Browse
Submit Data
Databases
API
Help

Dataset Information

28 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

A Major Role of DNA Polymerase ? in Replication of Both the Leading and Lagging DNA Strands.

ABSTRACT: Genetic studies with S. cerevisiae Pol? (pol3-L612M) and Pol? (pol2-M644G) mutant alleles, each of which display a higher rate for the generation of a specific mismatch, have led to the conclusion that Pol? is the primary leading strand replicase and that Pol? is restricted to replicating the lagging strand template. Contrary to this widely accepted view, here we show that Pol? plays a major role in the replication of both DNA strands, and that the paucity of pol3-L612M-generated errors on the leading strand results from their more proficient removal. Thus, the apparent lack of Pol? contribution to leading strand replication is due to differential mismatch removal rather than differential mismatch generation. Altogether, our genetic studies with Pol3 and Pol2 mutator alleles support the conclusion that Pol?, and not Pol?, is the major DNA polymerase for carrying out both leading and lagging DNA synthesis.

SUBMITTER: Johnson RE

PROVIDER: S-EPMC4517859 | biostudies-literature | 2015 Jul

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

A Major Role of DNA Polymerase δ in Replication of Both the Leading and Lagging DNA Strands.

Johnson Robert E RE Klassen Roland R Prakash Louise L Prakash Satya S

Molecular cell 20150702 2

Genetic studies with S. cerevisiae Polδ (pol3-L612M) and Polε (pol2-M644G) mutant alleles, each of which display a higher rate for the generation of a specific mismatch, have led to the conclusion that Polε is the primary leading strand replicase and that Polδ is restricted to replicating the lagging strand template. Contrary to this widely accepted view, here we show that Polδ plays a major role in the replication of both DNA strands, and that the paucity of pol3-L612M-generated errors on the l ...[more]

PMID: 26145172

Similar Datasets

DNA Polymerase Delta Synthesizes Both Strands during Break-Induced Replication.

Project description:Break-induced replication (BIR) is a pathway of homology-directed repair that repairs one-ended DNA breaks, such as those formed at broken replication forks or uncapped telomeres. In contrast to conventional S phase DNA synthesis, BIR proceeds by a migrating D-loop and results in conservative synthesis of the nascent strands. DNA polymerase delta (Pol δ) initiates BIR; however, it is not known whether synthesis of the invading strand switches to a different polymerase or how the complementary strand is synthesized. By using alleles of the replicative DNA polymerases that are permissive for ribonucleotide incorporation, thus generating a signature of their action in the genome that can be identified by hydrolytic end sequencing, we show that Pol δ replicates both the invading and the complementary strand during BIR. In support of this conclusion, we show that depletion of Pol δ from cells reduces BIR, whereas depletion of Pol ε has no effect.

| S-EPMC6862718 | biostudies-literature

DNA polymerase Δ synthesizes both strands during break-induced replication

Project description:We applied our ribonucleotide mapping technique HydEn-seq to study DNA polymerase usage during break-induced replication (BIR) in the budding yeast Saccharomyces cerevisiae.

2019-07-01 | GSE133558 | GEO

A perfect palindrome in the Escherichia coli chromosome forms DNA hairpins on both leading- and lagging-strands.

Project description:DNA palindromes are hotspots for DNA double strand breaks, inverted duplications and intra-chromosomal translocations in a wide spectrum of organisms from bacteria to humans. These reactions are mediated by DNA secondary structures such as hairpins and cruciforms. In order to further investigate the pathways of formation and cleavage of these structures, we have compared the processing of a 460 base pair (bp) perfect palindrome in the Escherichia coli chromosome with the same construct interrupted by a 20 bp spacer to form a 480 bp interrupted palindrome. We show here that the perfect palindrome can form hairpin DNA structures on the templates of the leading- and lagging-strands in a replication-dependent reaction. In the presence of the hairpin endonuclease SbcCD, both copies of the replicated chromosome containing the perfect palindrome are cleaved, resulting in the formation of an unrepairable DNA double-strand break and cell death. This contrasts with the interrupted palindrome, which forms a hairpin on the lagging-strand template that is processed to form breaks, which can be repaired by homologous recombination.

| S-EPMC4245961 | biostudies-literature

DNA polymerase Δ synthesizes both strands during break-induced replication

Project description:DNA polymerase Δ synthesizes both strands during break-induced replication

| PRJNA551884 | ENA

Roles for DNA polymerase δ in initiating and terminating leading strand DNA replication.

Project description:Most current evidence indicates that DNA polymerases ε and δ, respectively, perform the bulk of leading and lagging strand replication of the eukaryotic nuclear genome. Given that ribonucleotide and mismatch incorporation rates by these replicases influence somatic and germline patterns of variation, it is important to understand the details and exceptions to this overall division of labor. Using an improved method to map where these replicases incorporate ribonucleotides during replication, here we present evidence that DNA polymerase δ universally participates in initiating leading strand synthesis and that nascent leading strand synthesis switches from Pol ε to Pol δ during replication termination. Ribonucleotide maps from both the budding and fission yeast reveal conservation of these processes. These observations of replisome dynamics provide important insight into the mechanisms of eukaryotic replication and genome maintenance.

| S-EPMC6728351 | biostudies-literature

DNA Polymerase Delta Governs Parental Histone Transfer to DNA Replication Lagging Strand

Project description:Chromatin replication is intricately intertwined with DNA replication, and the recycling of parental histones is essential for epigenetic inheritance. The transfer of parental histones to both the DNA replication leading and lagging strands involves two distinct pathways: the leading strand utilizes DNA polymerase ε subunits Dpb3/Dpb4, while the lagging strand is facilitated by the MCM helicase subunit Mcm2. However, the process by which Mcm2, moving along the leading strand, facilitates the transfer of parental histones to the lagging strand remains unclear. Our study reveals that the deletion of Pol32, a non-essential subunit of major lagging-strand DNA polymerase δ, results in a predominant transfer of parental histone H3-H4 to the replication leading strand. Biochemical analyses further demonstrate that Pol32 can bind histone H3-H4 both in vivo and in vitro. The interaction of Pol32 with parental histone H3-H4 is disrupted by the mutation of Mcm2's histone H3-H4 binding domain. In conclusion, our findings identify the DNA polymerase delta subunit Pol32 as a novel key histone chaperone downstream of Mcm2, mediating the transfer of parental histones to the lagging strand during DNA replication.

2024-04-24 | GSE252049 | GEO

Mismatch repair balances leading and lagging strand DNA replication fidelity.

Project description:The two DNA strands of the nuclear genome are replicated asymmetrically using three DNA polymerases, ?, ?, and ?. Current evidence suggests that DNA polymerase ? (Pol ?) is the primary leading strand replicase, whereas Pols ? and ? primarily perform lagging strand replication. The fact that these polymerases differ in fidelity and error specificity is interesting in light of the fact that the stability of the nuclear genome depends in part on the ability of mismatch repair (MMR) to correct different mismatches generated in different contexts during replication. Here we provide the first comparison, to our knowledge, of the efficiency of MMR of leading and lagging strand replication errors. We first use the strand-biased ribonucleotide incorporation propensity of a Pol ? mutator variant to confirm that Pol ? is the primary leading strand replicase in Saccharomyces cerevisiae. We then use polymerase-specific error signatures to show that MMR efficiency in vivo strongly depends on the polymerase, the mismatch composition, and the location of the mismatch. An extreme case of variation by location is a T-T mismatch that is refractory to MMR. This mismatch is flanked by an AT-rich triplet repeat sequence that, when interrupted, restores MMR to > 95% efficiency. Thus this natural DNA sequence suppresses MMR, placing a nearby base pair at high risk of mutation due to leading strand replication infidelity. We find that, overall, MMR most efficiently corrects the most potentially deleterious errors (indels) and then the most common substitution mismatches. In combination with earlier studies, the results suggest that significant differences exist in the generation and repair of Pol ?, ?, and ? replication errors, but in a generally complementary manner that results in high-fidelity replication of both DNA strands of the yeast nuclear genome.

| S-EPMC3469411 | biostudies-literature

In vitro Assays for Eukaryotic Leading/Lagging Strand DNA Replication.

Project description:The eukaryotic replisome is a multiprotein complex that duplicates DNA. The replisome is sculpted to couple continuous leading strand synthesis with discontinuous lagging strand synthesis, primarily carried out by DNA polymerases ε and δ, respectively, along with helicases, polymerase α-primase, DNA sliding clamps, clamp loaders and many other proteins. We have previously established the mechanisms by which the polymerases ε and δ are targeted to their 'correct' strands, as well as quality control mechanisms that evict polymerases when they associate with an 'incorrect' strand. Here, we provide a practical guide to differentially assay leading and lagging strand replication in vitro using pure proteins.

| S-EPMC5659624 | biostudies-literature

Separable recruitment of DNA Polymerase α for leading- and lagging-strand replication initiation

Project description:Okazaki fragments were purified and sequenced from asynchronous S. cerevisiae at various DNA polymerase alpha levels in the presence or absence of interactions with Ctf4. The sequencing data was used to quantify efficiencies of replication origins.

2018-06-16 | GSE115897 | GEO

The proofreading exonuclease of leading-strand DNA polymerase epsilon prevents replication fork collapse at broken template strands.

Project description:Leading-strand DNA replication by polymerase epsilon (Polϵ) across single-strand breaks (SSBs) causes single-ended double-strand breaks (seDSBs), which are repaired via homology-directed repair (HDR) and suppressed by fork reversal (FR). Although previous studies identified many molecules required for hydroxyurea-induced FR, FR at seDSBs is poorly understood. Here, we identified molecules that specifically mediate FR at seDSBs. Because FR at seDSBs requires poly(ADP ribose)polymerase 1 (PARP1), we hypothesized that seDSB/FR-associated molecules would increase tolerance to camptothecin (CPT) but not the PARP inhibitor olaparib, even though both anti-cancer agents generate seDSBs. Indeed, we uncovered that Polϵ exonuclease and CTF18, a Polϵ cofactor, increased tolerance to CPT but not olaparib. To explore potential functional interactions between Polϵ exonuclease, CTF18, and PARP1, we created exonuclease-deficient POLE1exo-/-, CTF18-/-, PARP1-/-, CTF18-/-/POLE1exo-/-, PARP1-/-/POLE1exo-/-, and CTF18-/-/PARP1-/- cells. Epistasis analysis indicated that Polϵ exonuclease and CTF18 were interdependent and required PARP1 for CPT tolerance. Remarkably, POLE1exo-/- and HDR-deficient BRCA1-/- cells exhibited similar CPT sensitivity. Moreover, combining POLE1exo-/- with BRCA1-/- mutations synergistically increased CPT sensitivity. In conclusion, the newly identified PARP1-CTF18-Polϵ exonuclease axis and HDR act independently to prevent fork collapse at seDSBs. Olaparib inhibits this axis, explaining the pronounced cytotoxic effects of olaparib on HDR-deficient cells.

| S-EPMC10711444 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data