scholarly journals Restarted replication forks are error-prone and cause CAG repeat expansions and contractions

PLoS Genetics ◽  
2021 ◽  
Vol 17 (10) ◽  
pp. e1009863
Author(s):  
Michaela A. Gold ◽  
Jenna M. Whalen ◽  
Karine Freon ◽  
Zixin Hong ◽  
Ismail Iraqui ◽  
...  
Keyword(s):  
Repetitive Sequences ◽  
Trinucleotide Repeats ◽  
Cag Repeat ◽  
Repeat Instability ◽  
D Loop ◽  
Repeat Expansions ◽  
Fork Stalling ◽  
Secondary Dna Structures ◽  
The Cost ◽  
Stalled Fork

Disease-associated trinucleotide repeats form secondary DNA structures that interfere with replication and repair. Replication has been implicated as a mechanism that can cause repeat expansions and contractions. However, because structure-forming repeats are also replication barriers, it has been unclear whether the instability occurs due to slippage during normal replication progression through the repeat, slippage or misalignment at a replication stall caused by the repeat, or during subsequent replication of the repeat by a restarted fork that has altered properties. In this study, we have specifically addressed the fidelity of a restarted fork as it replicates through a CAG/CTG repeat tract and its effect on repeat instability. To do this, we used a well-characterized site-specific replication fork barrier (RFB) system in fission yeast that creates an inducible and highly efficient stall that is known to restart by recombination-dependent replication (RDR), in combination with long CAG repeat tracts inserted at various distances and orientations with respect to the RFB. We find that replication by the restarted fork exhibits low fidelity through repeat sequences placed 2–7 kb from the RFB, exhibiting elevated levels of Rad52- and Rad8ScRad5/HsHLTF-dependent instability. CAG expansions and contractions are not elevated to the same degree when the tract is just in front or behind the barrier, suggesting that the long-traveling Polδ-Polδ restarted fork, rather than fork reversal or initial D-loop synthesis through the repeat during stalling and restart, is the greatest source of repeat instability. The switch in replication direction that occurs due to replication from a converging fork while the stalled fork is held at the barrier is also a significant contributor to the repeat instability profile. Our results shed light on a long-standing question of how fork stalling and RDR contribute to expansions and contractions of structure-forming trinucleotide repeats, and reveal that tolerance to replication stress by fork restart comes at the cost of increased instability of repetitive sequences.

scholarly journals Convergent Transcription through a Long CAG Tract Destabilizes Repeats and Induces Apoptosis

2010 ◽  
Vol 30 (18) ◽  
pp. 4435-4451 ◽  
Author(s):  
Yunfu Lin ◽  
Mei Leng ◽  
Ma Wan ◽  
John H. Wilson
Keyword(s):  
Cell Death ◽  
Signaling Pathway ◽  
Cellular Response ◽  
Excision Repair ◽  
Repetitive Sequences ◽  
Antisense Transcription ◽  
Cag Repeat ◽  
Cellular Stress Response ◽  
Repeat Instability ◽  
Convergent Transcription

ABSTRACT Short repetitive sequences are common in the human genome, and many fall within transcription units. We have previously shown that transcription through CAG repeat tracts destabilizes them in a way that depends on transcription-coupled nucleotide excision repair and mismatch repair. Recent observations that antisense transcription accompanies sense transcription in many human genes led us to test the effects of antisense transcription on triplet repeat instability in human cells. Here, we report that simultaneous sense and antisense transcription (convergent transcription) initiated from two inducible promoters flanking a CAG95 tract in a nonessential gene enhances repeat instability synergistically, arrests the cell cycle, and causes massive cell death via apoptosis. Using chemical inhibitors and small interfering RNA (siRNA) knockdowns, we identified the ATR (ataxia-telangiectasia mutated [ATM] and Rad3 related) signaling pathway as a key mediator of this cellular response. RNA polymerase II, replication protein A (RPA), and components of the ATR signaling pathway accumulate at convergently transcribed repeat tracts, accompanied by phosphorylation of ATR, CHK1, and p53. Cell death depends on simultaneous sense and antisense transcription and is proportional to their relative levels, it requires the presence of the repeat tract, and it occurs in both proliferating and nonproliferating cells. Convergent transcription through a CAG repeat represents a novel mechanism for triggering a cellular stress response, one that is initiated by events at a single locus in the genome and resembles the response to DNA damage.

scholarly journals Mechanism for inverted-repeat recombination induced by a replication fork barrier

2022 ◽  
Vol 13 (1) ◽  
Author(s):  
Léa Marie ◽  
Lorraine S. Symington
Keyword(s):  
Replication Fork ◽  
Repetitive Sequences ◽  
Replication Stress ◽  
Yeast Genome ◽  
Physical Analysis ◽  
Srs2 Helicase ◽  
Fork Stalling ◽  
Strand Annealing ◽  
Recombination Pathway ◽  
Stalled Replication Forks

AbstractReplication stress and abundant repetitive sequences have emerged as primary conditions underlying genomic instability in eukaryotes. To gain insight into the mechanism of recombination between repeated sequences in the context of replication stress, we used a prokaryotic Tus/Ter barrier designed to induce transient replication fork stalling near inverted repeats in the budding yeast genome. Our study reveals that the replication fork block stimulates a unique recombination pathway dependent on Rad51 strand invasion and Rad52-Rad59 strand annealing activities, Mph1/Rad5 fork remodelers, Mre11/Exo1/Dna2 resection machineries, Rad1-Rad10 nuclease and DNA polymerase δ. Furthermore, we show recombination at stalled replication forks is limited by the Srs2 helicase and Mus81-Mms4/Yen1 nucleases. Physical analysis of the replication-associated recombinants revealed that half are associated with an inversion of sequence between the repeats. Based on our extensive genetic characterization, we propose a model for recombination of closely linked repeats that can robustly generate chromosome rearrangements.

Progressive Ataxia with Hemiplegic Migraines: a Phenotype of CACNA1A Missense Mutations, Not CAG Repeat Expansions

2020 ◽  
Author(s):  
Kevin R. Duque ◽  
Luca Marsili ◽  
Andrea Sturchio ◽  
Abhimanyu Mahajan ◽  
Aristide Merola ◽  
...  
Keyword(s):  
Cag Repeat ◽  
Missense Mutations ◽  
Progressive Ataxia ◽  
Repeat Expansions

Pathogenic Mechanisms

2014 ◽  
Author(s):  
Alis Hughes ◽  
Lesley Jones
Keyword(s):  
Cag Repeat ◽  
Therapeutic Interventions ◽  
Mutant Huntingtin ◽  
Repeat Instability ◽  
Disease Etiology ◽  
Pathogenic Mechanisms ◽  
Rna Toxicity ◽  
Cellular Processes ◽  
Protein Clearance ◽  
Individual Importance

Huntington’s disease (HD) pathogenesis is complex. In the two decades since the gene and its mutation were discovered, there has been extensive exploration of how the expanded CAG repeat in HTT leads to neurodegeneration in HD. This chapter focuses on the mechanisms that potentially contribute to the dysfunction and death of cells in HD. These include repeat instability and RNA toxicity and the production, processing, modification, and degradation of mutant huntingtin. The effects of mutant HTT on cellular processes such as transcription, transport, neurotransmission, and protein clearance are also described. The interdependence and individual importance of these mechanisms in disease etiology remains to be clarified; however, consideration of each could be important for the development of therapeutic interventions in HD.

CAG repeat expansions in patients with sporadic cerebellar ataxia

1998 ◽  
Vol 98 (1) ◽  
pp. 55-59 ◽  
Author(s):  
N. Futamura ◽  
R. Matsumura ◽  
Y. Fujimoto ◽  
H. Horikawa ◽  
A. Suzumura ◽  
...  
Keyword(s):  
Cerebellar Ataxia ◽  
Cag Repeat ◽  
Repeat Expansions

scholarly journals Postreplication Repair Inhibits CAG · CTG Repeat Expansions in Saccharomyces cerevisiae

2006 ◽  
Vol 27 (1) ◽  
pp. 102-110 ◽  
Author(s):  
Danielle L. Daee ◽  
Tony Mertz ◽  
Robert S. Lahue
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Trinucleotide Repeats ◽  
Dna Microsatellites ◽  
Dinucleotide Repeats ◽  
Replication Slippage ◽  
Postreplication Repair ◽  
Epistasis Analysis ◽  
Repeat Expansions ◽  
The Relationship ◽  
Ctg Repeat

ABSTRACTTrinucleotide repeats (TNRs) are unique DNA microsatellites that can expand to cause human disease. Recently, Srs2 was identified as a protein that inhibits TNR expansions inSaccharomyces cerevisiae. Here, we demonstrate that Srs2 inhibits CAG · CTG expansions in conjunction with the error-free branch of postreplication repair (PRR). Likesrs2mutants, expansions are elevated inrad18andrad5mutants, as well as the PRR-specific PCNA allelespol30-K164Randpol30-K127/164R. Epistasis analysis indicates that Srs2 acts upstream of these PRR proteins. Also, likesrs2mutants, thepol30-K127/164Rphenotype is specific for expansions, as this allele does not alter mutation rates at dinucleotide repeats, at nonrepeating sequences, or for CAG · CTG repeat contractions. Our results suggest that Srs2 action and PRR processing inhibit TNR expansions. We also investigated the relationship between PRR and Rad27 (Fen1), a well-established inhibitor of TNR expansions that acts at 5′ flaps. Our results indicate that PRR protects against expansions arising from the 3′ terminus, presumably replication slippage events. This work provides the first evidence that CAG · CTG expansions can occur by 3′ slippage, and our results help define PRR as a key cellular mechanism that protects against expansions.

scholarly journals Genetic Contributors to Intergenerational CAG Repeat Instability in Huntington’s Disease Knock-In Mice

Genetics ◽  
2016 ◽  
Vol 205 (2) ◽  
pp. 503-516 ◽  
Author(s):  
João Luís Neto ◽  
Jong-Min Lee ◽  
Ali Afridi ◽  
Tammy Gillis ◽  
Jolene R. Guide ◽  
...  
Keyword(s):  
Huntington's Disease ◽  
Huntington’S Disease ◽  
Cag Repeat ◽  
Repeat Instability
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