no information about sequence variants). Genotyping-by-sequencing could allow for some of these limitations to be overcome. Objective: To investigate the utility of PCR sequencing to genotype large (>50 CAGs) HD alleles and to quantify the associated somatic mosaicism. Methods: We have applied MiSeq and PacBio sequencing to PCR products of the HTT CAG repeat in transgenic R6/2 mice carrying ∼55, ∼110, ∼255 and ∼470 CAGs. For each of these alleles, we compared the repeat length distributions generated for different tissues at two ages. Results: We were able to sequence the CAG repeat full length in all samples. However, the repeat length distributions for samples with ∼470 CAGs were biased towards shorter repeat lengths. Conclusion: PCR sequencing can be used to sequence all the HD alleles considered, but this approach cannot be used to estimate modal allele size or quantify somatic expansions for alleles ⪢250 CAGs. We review the limitations of PCR sequencing and alternative approaches that may allow the quantification of somatic contractions and very large somatic expansions.

scholarly journals Library preparation and MiSeq sequencing for the genotyping-by-sequencing of the Huntington disease HTT exon one trinucleotide repeat and the quantification of somatic mosaicism

2020 ◽  
Author(s):  
Marc Ciosi ◽  
Sarah A. Cumming ◽  
Asma M. Alshammari ◽  
Efthymia Symeonidi ◽  
Pawel Herzyk ◽  
...  
Keyword(s):  
Huntington Disease ◽  
Trinucleotide Repeat ◽  
Neurodegenerative Disorder ◽  
Fragment Size ◽  
Somatic Mosaicism ◽  
Genotyping By Sequencing ◽  
Amplicon Sequencing ◽  
Cag Repeat ◽  
Miseq Sequencing ◽  
Flanking Sequence

Abstract Huntington disease \(HD) is an autosomal dominant neurodegenerative disorder caused by the expansion of a CAG repeat in the first exon of the _HTT_ gene. Affected individuals inherit more than 40 repeats and the CAG repeat is genetically unstable in both the germline and soma. Molecular diagnosis and genotyping of the CAG repeat is traditionally performed by estimation of PCR fragment size. However, this approach is complicated by the presence of an adjacent polymorphic CCG repeat and provides no information on the presence of variant repeats, flanking sequence variants or on the degree of somatic mosaicism. To overcome these limitations, we have developed an amplicon-sequencing protocol that allows the sequencing of hundreds of samples in a single MiSeq run. The composition of the _HTT_ exon one trinucleotide repeat locus can be determined from the MiSeq sequencing reads generated. With sufficient sequencing depth, such MiSeq data can also be used to quantify the degree of somatic mosaicism of the _HTT_ CAG repeat in the tissue analysed.

scholarly journals Predicting Disease Onset from Mutation Status Using Proband and Relative Data with Applications to Huntington's Disease

2012 ◽  
Vol 2012 ◽  
pp. 1-19 ◽  
Author(s):  
Tianle Chen ◽  
Yuanjia Wang ◽  
Yanyuan Ma ◽  
Karen Marder ◽  
Douglas R. Langbehn
Keyword(s):  
Huntington's Disease ◽  
Huntington’S Disease ◽  
Family Member ◽  
Disease Onset ◽  
Repeat Length ◽  
Cag Repeat ◽  
Cag Repeats ◽  
Observational Research ◽  
Accurate Estimation ◽  
Combined Data

Huntington's disease (HD) is a progressive neurodegenerative disorder caused by an expansion of CAG repeats in the IT15 gene. The age-at-onset (AAO) of HD is inversely related to the CAG repeat length and the minimum length thought to cause HD is 36. Accurate estimation of the AAO distribution based on CAG repeat length is important for genetic counseling and the design of clinical trials. In the Cooperative Huntington's Observational Research Trial (COHORT) study, the CAG repeat length is known for the proband participants. However, whether a family member shares the huntingtin gene status (CAG expanded or not) with the proband is unknown. In this work, we use the expectation-maximization (EM) algorithm to handle the missing huntingtin gene information in first-degree family members in COHORT, assuming that a family member has the same CAG length as the proband if the family member carries a huntingtin gene mutation. We perform simulation studies to examine performance of the proposed method and apply the methods to analyze COHORT proband and family combined data. Our analyses reveal that the estimated cumulative risk of HD symptom onset obtained from the combined data is slightly lower than the risk estimated from the proband data alone.

scholarly journals CAG repeat length variation in the polymerase gamma (POLG) gene: effect on semen quality

2008 ◽  
Vol 14 (4) ◽  
pp. 245-249 ◽  
Author(s):  
G.H. Westerveld ◽  
L. Kaaij-Visser ◽  
M. Tanck ◽  
F. van der Veen ◽  
S. Repping
Keyword(s):  
Semen Quality ◽  
Repeat Length ◽  
Cag Repeat ◽  
Length Variation ◽  
Polymerase Gamma ◽  
Gene Effect ◽  
Polg Gene

scholarly journals Expanded huntingtin CAG repeats disrupt the balance between neural progenitor expansion and differentiation in human cerebral organoids

2019 ◽  
Author(s):  
Jinqiu Zhang ◽  
Jolene Ooi ◽  
Kagistia Hana Utami ◽  
Sarah R. Langley ◽  
Obed Akwasi Aning ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Neurodegenerative Disorder ◽  
Ventricular Zone ◽  
Neural Progenitor ◽  
Repeat Length ◽  
Cag Repeat ◽  
Cag Repeats ◽  
Normal Brain ◽  
Dependent Manner ◽  
The Impact

AbstractHuntington disease (HD) manifests in both adult and juvenile forms. Mutant HTT gene carriers are thought to undergo normal brain development followed by a degenerative phase, resulting in progressive clinical manifestations. However, recent studies in children and prodromal individuals at risk for HD have raised the possibility of abnormal neurodevelopment. Although key findings in rodent models support this notion, direct evidence in the context of human physiology remains lacking. Using a panel of isogenic HD human embryonic pluripotent stem cells and cerebral organoids, we investigated the impact of mutant HTT on early neurodevelopment. We find that ventricular zone-like neuroepithelial progenitor layer expansion is blunted in an HTT CAG repeat length-dependent manner due to premature neurogenesis in HD cerebral organoids, driven by cell intrinsic processes. Transcriptional profiling and imaging analysis revealed impaired cell cycle regulatory processes, increased G1 length, and increased asymmetric division of apical progenitors, collectively contributing to premature neuronal differentiation. We demonstrate increased activity of the ATM-p53 pathway, an up-stream regulator of cell cycle processes, and show that treatment with ATM antagonists partially rescues the blunted neuroepithelial progenitor expansion in HD organoids. Our findings suggest that CAG repeat length regulates the balance between neural progenitor expansion and differentiation during early neurodevelopment. Our results further support the view that HD, at least in its early-onset forms, may not be a purely neurodegenerative disorder, and that abnormal neurodevelopment may be a component of HD pathophysiology.

CAG repeat length variation in sperm from a patient with Kennedy's disease

1995 ◽  
Vol 4 (2) ◽  
pp. 303-305 ◽  
Author(s):  
Lin Zhang ◽  
Kenneth H. Fischbeck ◽  
Norman Arnhelm
Keyword(s):  
Repeat Length ◽  
Cag Repeat ◽  
Length Variation ◽  
Kennedy’S Disease ◽  
Kennedy's Disease

CAG repeat length variation in the Androgen Receptor gene is not associated with spermatogenic failure

2008 ◽  
Vol 89 (1) ◽  
pp. 253-259 ◽  
Author(s):  
Henrike Westerveld ◽  
Liesbeth Visser ◽  
Michael Tanck ◽  
Fulco van der Veen ◽  
Sjoerd Repping
Keyword(s):  
Androgen Receptor ◽  
Receptor Gene ◽  
Androgen Receptor Gene ◽  
Repeat Length ◽  
Cag Repeat ◽  
Length Variation ◽  
Spermatogenic Failure

An investigation into CAG repeat length variation and N/C terminal interactions in the T877A mutant androgen receptor found in prostate cancer

2008 ◽  
Vol 111 (1-2) ◽  
pp. 138-146 ◽  
Author(s):  
Jason Southwell ◽  
Shafinaz F. Chowdhury ◽  
Bruce Gottlieb ◽  
Lenore K. Beitel ◽  
Rose Lumbroso ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Repeat Length ◽  
Cag Repeat ◽  
Length Variation

scholarly journals Molecular pathology of dentatorubral--pallidoluysian atrophy

1999 ◽  
Vol 354 (1386) ◽  
pp. 1069-1074 ◽  
Author(s):  
Ichiro Kanazawa
Keyword(s):  
Age Of Onset ◽  
Neuronal Cell Death ◽  
Somatic Mosaicism ◽  
Neuronal Cell ◽  
Repeat Length ◽  
Cag Repeat ◽  
Autosomal Dominant Disorder ◽  
Myoclonus Epilepsy ◽  
Chromosome 12P ◽  
Sporadic Cases

Dentatorubral–pallidoluysian atrophy (DRPLA) is an autosomal dominant disorder characterized clinically by myoclonus, epilepsy, cerebellar ataxia, choreoathetosis and dementia. Cardinal pathological features of DRPLA are a combined degeneration of both the dentatorubral and the pallidoluysian systems. Although the early sporadic cases were reported by Western neuropathologists, a strong heritability and an age of onset–dependent variability of the clinical features were carefully deduced by Japanese clinicians. The disease is fairly common in Japan, but extremely rare in Caucasians. Since the gene was identified in 1994, DRPLA is known as one of the CAG repeat expansion diseases, in which the responsible gene is located on chromosome 12p and its product is called atrophin 1. DRPLA shows prominent ‘anticipation’, which is genetically clearly explained by a marked instability of the expanded CAG repeat length during spermatogenesis. Moreover, the instability of the CAG repeat length also seems to occur in the somatic cells, resulting in ‘somatic mosaicism’. Possible mechanism(s) underlying the neuronal cell death in DRPLA are discussed in terms of molecular pathological points of view.

scholarly journals Identification of Differentially Expressed Genes in Prostatic Epithelium in Relation to Androgen Receptor CAG Repeat Length

2006 ◽  
Vol 21 (2) ◽  
pp. 96-105 ◽  
Author(s):  
C.M. Coutinho-Camillo ◽  
E.C. Miracca ◽  
M.L. Dos Santos ◽  
S. Salaorni ◽  
A.S. Sarkis ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Differentially Expressed Genes ◽  
Repeat Length ◽  
Cag Repeat ◽  
Differentially Expressed ◽  
Cag Repeats ◽  
Pcr Analysis ◽  
Length Variation ◽  
Rt Pcr

The CAG repeat within exon 1 of the androgen receptor (AR) has been associated with the development of prostate cancer. The shorter number of glutamine residues in the protein has been associated with a higher transcriptional activity of the AR and increased relative risk for prostate cancer. In an attempt to identify differentially expressed genes in prostate cancer in relation to AR CAG repeat length variation, in this study we used total mRNA from normal and tumor tissues from 2 prostate cancer patients with AR alleles containing 19 and 26 CAG repeats to perform differential-display RT-PCR analysis. We were able to identify 48 different transcripts that showed homology to several known genes associated with different biological pathways. Among the differentially expressed genes, ATRX and SFRP1 were further validated by quantitative RT-PCR. The transcripts of both ATRX and SFRP1 genes proved to be down-regulated in most of the prostate tumors analyzed by quantitative RT-PCR. Hypermethylation of the promoter region of the SFRP1 gene was found in 17.5% (7/40) of the cases analyzed and was associated with the loss of SFRP1 expression (p=0.014). The differentially expressed genes identified in this study are implicated in several cellular pathways that, when up- or down-regulated, might play a role in the tumorigenic process of the prostate.

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