scholarly journals mlh3 separation of function and endonuclease defective mutants display an unexpected effect on meiotic recombination outcomes

2017 ◽  
Author(s):  
Najla Al-Sweel ◽  
Vandana Raghavan ◽  
Abhishek Dutta ◽  
V. P. Ajith ◽  
Luigi Di Vietro ◽  
...  
Keyword(s):  
Dna Replication ◽  
Mismatch Repair ◽  
Protein Interactions ◽  
Meiotic Recombination ◽  
Meiotic Division ◽  
Dna Mismatch Repair ◽  
Crossing Over ◽  
Baker's Yeast ◽  
Protein Protein Interactions ◽  
Dna Mismatch

AbstractMlh1-Mlh3 is an endonuclease hypothesized to act in meiosis to resolve double Holliday junctions into crossovers. It also plays a minor role in eukaryotic DNA mismatch repair (MMR). To understand how Mlh1-Mlh3 functions in both meiosis and MMR, we analyzed in baker’s yeast 60 new mlh3 alleles. Five alleles specifically disrupted MMR, whereas one (mlh3-32) specifically disrupted meiotic crossing over. Mlh1-mlh3 representatives for each separation of function class were purified and characterized. Both Mlh1-mlh3-32 (MMR+, crossover-) and Mlh1-mlh3-45 (MMR-, crossover+) displayed wild-type endonuclease activities in vitro. Msh2-Msh3, an MSH complex that acts with Mlh1-Mlh3 in MMR, stimulated the endonuclease activity of Mlh1-mlh3-32 but not Mlh1-mlh3-45, suggesting that Mlh1-mlh3-45 is defective in MSH interactions. Whole genome recombination maps were constructed for two mlh3 mutants with opposite separation of function phenotypes, and an endonuclease defective mutant. Unexpectedly, all three showed increases in the number of non-crossover events that were not observed in mlh3Δ. Our observations provide a structure-function map for Mlh3 that reveals the importance of protein-protein interactions in regulating Mlh1-Mlh3’s enzymatic activity. They also illustrate how defective meiotic components can alter the fate of meiotic recombination intermediates, providing new insights for how meiotic recombination pathways are regulated.Author SummaryDuring meiosis, diploid germ cells that become eggs or sperm undergo a single round of DNA replication followed by two consecutive chromosomal divisions. The segregation of chromosomes at the first meiotic division is dependent in most organisms on at least one genetic exchange, or crossover event, between chromosome homologs. Homologs that do not receive a crossover frequently undergo non-disjunction at the first meiotic division, yielding gametes that lack chromosomes or contain additional copies. Such events have been linked to human disease and infertility. Recent studies suggest that the Mlh1-Mlh3 complex is an endonuclease that resolves recombination intermediates into crossovers. Interestingly, this complex also acts as a matchmaker in DNA mismatch repair (MMR) to remove DNA replication errors. How does one complex act in two different processes? We investigated this question by performing a mutational analysis of the baker’s yeast Mlh3 protein. Five mutations were identified that disrupted MMR but not crossing over, and one mutation disrupted crossing over while maintaining MMR. Using a combination of biochemical and genetic analyses to further characterize these mutants we illustrate the importance of protein-protein interactions for Mlh1-Mlh3’s activity. Importantly, we illustrate how defective meiotic components can alter the outcome of meiotic recombination events. They also provide new insights in our understanding of the basis of infertility syndromes.

Protein-protein interactions in DNA mismatch repair

DNA Repair ◽  
2016 ◽  
Vol 38 ◽  
pp. 50-57 ◽  
Author(s):  
Peter Friedhoff ◽  
Pingping Li ◽  
Julia Gotthardt
Keyword(s):  
Mismatch Repair ◽  
Protein Interactions ◽  
Dna Mismatch Repair ◽  
Protein Protein Interactions ◽  
Dna Mismatch

scholarly journals Mispair-bound human MutS–MutL complex triggers DNA incisions and activates mismatch repair

Cell Research ◽  
2021 ◽  
Author(s):  
Janice Ortega ◽  
Grace Sanghee Lee ◽  
Liya Gu ◽  
Wei Yang ◽  
Guo-Min Li
Keyword(s):  
Mismatch Repair ◽  
Protein Interactions ◽  
Dna Mismatch Repair ◽  
Protein Protein Interactions ◽  
Dna Mismatch ◽  
Exonuclease 1 ◽  
Mismatch Recognition ◽  
Mispaired Bases

AbstractDNA mismatch repair (MMR) relies on MutS and MutL ATPases for mismatch recognition and strand-specific nuclease recruitment to remove mispaired bases in daughter strands. However, whether the MutS–MutL complex coordinates MMR by ATP-dependent sliding on DNA or protein–protein interactions between the mismatch and strand discrimination signal is ambiguous. Using functional MMR assays and systems preventing proteins from sliding, we show that sliding of human MutSα is required not for MMR initiation, but for final mismatch removal. MutSα recruits MutLα to form a mismatch-bound complex, which initiates MMR by nicking the daughter strand 5′ to the mismatch. Exonuclease 1 (Exo1) is then recruited to the nick and conducts 5′ → 3′ excision. ATP-dependent MutSα dissociation from the mismatch is necessary for Exo1 to remove the mispaired base when the excision reaches the mismatch. Therefore, our study has resolved a long-standing puzzle, and provided new insights into the mechanism of MMR initiation and mispair removal.

Involvement of mouse Mlh1 in DNA mismatch repair and meiotic crossing over

1996 ◽  
Vol 13 (3) ◽  
pp. 336-342 ◽  
Author(s):  
Sean M. Baker ◽  
Annemieke W. Plug ◽  
Tomas A. Prolla ◽  
C. Eric Bronner ◽  
Allie C. Harris ◽  
...  
Keyword(s):  
Mismatch Repair ◽  
Dna Mismatch Repair ◽  
Crossing Over ◽  
Dna Mismatch

scholarly journals Somatic Hypermutation in Muts Homologue (Msh)3-, Msh6-, and Msh3/Msh6-Deficient Mice Reveals a Role for the Msh2–Msh6 Heterodimer in Modulating the Base Substitution Pattern

2000 ◽  
Vol 191 (3) ◽  
pp. 579-584 ◽  
Author(s):  
Margrit Wiesendanger ◽  
Burkhard Kneitz ◽  
Winfried Edelmann ◽  
Matthew D. Scharff
Keyword(s):  
Immune Response ◽  
Dna Replication ◽  
Mismatch Repair ◽  
Dna Mismatch Repair ◽  
Somatic Hypermutation ◽  
Hot Spot ◽  
Base Substitution ◽  
Substitution Pattern ◽  
Dna Mismatch ◽  
Deficient Mice

Although the primary function of the DNA mismatch repair (MMR) system is to identify and correct base mismatches that have been erroneously introduced during DNA replication, recent studies have further implicated several MMR components in somatic hypermutation of immunoglobulin (Ig) genes. We studied the immune response in mice deficient in MutS homologue (MSH)3 and MSH6, two mutually exclusive partners of MSH2 that have not been examined previously for their role in Ig hypermutation. In Msh6−/− and Msh3−/−/Msh6−/− mice, base substitutions are preferentially targeted to G and C nucleotides and to an RGYW hot spot, as has been shown previously in Msh2−/− mice. In contrast, Msh3−/− mice show no differences from their littermate controls. These findings indicate that the MSH2–MSH6 heterodimer, but not the MSH2–MSH3 complex, is responsible for modulating Ig hypermutation.

scholarly journals DNA Mismatch Repair Catalyzed by Extracts of Mitotic, Postmitotic, and Senescent Drosophila Tissues and Involvement of mei-9 Gene Function for Full Activity

1998 ◽  
Vol 18 (3) ◽  
pp. 1436-1443 ◽  
Author(s):  
Arvinder Bhui-Kaur ◽  
Myron F. Goodman ◽  
John Tower
Keyword(s):  
Mismatch Repair ◽  
Dna Mismatch Repair ◽  
Specific Activity ◽  
Excision Repair ◽  
Crossing Over ◽  
Dna Mismatch ◽  
Nucleotide Excision ◽  
Repair Activity ◽  
Development And Aging

ABSTRACT Extracts of Drosophila embryos and adults have been found to catalyze highly efficient DNA mismatch repair, as well as repair of 1- and 5-bp loops. For mispairs T · G and G · G, repair is nick dependent and is specific for the nicked strand of heteroduplex DNA. In contrast, repair of A · A, C · A, G · A, C · T, T · T, and C · C is not nick dependent, suggesting the presence of glycosylase activities. For nick-dependent repair, the specific activity of embryo extracts was similar to that of extracts derived from the entirely postmitotic cells of young and senescent adults. Thus, DNA mismatch repair activity is expressed in Drosophila cells during both development and aging, suggesting that there may be a function or requirement for mismatch repair throughout the Drosophila life span. Nick-dependent repair was reduced in extracts of animals mutant for themei-9 gene. mei-9 has been shown to be required in vivo for certain types of DNA mismatch repair, nucleotide excision repair (NER), and meiotic crossing over and is theDrosophila homolog of the yeast NER gene rad1.

scholarly journals MutS/MutL crystal structure reveals that the MutS sliding clamp loads MutL onto DNA

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Flora S Groothuizen ◽  
Ines Winkler ◽  
Michele Cristóvão ◽  
Alexander Fish ◽  
Herrie HK Winterwerp ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Dna Replication ◽  
Mismatch Repair ◽  
Conformational Changes ◽  
Dna Mismatch Repair ◽  
Transient State ◽  
Dna Mismatch ◽  
Sliding Clamp ◽  
Downstream Effectors ◽  
A Site

To avoid mutations in the genome, DNA replication is generally followed by DNA mismatch repair (MMR). MMR starts when a MutS homolog recognizes a mismatch and undergoes an ATP-dependent transformation to an elusive sliding clamp state. How this transient state promotes MutL homolog recruitment and activation of repair is unclear. Here we present a crystal structure of the MutS/MutL complex using a site-specifically crosslinked complex and examine how large conformational changes lead to activation of MutL. The structure captures MutS in the sliding clamp conformation, where tilting of the MutS subunits across each other pushes DNA into a new channel, and reorientation of the connector domain creates an interface for MutL with both MutS subunits. Our work explains how the sliding clamp promotes loading of MutL onto DNA, to activate downstream effectors. We thus elucidate a crucial mechanism that ensures that MMR is initiated only after detection of a DNA mismatch.

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