Mapping the Polarity of Changes That Occur in Interrupted CAG Repeat Tracts in Yeast

Maurer, Debra J.; O’Callaghan, Brennon; Livingston, Dennis

doi:10.1128/mcb.18.8.4597

Cited by 25 publications

(17 citation statements)

References 32 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Contractions were markedly higher than expansions and increased with increasing tract length (8.8% for CTG-85; 23% for CTG-155) ( Table 2). This result is consistent with the known bias toward contractions in yeast cells (16,38,39). As has been seen previously, repeat sequence instability was dramatically increased in the rad27⌬ strain with a strong bias towards expansion (Table 2) (8,15,52,56).…”

Section: Mutation Of Fen1 Endonuclease Leads To Ctg Tract Expansions supporting

confidence: 79%

Saccharomyces cerevisiae Flap Endonuclease 1 Uses Flap Equilibration To Maintain Triplet Repeat Stability

Liu

Zhang²,

Veeraraghavan

et al. 2004

Molecular and Cellular Biology

View full text Add to dashboard Cite

Flap endonuclease 1 (FEN1) is a central component of Okazaki fragment maturation in eukaryotes.Genetic analysis of Saccharomyces cerevisiae FEN1 (RAD27) also reveals its important role in preventing trinucleotide repeat (TNR) expansion. In humans such expansion is associated with neurodegenerative diseases. In vitro, FEN1 can inhibit TNR expansion by employing its endonuclease activity to compete with DNA ligase I. Here we employed two yeast FEN1 nuclease mutants, rad27-G67S and rad27-G240D, to further define the mechanism by which FEN1 prevents TNR expansion. Using a yeast artificial chromosome system that can detect both TNR instability and fragility, we demonstrate that the G240D but not the G67S mutation increases both the expansion and fragility of a CTG tract in vivo. In vitro, the G240D nuclease is proficient in cleaving a fixed nonrepeat double flap; however, it exhibits severely impaired cleavage of both nonrepeat and CTG-containing equilibrating flaps. In contrast, wild-type FEN1 and the G67S mutant exhibit more efficient cleavage on an equilibrating flap than on a fixed CTG flap. The degree of TNR expansion and the amount of chromosome fragility observed in the mutant strains correlate with the severity of defective flap cleavage in vitro. We present a model to explain how flap equilibration and the unique tracking mechanism of FEN1 can collaborate to remove TNR flaps and prevent repeat expansion.Eukaryotic flap endonuclease 1 (FEN1) is critical for removing initiator RNA primers from Okazaki fragments during DNA lagging-strand synthesis (20,50,60). The enzyme belongs to a highly conserved 5Ј endo-exonuclease superfamily (53). FEN1 is a structure-specific nuclease in that it specifically removes the 5Ј unannealed flap in a branch structure resulting from strand displacement synthesis. This property allows the enzyme to actively participate in both Okazaki fragment processing and DNA long-patch base excision repair (12, 37) since both of these processes involve creation of a 5Ј flap by strand displacement synthesis. The removal of initiator RNA primers in Okazaki fragments has been proposed as a two-step mechanism in which an endonuclease, Dna2 protein, removes a part of the flap (2). The remainder of the flap is then cut off by FEN1, leaving a nick to be ligated. Other studies suggest that FEN1 alone is sufficient for removal of most flaps (1). Studies in vivo demonstrate a critical role for FEN1 since the absence of enzyme activity in mice leads to embryonic lethality (30). Knocking out one copy of FEN1 along with one of the adenomatous polyposis coli genes in mice results in an increased number of adenocarcinomas, presumably due to the partial loss of FEN1 function in mammalian DNA replication and repair (30).In Saccharomyces cerevisiae the FEN1 homologue, RAD27/ RTH1 (47) demonstrates a high homology (79% conserved and 60% identical) with human FEN1 (53). A deletion of RAD27/ RTH1 leads to conditional lethality phenotypes (54). The mutant cells exhibit temperature sensitivity and cell cycle arrest ...

show abstract

Section: Mutation Of Fen1 Endonuclease Leads To Ctg Tract Expansions supporting

confidence: 79%

Saccharomyces cerevisiae Flap Endonuclease 1 Uses Flap Equilibration To Maintain Triplet Repeat Stability

Liu

Zhang²,

Veeraraghavan

et al. 2004

Molecular and Cellular Biology

View full text Add to dashboard Cite

show abstract

“…In another yeast study, Maurer et al (15) used tracts of 90 to 97 CAG repeats containing a single CAT interruption to investigate the polarity of repeat mutations. They observed only contractions in unselected populations of wild-type cells.…”

Section: Discussionmentioning

confidence: 99%

Stabilizing Effects of Interruptions on Trinucleotide Repeat Expansions in Saccharomyces cerevisiae

Rolfsmeier

Lahue

2000

Molecular and Cellular Biology

View full text Add to dashboard Cite

In most trinucleotide repeat (TNR) diseases, the primary factor determining the likelihood of expansions is the length of the TNR. In some diseases, however, stable alleles contain one to three base pair substitutions that interrupt the TNR tract. The unexpected stability of these alleles compared to the frequent expansions of perfect TNRs suggested that interruptions somehow block expansions and that expansions occur only upon loss of at least one interruption. The work in this study uses a yeast genetic assay to examine the mechanism of stabilization conferred by two interruptions of a 25-repeat tract. Expansion rates are reduced up to 90-fold compared to an uninterrupted allele. Stabilization is greatest when the interruption is replicated early on the lagging strand, relative to the rest of the TNR. Although expansions are infrequent, they are often polar, gaining new DNA within the largest available stretch of perfect repeats. Surprisingly, interruptions are always retained and sometimes even duplicated, suggesting that expansion in yeast cells can proceed without loss of the interruption. These findings support a stabilization model in which interruptions contribute in cis to reduce hairpin formation during TNR replication and thus inhibit expansion rates.Trinucleotide repeat (TNR) instability has been found in more than 12 human neurological diseases that have many common genetic traits (1,7,18). One of the most important predictors of TNR expansions is the length of the triplet tract. In normal populations, the TNR is highly polymorphic but relatively short. In affected individuals the TNR is expanded anywhere from 5 to 2,000 repeats, depending on the disease locus (1, 7, 18). Not only does expansion often lead to disease but longer repeats are also further destabilized, expanding with even higher frequency upon subsequent transmissions. The cutoff between short, stable alleles and long, unstable alleles has been termed the threshold (1,7,18). Once a threshold of about 35 repeats is reached, the likelihood of expansion in the next generation is greatly increased.The purity of the TNR tract also influences its mutability. There are three examples of TNR diseases in which most normal, stable alleles contain one to three point mutations, or interruptions, interspersed within the perfect repeat tract. SCA1 (spinocerebellar ataxia type 1 gene) contains CAT interruptions in a CAG tract (2), the CGG tract of the fragile X syndrome gene FMR1 is punctuated with AGGs (3, 9, 13, 25), and CAAs are dispersed through the CAG tract of SCA2 (10, 21, 23). About 1% of normal Friedreich's ataxia alleles have five to eight GAGGAA hexanucleotide interruptions within a GAA tract (17). In contrast, expanded alleles of these genes have fewer or no interruptions. This correlation has led to the suggestion that one or more interruptions must be lost before expansion can occur (2,3,9,10,13,22,25). The most direct test of this hypothesis would be to look at expansions that arise directly from interrupted alleles. However, it has no...

show abstract

“…Studies in budding yeast have supported a number of different models for the expansion and contraction of TNR tracts including classical ''slippage'' and recombination (Maurer et al 1996(Maurer et al , 1998Freudenreich et al 1997Freudenreich et al , 1998Miret et al 1997Miret et al , 1998Cohen et al 1999;Richard et al 1999Richard et al , 2000Richard et al , 2003Schweitzer and Livingston 1999;Ireland et al 2000;Jankowski et al 2000;Schweitzer et al 2001;Jankowski and Nag 2002;Callahan et al 2003;Cleary and Pearson 2003;Lenzmeier and Freudenreich 2003). In all models, the ability of TNR sequences to produce intermediates stabilized by secondary structure formation is thought to be critical to the mutation process (reviewed in Mitas 1997;Cleary and Pearson 2003;Lenzmeier and Freudenreich 2003).…”

Section: E Xpansion Of Trinucleotide Repeat (Tnr) Tractsmentioning

confidence: 99%

Genetic Instability Induced by Overexpression of DNA Ligase I in Budding Yeast

et al. 2005

View full text Add to dashboard Cite

Recombination and microsatellite mutation in humans contribute to disorders including cancer and trinucleotide repeat (TNR) disease. TNR expansions in wild-type yeast may arise by flap ligation during lagging-strand replication. Here we show that overexpression of DNA ligase I (CDC9) increases the rates of TNR expansion, of TNR contraction, and of mitotic recombination. Surprisingly, this effect is observed with catalytically inactive forms of Cdc9p protein, but only if they possess a functional PCNA-binding site. Furthermore, in vitro analysis indicates that the interaction of PCNA with Cdc9p and Rad27p (Fen1) is mutually exclusive. Together our genetic and biochemical analysis suggests that, although DNA ligase I seals DNA nicks during replication, repair, and recombination, higher than normal levels can yield genetic instability by disrupting the normal interplay of PCNA with other proteins such as Fen1.

show abstract

Mapping the Polarity of Changes That Occur in Interrupted CAG Repeat Tracts in Yeast

Cited by 25 publications

References 32 publications

Saccharomyces cerevisiae Flap Endonuclease 1 Uses Flap Equilibration To Maintain Triplet Repeat Stability

Saccharomyces cerevisiae Flap Endonuclease 1 Uses Flap Equilibration To Maintain Triplet Repeat Stability

Stabilizing Effects of Interruptions on Trinucleotide Repeat Expansions in Saccharomyces cerevisiae

Genetic Instability Induced by Overexpression of DNA Ligase I in Budding Yeast

Contact Info

Product

Resources

About