New insights into how MutS separates its function in the regulation of the Pol IV access to replication sites from that in the conserved MMR pathway (original) (raw)
Related papers
New insights into the mechanism of DNA mismatch repair
Chromosoma, 2015
The genome of all organisms is constantly being challenged by endogenous and exogenous sources of DNA damage. Errors like base:base mismatches or small insertions and deletions, primarily introduced by DNA polymerases during DNA replication are repaired by an evolutionary conserved DNA mismatch repair (MMR) system. The MMR system, together with the DNA replication machinery, promote repair by an excision and resynthesis mechanism during or after DNA replication, increasing replication fidelity by up-to-three orders of magnitude. Consequently, inactivation of MMR genes results in elevated mutation rates that can lead to increased cancer susceptibility in humans. In this review, we summarize our current understanding of MMR with a focus on the different MMR protein complexes, their function and structure. We also discuss how recent findings have provided new insights in the spatio-temporal regulation and mechanism of MMR.
Nucleic acids research, 2016
Translesion DNA polymerases (Pol) function in the bypass of template lesions to relieve stalled replication forks but also display potentially deleterious mutagenic phenotypes that contribute to antibiotic resistance in bacteria and lead to human disease. Effective activity of these enzymes requires association with ring-shaped processivity factors, which dictate their access to sites of DNA synthesis. Here, we show for the first time that the mismatch repair protein MutS plays a role in regulating access of the conserved Y-family Pol IV to replication sites. Our biochemical data reveals that MutS inhibits the interaction of Pol IV with the β clamp processivity factor by competing for binding to the ring. Moreover, the MutS-β clamp association is critical for controlling Pol IV mutagenic replication under normal growth conditions. Thus, our findings reveal important insights into a non-canonical function of MutS in the regulation of a replication activity.
Stoichiometry of MutS and MutL at unrepaired mismatches in vivo suggests a mechanism of repair
Nucleic Acids Research, 2012
Mismatch repair (MMR) is an evolutionarily conserved DNA repair system, which corrects mismatched bases arising during DNA replication. MutS recognizes and binds base pair mismatches, while the MutL protein interacts with MutSmismatch complex and triggers MutH endonuclease activity at a distal-strand discrimination site on the DNA. The mechanism of communication between these two distal sites on the DNA is not known. We used functional fluorescent MMR proteins, MutS and MutL, in order to investigate the formation of the fluorescent MMR protein complexes on mismatches in real-time in growing Escherichia coli cells. We found that MutS and MutL proteins co-localize on unrepaired mismatches to form fluorescent foci. MutL foci were, on average, 2.7 times more intense than the MutS foci co-localized on individual mismatches. A steric block on the DNA provided by the MutHE56A mutant protein, which binds to but does not cut the DNA at the strand discrimination site, decreased MutL foci fluorescence 3-fold. This indicates that MutL accumulates from the mismatch site toward strand discrimination site along the DNA. Our results corroborate the hypothesis postulating that MutL accumulation assures the coordination of the MMR activities between the mismatch and the strand discrimination site.
Roles for mismatch repair factors in regulating genetic recombination
Molecular and cellular biology, 2000
Mismatch repair (MMR) systems are evolutionarily conserved and play a primary role in mutation avoidance by removing base-base and small insertion-deletion mismatches that arise during DNA replication (31). In addition, MMR factors are required for the repair of mismatches in heteroduplex DNA (hDNA) that form as a result of sequence heterologies between recombining sequences . MMR also acts to inhibit recombination between moderately divergent (homeologous) sequences . The roles of MMR during recombination are believed to reflect the interaction of MMR factors with mismatches that arise in hDNA or possibly with other structures such as Holliday junctions (2, 33). The full range of effects that MMR can exert on mitotic and meiotic recombination have been discussed elsewhere (11) and will only be summarized briefly here. The purpose of this review is to highlight recent results that have furthered our understanding of interactions between MMR factors and mitotic recombination intermediates.
Mismatch repair and DNA damage signalling
DNA Repair, 2004
Postreplicative mismatch repair (MMR) increases the fidelity of DNA replication by up to three orders of magnitude, through correcting DNA polymerase errors that escaped proofreading. MMR also controls homologous recombination (HR) by aborting strand exchange between divergent DNA sequences. In recent years, MMR has also been implicated in the response of mammalian cells to DNA damaging agents. Thus, MMR-deficient cells were shown to be around 100-fold more resistant to killing by methylating agents of the S N 1type than cells with functional MMR. In the case of cisplatin, the sensitivity difference was lower, typically two-to three-fold, but was observed in all matched MMR-proficient and -deficient cell pairs. More controversial is the role of MMR in cellular response to other DNA damaging agents, such as ionizing radiation (IR), topoisomerase poisons, antimetabolites, UV radiation and DNA intercalators. The MMR-dependent DNA damage signalling pathways activated by the above agents are also ill-defined. To date, signalling cascades involving the Ataxia telangiectasia mutated (ATM), ATM-and Rad3-related (ATR), as well as the stress-activated kinases JNK/SAPK and p38␣ have been linked with methylating agent and 6-thioguanine (TG) treatments, while cisplatin damage was reported to activate the c-Abl and JNK/SAPK kinases in MMR-dependent manner. MMR defects are found in several different cancer types, both familiar and sporadic, and it is possible that the involvement of the MMR system in DNA damage signalling play an important role in transformation. The scope of this article is to provide a brief overview of the recent literature on this subject and to raise questions that could be addressed in future studies.
MutSΒ exceeds MutSα in dinucleotide loop repair
British Journal of Cancer, 2010
The target substrates of DNA mismatch recognising factors MutSalpha (MSH2+MSH6) and MutSbeta (MSH2+MSH3) have already been widely researched. However, the extent of their functional redundancy and clinical substance remains unclear. Mismatch repair (MMR)-deficient tumours are strongly associated with microsatellite instability (MSI) and the degree and type of MSI seem to be dependent on the MMR gene affected, and is linked to its substrate specificities. Deficiency in MSH2 and MSH6 is associated with both mononucleotide and dinucleotide repeat instability. Although no pathogenic MSH3 mutations have been reported, its deficiency is also suggested to cause low dinucleotide repeat instability. To assess the substrate specificities and functionality of MutSalpha and MutSbeta we performed an in vitro MMR assay using three substrate constructs, GT mismatch, 1 and 2 nucleotide insertion/deletion loops (IDLs) in three different cell lines. Our results show that though MutSalpha alone seems to be responsible for GT and IDL1 repair, MutSalpha and MutSbeta indeed have functional redundancy in IDL2 repair and in contrast with earlier studies, MutSbeta seems to exceed MutSalpha. The finding is clinically relevant because the strong role of MutSbeta in IDL2 repair indicates MSH3 deficiency in tumours with low dinucleotide and no mononucleotide repeat instability.
Structure, 2000
The Scripps Research Institute been established in E. coli. MMR is initiated when MutS recognizes and binds to mis-La Jolla, California † Cell and Molecular Biology matched DNA. Prokaryotic MutS forms homodimers that can recognize a variety of DNA substrates, including mispairs and Life Sciences Division Lawrence Berkeley National Laboratory small insertion/deletion loops. Eukaryotic MutS homologs (MSHs) form different heterodimers that can recognize different Berkeley, California DNA substrates, including not only mispairs and loops, but also single-strand flaps, Holliday junctions and various chemical lesions [3, 11]. MSHs contain nucleotide binding Walker A and Summary B motifs and extensive research was aimed at determining the role of ATP in MutS function and MMR. A defining observation DNA mismatch repair (MMR) is initiated when the MutS prowas that MSH2/6 binds to damaged DNA in the ADP-bound tein recognizes damaged DNA. Crystal structures of MutS state and that binding to mismatched DNA stimulates ATP bound to mispaired and unpaired DNA show how MutS distinbinding and hydrolysis [12]. Furthermore, ATP binding induces guishes damaged from undamaged DNA and explain how a a conformational change in MSH2/6, which can diffuse along broad variety of DNA mismatch lesions can be detected. the DNA [13] and was suggested to form a sliding clamp and The structures suggest mechanisms for the ATP-induced to promote ␣-loop formation [14, 15]. However, the role of ATP structural regulation of multistep DNA repair processes.
Unexpected moves: a conformational change in MutSα enables high-affinity DNA mismatch binding
Nucleic Acids Research
The DNA mismatch repair protein MutSα recognizes wrongly incorporated DNA bases and initiates their correction during DNA replication. Dysfunctions in mismatch repair lead to a predisposition to cancer. Here, we study the homozygous mutation V63E in MSH2 that was found in the germline of a patient with suspected constitutional mismatch repair deficiency syndrome who developed colorectal cancer before the age of 30. Characterization of the mutant in mouse models, as well as slippage and repair assays, shows a mildly pathogenic phenotype. Using cryogenic electron microscopy and surface plasmon resonance, we explored the mechanistic effect of this mutation on MutSα function. We discovered that V63E disrupts a previously unappreciated interface between the mismatch binding domains (MBDs) of MSH2 and MSH6 and leads to reduced DNA binding. Our research identifies this interface as a ‘safety lock’ that ensures high-affinity DNA binding to increase replication fidelity. Our mechanistic mode...