Msh2-Msh3 Interferes with Okazaki Fragment Processing to Promote Trinucleotide Repeat Expansions

Kantartzis, Athena; Williams, Gregory M.; Balakrishnan, Lata; Roberts, Rick L.; Surtees, Jennifer A.; Bambara, Robert A.

doi:10.1016/j.celrep.2012.06.020

Cited by 43 publications

(90 citation statements)

References 39 publications

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“…Since no expansion was detected in the CTG orientation, and only one in the CAG orientation, it was not possible to address the effect of replication pausing on triplet repeat expansions. This low level of expansions is similar to what was observed by other authors in recent publications [40,49].…”

Section: Replication Fork Stalling Occurs In Both Stable and Unstablesupporting

confidence: 80%

“…All these data point to a model in which MMR complexes recognize and bind secondary structures associated to CAG/CTG repeats, maybe stabilizing these structures, and that such binding leads to expansions, by a process requiring a functional MMR activity. In support of this model, it was shown that purified yeast inhibits Rad27p cleavage at the base of repeat-containing 5' flaps, suggesting that this inhibition eventually leads to repeat expansions in vivo [40].…”

Section: Introductionmentioning

confidence: 83%

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Replication stalling and heteroduplex formation within CAG/CTG trinucleotide repeats by mismatch repair

et al. 2016

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2 Highlights-CAG/CTG triplet repeats block replication in both orientations on a yeast chromosome -Replication fork stalling depends on mismatch repair integrity -Msh2p is enriched at CAG/CTG triplet repeats in both orientations -MSH2 overexpression favors or stabilizes the formation of heteroduplex molecules 3 AbstractTrinucleotide repeat expansions are responsible for at least two dozen neurological disorders.Mechanisms leading to these large expansions of repeated DNA are still poorly understood. It was proposed that transient stalling of the replication fork by the repeat tract might trigger slippage of the newly-synthesized strand over its template, leading to expansions or contractions of the triplet repeat. However, such mechanism was never formally proven. Here we show that replication fork pausing and CAG/CTG trinucleotide repeat instability are not linked, stable and unstable repeats exhibiting the same propensity to stall replication forks when integrated in a yeast natural chromosome. We found that replication fork stalling was dependent on the integrity of the mismatch-repair system, especially the Msh2p-Msh6p complex, suggesting that direct interaction of MMR proteins with secondary structures formed by trinucleotide repeats in vivo, triggers replication fork pauses. We also show by chromatin immunoprecipitation that Msh2p is enriched at trinucleotide repeat tracts, in both stable and unstable orientations, this enrichment being dependent on MSH3 and MSH6.Finally, we show that overexpressing MSH2 favors the formation of heteroduplex regions, leading to an increase in contractions and expansions of CAG/CTG repeat tracts during replication, these heteroduplexes being dependent on both MSH3 and MSH6. These heteroduplex regions were not detected when a mutant msh2-E768A gene in which the ATPase domain was mutated was overexpressed. Our results unravel two new roles for mismatch-repair proteins: stabilization of heteroduplex regions and transient blocking of replication forks passing through such repeats. Both roles may involve direct interactions between MMR proteins and secondary structures formed by trinucleotide repeat tracts, although indirect interactions may not be formally excluded.

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Section: Replication Fork Stalling Occurs In Both Stable and Unstablesupporting

confidence: 80%

Section: Introductionmentioning

confidence: 83%

Replication stalling and heteroduplex formation within CAG/CTG trinucleotide repeats by mismatch repair

et al. 2016

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“…As the tract length increased in sperm cells, there was significant bias toward expansions vs. contractions. The sizes of the observed expansion and contraction events were similar to our in vitro observations (one-two repeats) (Kantartzis et al 2012), in contrast to larger expansion events observed in nondividing cells or postmitotic neurons (McMurray 2010).One factor known to contribute to (CNG) n tract expansions is the mismatch repair (MMR) complex Msh2-Msh3 (MutSb in mammals). Typically, Msh2-Msh3 recognizes and binds insertion-deletion loops (IDLs) that result from DNA polymerase slippage events, often within repetitive sequences (Lovett 2004;Li 2008).…”

supporting

confidence: 84%

“…Similarly, Msh3 promotes expansions in human cells (Gannon et al 2012;Halabi et al 2012). We recently demonstrated a significant decrease in expansion of both CAG and CTG repeat tracts in Saccharomyces cerevisiae in an msh3D background (Kantartzis et al 2012).The role that Msh2-Msh3 plays in promoting TNR expansions remains unclear. Our in vitro results indicate that yeast Msh2-Msh3 interferes with proper Okazaki fragment processing by Rad27 (Fen1) and Cdc9 (Lig1) in the presence of a dynamic CTG or CAG repeat tract, leading to small incremental expansions and providing mechanistic insight into the role of Msh2-Msh3 in replication-dependent TNR expansions (Kantartzis et al 2012).…”

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confidence: 99%

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MSH3 Promotes Dynamic Behavior of Trinucleotide Repeat Tracts In Vivo

Williams

Surtees

2015

Genetics

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Trinucleotide repeat (TNR) expansions are the underlying cause of more than 40 neurodegenerative and neuromuscular diseases, including myotonic dystrophy and Huntington's disease, yet the pathway to expansion remains poorly understood. An important step in expansion is the shift from a stable TNR sequence to an unstable, expanding tract, which is thought to occur once a TNR attains a threshold length. Modeling of human data has indicated that TNR tracts are increasingly likely to expand as they increase in size and to do so in increments that are smaller than the repeat itself, but this has not been tested experimentally. Genetic work has implicated the mismatch repair factor MSH3 in promoting expansions. Using Saccharomyces cerevisiae as a model for CAG and CTG tract dynamics, we examined individual threshold-length TNR tracts in vivo over time in MSH3 and msh3D backgrounds. We demonstrate, for the first time, that these TNR tracts are highly dynamic. Furthermore, we establish that once such a tract has expanded by even a few repeat units, it is significantly more likely to expand again. Finally, we show that threshold-length TNR sequences readily accumulate net incremental expansions over time through a series of small expansion and contraction events. Importantly, the tracts were substantially stabilized in the msh3D background, with a bias toward contractions, indicating that Msh2-Msh3 plays an important role in shifting the expansioncontraction equilibrium toward expansion in the early stages of TNR tract expansion.KEYWORDS Saccharomyces cerevisiae; Msh2-Msh3; mismatch repair; trinucleotide repeat tract T HE EXPANSION of trinucleotide repeat (TNR) sequences is the underlying cause of over 40 neurodegenerative and neuromuscular diseases (Castel et al. 2010;McMurray 2010). TNR sequences made of (CNG) n repeats are of particular interest because of their role in causing Huntington's disease (HD) and myotonic dystrophy type 1 (DM1), as well as a number of other diseases (McMurray 2010). TNR tracts within the normal range (which is tract dependent) are stably maintained within that range (Figure 1). However, through a mechanism(s) that remains unclear, a TNR tract can expand, increasing the number of repeats within the tract. Initially, this brings the tract into a threshold-length range (Gannon et al. 2012;Concannon and Lahue 2014) (Figure 1 and Figure 2), in which these somewhat longer tracts are not pathogenic but are increasingly susceptible to expansion; individuals with this phenomenon are carriers for disease. Once a tract has expanded sufficiently, it crosses a threshold; tracts above this threshold (which is disease specific) are pathogenic and cause disease ( Figure 1). As the size of the tract increases, it becomes increasingly unstable and prone to changes in length, particularly expansions.The dynamic behavior of TNR tracts that are within the threshold range (i.e., more susceptible to expansions) and the manner in which they occur in vivo remain unclear. Studies of TNR tract length changes...

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Measuring Dynamic Behavior of Trinucleotide Repeat Tracts In Vivo in Saccharomyces cerevisiae

Williams

Surtees

2017

Methods in Molecular Biology

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Trinucleotide repeat (TNR) tracts are inherently unstable during DNA replication, leading to repeat expansions and/or contractions. Expanded tracts are the cause of over 40 neurodegenerative and neuromuscular diseases. In this chapter, we focus on the (CNG) repeat sequences that, when expanded, lead to Huntington's disease (HD), myotonic dystrophy type 1 (DM1), and a number of other neurodegenerative diseases. We describe a series of in vivo assays, using the model system Saccharomyces cerevisiae, to determine and characterize the dynamic behavior of TNR tracts that are in the early stages of expansion, i.e., the so-called threshold range. Through a series of time courses and PCR-based assays, dynamic changes in tract length can be observed as a function of time. These assays can ultimately be used to determine how genetic factors influence the process of tract expansion in these early stages.

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Msh2-Msh3 Interferes with Okazaki Fragment Processing to Promote Trinucleotide Repeat Expansions

Cited by 43 publications

References 39 publications

Replication stalling and heteroduplex formation within CAG/CTG trinucleotide repeats by mismatch repair

Replication stalling and heteroduplex formation within CAG/CTG trinucleotide repeats by mismatch repair

MSH3 Promotes Dynamic Behavior of Trinucleotide Repeat Tracts In Vivo

Measuring Dynamic Behavior of Trinucleotide Repeat Tracts In Vivo in Saccharomyces cerevisiae

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