scholarly journals CRISPR-Associated Primase-Polymerases are implicated in prokaryotic CRISPR-Cas adaptation

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Katerina Zabrady ◽  
Matej Zabrady ◽  
Peter Kolesar ◽  
Arthur W. H. Li ◽  
Aidan J. Doherty
Keyword(s):  
Dna Repair ◽  
Dna Synthesis ◽  
Genome Stability ◽  
Dna Polymerases ◽  
Acquired Immunity ◽  
Strand Displacement ◽  
Genetic Studies ◽  
Type Iiia ◽  
Maintain Genome Stability ◽  
Cas Proteins

AbstractCRISPR-Cas pathways provide prokaryotes with acquired “immunity” against foreign genetic elements, including phages and plasmids. Although many of the proteins associated with CRISPR-Cas mechanisms are characterized, some requisite enzymes remain elusive. Genetic studies have implicated host DNA polymerases in some CRISPR-Cas systems but CRISPR-specific replicases have not yet been discovered. We have identified and characterised a family of CRISPR-Associated Primase-Polymerases (CAPPs) in a range of prokaryotes that are operonically associated with Cas1 and Cas2. CAPPs belong to the Primase-Polymerase (Prim-Pol) superfamily of replicases that operate in various DNA repair and replication pathways that maintain genome stability. Here, we characterise the DNA synthesis activities of bacterial CAPP homologues from Type IIIA and IIIB CRISPR-Cas systems and establish that they possess a range of replicase activities including DNA priming, polymerisation and strand-displacement. We demonstrate that CAPPs operonically-associated partners, Cas1 and Cas2, form a complex that possesses spacer integration activity. We show that CAPPs physically associate with the Cas proteins to form bespoke CRISPR-Cas complexes. Finally, we propose how CAPPs activities, in conjunction with their partners, may function to undertake key roles in CRISPR-Cas adaptation.

scholarly journals Tim–Tipin dysfunction creates an indispensible reliance on the ATR–Chk1 pathway for continued DNA synthesis

2009 ◽  
Vol 187 (1) ◽  
pp. 15-23 ◽  
Author(s):  
Kevin D. Smith ◽  
Michael A. Fu ◽  
Eric J. Brown
Keyword(s):  
Dna Synthesis ◽  
Genome Stability ◽  
Double Strand Breaks ◽  
Dna Unwinding ◽  
Replication Forks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Nucleotide Incorporation ◽  
Pathway Activation ◽  
Maintain Genome Stability

The Tim (Timeless)–Tipin complex has been proposed to maintain genome stability by facilitating ATR-mediated Chk1 activation. However, as a replisome component, Tim–Tipin has also been suggested to couple DNA unwinding to synthesis, an activity expected to suppress single-stranded DNA (ssDNA) accumulation and limit ATR–Chk1 pathway engagement. We now demonstrate that Tim–Tipin depletion is sufficient to increase ssDNA accumulation at replication forks and stimulate ATR activity during otherwise unperturbed DNA replication. Notably, suppression of the ATR–Chk1 pathway in Tim–Tipin-deficient cells completely abrogates nucleotide incorporation in S phase, indicating that the ATR-dependent response to Tim–Tipin depletion is indispensible for continued DNA synthesis. Replication failure in ATR/Tim-deficient cells is strongly associated with synergistic increases in H2AX phosphorylation and DNA double-strand breaks, suggesting that ATR pathway activation preserves fork stability in instances of Tim–Tipin dysfunction. Together, these experiments indicate that the Tim–Tipin complex stabilizes replication forks both by preventing the accumulation of ssDNA upstream of ATR–Chk1 function and by facilitating phosphorylation of Chk1 by ATR.

scholarly journals Plant Organellar DNA Polymerases Evolved Multifunctionality through the Acquisition of Novel Amino Acid Insertions

Genes ◽  
2020 ◽  
Vol 11 (11) ◽  
pp. 1370
Author(s):  
Antolín Peralta-Castro ◽  
Paola L. García-Medel ◽  
Noe Baruch-Torres ◽  
Carlos H. Trasviña-Arenas ◽  
Víctor Juarez-Quintero ◽  
...  
Keyword(s):  
Dna Repair ◽  
Amino Acid ◽  
Dna Replication ◽  
Excision Repair ◽  
Dna Polymerases ◽  
Strand Displacement ◽  
End Joining ◽  
Apparent Contradiction ◽  
Organellar Dna ◽  
Base Excision

The majority of DNA polymerases (DNAPs) are specialized enzymes with specific roles in DNA replication, translesion DNA synthesis (TLS), or DNA repair. The enzymatic characteristics to perform accurate DNA replication are in apparent contradiction with TLS or DNA repair abilities. For instance, replicative DNAPs incorporate nucleotides with high fidelity and processivity, whereas TLS DNAPs are low-fidelity polymerases with distributive nucleotide incorporation. Plant organelles (mitochondria and chloroplast) are replicated by family-A DNA polymerases that are both replicative and TLS DNAPs. Furthermore, plant organellar DNA polymerases from the plant model Arabidopsis thaliana (AtPOLIs) execute repair of double-stranded breaks by microhomology-mediated end-joining and perform Base Excision Repair (BER) using lyase and strand-displacement activities. AtPOLIs harbor three unique insertions in their polymerization domain that are associated with TLS, microhomology-mediated end-joining (MMEJ), strand-displacement, and lyase activities. We postulate that AtPOLIs are able to execute those different functions through the acquisition of these novel amino acid insertions, making them multifunctional enzymes able to participate in DNA replication and DNA repair.

scholarly journals Overview of DNA Repair inTrypanosoma cruzi, Trypanosoma brucei,andLeishmania major

2010 ◽  
Vol 2010 ◽  
pp. 1-14 ◽  
Author(s):  
Danielle Gomes Passos-Silva ◽  
Matheus Andrade Rajão ◽  
Pedro Henrique Nascimento de Aguiar ◽  
João Pedro Vieira-da-Rocha ◽  
Carlos Renato Machado ◽  
...  
Keyword(s):  
Dna Repair ◽  
Trypanosoma Brucei ◽  
Genome Stability ◽  
Leishmania Major ◽  
Experimental Studies ◽  
Life Cycles ◽  
Dna Lesions ◽  
Cellular Mechanisms ◽  
Environmental Agents ◽  
Maintain Genome Stability

A wide variety of DNA lesions arise due to environmental agents, normal cellular metabolism, or intrinsic weaknesses in the chemical bonds of DNA. Diverse cellular mechanisms have evolved to maintain genome stability, including mechanisms to repair damaged DNA, to avoid the incorporation of modified nucleotides, and to tolerate lesions (translesion synthesis). Studies of the mechanisms related to DNA metabolism in trypanosomatids have been very limited. Together with recent experimental studies, the genome sequencing ofTrypanosoma brucei, Trypanosoma cruzi,andLeishmania major, three related pathogens with different life cycles and disease pathology, has revealed interesting features of the DNA repair mechanism in these protozoan parasites, which will be reviewed here.

scholarly journals Unravelling roles of error-prone DNA polymerases in shaping cancer genomes

Oncogene ◽  
2021 ◽  
Author(s):  
Cyrus Vaziri ◽  
Igor B. Rogozin ◽  
Qisheng Gu ◽  
Di Wu ◽  
Tovah A. Day
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Cancer Cells ◽  
Dna Polymerase ◽  
Genome Stability ◽  
Dna Polymerases ◽  
The Cancer Genome Atlas ◽  
Cancer Genomes ◽  
Recent Developments ◽  
Comprehensive Picture

AbstractMutagenesis is a key hallmark and enabling characteristic of cancer cells, yet the diverse underlying mutagenic mechanisms that shape cancer genomes are not understood. This review will consider the emerging challenge of determining how DNA damage response pathways—both tolerance and repair—act upon specific forms of DNA damage to generate mutations characteristic of tumors. DNA polymerases are typically the ultimate mutagenic effectors of DNA repair pathways. Therefore, understanding the contributions of DNA polymerases is critical to develop a more comprehensive picture of mutagenic mechanisms in tumors. Selection of an appropriate DNA polymerase—whether error-free or error-prone—for a particular DNA template is critical to the maintenance of genome stability. We review different modes of DNA polymerase dysregulation including mutation, polymorphism, and over-expression of the polymerases themselves or their associated activators. Based upon recent findings connecting DNA polymerases with specific mechanisms of mutagenesis, we propose that compensation for DNA repair defects by error-prone polymerases may be a general paradigm molding the mutational landscape of cancer cells. Notably, we demonstrate that correlation of error-prone polymerase expression with mutation burden in a subset of patient tumors from The Cancer Genome Atlas can identify mechanistic hypotheses for further testing. We contrast experimental approaches from broad, genome-wide strategies to approaches with a narrower focus on a few hundred base pairs of DNA. In addition, we consider recent developments in computational annotation of patient tumor data to identify patterns of mutagenesis. Finally, we discuss the innovations and future experiments that will develop a more comprehensive portrait of mutagenic mechanisms in human tumors.

scholarly journals DNA Replication Through Strand Displacement During Lagging Strand DNA Synthesis in Saccharomyces cerevisiae

Genes ◽  
2019 ◽  
Vol 10 (2) ◽  
pp. 167 ◽  
Author(s):  
Michele Giannattasio ◽  
Dana Branzei
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Replication ◽  
Dna Synthesis ◽  
Dna Polymerases ◽  
Experimental Results ◽  
Genome Integrity ◽  
Strand Displacement ◽  
Viral Dna ◽  
Okazaki Fragment Processing ◽  
Lagging Strand

This review discusses a set of experimental results that support the existence of extended strand displacement events during budding yeast lagging strand DNA synthesis. Starting from introducing the mechanisms and factors involved in leading and lagging strand DNA synthesis and some aspects of the architecture of the eukaryotic replisome, we discuss studies on bacterial, bacteriophage and viral DNA polymerases with potent strand displacement activities. We describe proposed pathways of Okazaki fragment processing via short and long flaps, with a focus on experimental results obtained in Saccharomyces cerevisiae that suggest the existence of frequent and extended strand displacement events during eukaryotic lagging strand DNA synthesis, and comment on their implications for genome integrity.

scholarly journals Polynucleotide kinase–phosphatase enables neurogenesis via multiple DNA repair pathways to maintain genome stability

2015 ◽  
Vol 34 (19) ◽  
pp. 2465-2480 ◽  
Author(s):  
Mikio Shimada ◽  
Lavinia C Dumitrache ◽  
Helen R Russell ◽  
Peter J McKinnon
Keyword(s):  
Dna Repair ◽  
Genome Stability ◽  
Polynucleotide Kinase ◽  
Repair Pathways ◽  
Maintain Genome Stability

scholarly journals Impact of G-Quadruplexes and Chronic Inflammation on Genome Instability: Additive Effects during Carcinogenesis

Genes ◽  
2021 ◽  
Vol 12 (11) ◽  
pp. 1779
Author(s):  
MaryElizabeth Stein ◽  
Kristin A. Eckert
Keyword(s):  
Chronic Inflammation ◽  
Genome Stability ◽  
Large Scale ◽  
Oxidative Dna Damage ◽  
Genome Instability ◽  
Physiological State ◽  
Dna Polymerases ◽  
Small Scale ◽  
Gene Promoters ◽  
Maintain Genome Stability

Genome instability is an enabling characteristic of cancer, essential for cancer cell evolution. Hotspots of genome instability, from small-scale point mutations to large-scale structural variants, are associated with sequences that potentially form non-B DNA structures. G-quadruplex (G4) forming motifs are enriched at structural variant endpoints in cancer genomes. Chronic inflammation is a physiological state underlying cancer development, and oxidative DNA damage is commonly invoked to explain how inflammation promotes genome instability. We summarize where G4s and oxidative stress overlap, with a focus on DNA replication. Guanine has low ionization potential, making G4s vulnerable to oxidative damage. Impacts to G4 structure are dependent upon lesion type, location, and G4 conformation. Occasionally, G4s pose a challenge to replicative DNA polymerases, requiring specialized DNA polymerases to maintain genome stability. Therefore, chronic inflammation creates a dual challenge for DNA polymerases to maintain genome stability: faithful G4 synthesis and bypassing unrepaired oxidative lesions. Inflammation is also accompanied by global transcriptome changes that may impact mutagenesis. Several studies suggest a regulatory role for G4s within cancer- and inflammatory-related gene promoters. We discuss the extent to which inflammation could influence gene regulation by G4s, thereby impacting genome instability, and highlight key areas for new investigation.

scholarly journals When DNA Polymerases Multitask: Functions Beyond Nucleotidyl Transfer

2022 ◽  
Vol 8 ◽  
Author(s):  
Denisse Carvajal-Maldonado ◽  
Lea Drogalis Beckham ◽  
Richard D. Wood ◽  
Sylvie Doublié
Keyword(s):  
Dna Repair ◽  
Dna Replication ◽  
Dna Synthesis ◽  
Extracurricular Activities ◽  
Damage Tolerance ◽  
Dna Polymerases ◽  
Translesion Dna Synthesis ◽  
Dna Strands ◽  
Translesion Dna ◽  
Central Reaction

DNA polymerases catalyze nucleotidyl transfer, the central reaction in synthesis of DNA polynucleotide chains. They function not only in DNA replication, but also in diverse aspects of DNA repair and recombination. Some DNA polymerases can perform translesion DNA synthesis, facilitating damage tolerance and leading to mutagenesis. In addition to these functions, many DNA polymerases conduct biochemically distinct reactions. This review presents examples of DNA polymerases that carry out nuclease (3ʹ—5′ exonuclease, 5′ nuclease, or end-trimming nuclease) or lyase (5′ dRP lyase) extracurricular activities. The discussion underscores how DNA polymerases have a remarkable ability to manipulate DNA strands, sometimes involving relatively large intramolecular movement.

scholarly journals Acquisition of meiotic DNA repair regulators maintain genome stability in glioblastoma

2015 ◽  
Vol 6 (4) ◽  
pp. e1732-e1732 ◽  
Author(s):  
M Rivera ◽  
Q Wu ◽  
P Hamerlik ◽  
A B Hjelmeland ◽  
S Bao ◽  
...  
Keyword(s):  
Dna Repair ◽  
Genome Stability ◽  
Maintain Genome Stability

scholarly journals Bacteriophage-Encoded DNA Polymerases—Beyond the Traditional View of Polymerase Activities

2022 ◽  
Vol 23 (2) ◽  
pp. 635
Author(s):  
Joanna Morcinek-Orłowska ◽  
Karolina Zdrojewska ◽  
Alicja Węgrzyn
Keyword(s):  
Dna Repair ◽  
Dna Replication ◽  
Genetic Engineering ◽  
Dna Synthesis ◽  
Genetic Recombination ◽  
Dna Polymerases ◽  
Traditional View ◽  
Even Start ◽  
Viral Dna ◽  
Viral Dna Replication

DNA polymerases are enzymes capable of synthesizing DNA. They are involved in replication of genomes of all cellular organisms as well as in processes of DNA repair and genetic recombination. However, DNA polymerases can also be encoded by viruses, including bacteriophages, and such enzymes are involved in viral DNA replication. DNA synthesizing enzymes are grouped in several families according to their structures and functions. Nevertheless, there are examples of bacteriophage-encoded DNA polymerases which are significantly different from other known enzymes capable of catalyzing synthesis of DNA. These differences are both structural and functional, indicating a huge biodiversity of bacteriophages and specific properties of their enzymes which had to evolve under certain conditions, selecting unusual properties of the enzymes which are nonetheless crucial for survival of these viruses, propagating as special kinds of obligatory parasites. In this review, we present a brief overview on DNA polymerases, and then we discuss unusual properties of different bacteriophage-encoded enzymes, such as those able to initiate DNA synthesis using the protein-priming mechanisms or even start this process without any primer, as well as able to incorporate untypical nucleotides. Apart from being extremely interesting examples of biochemical biodiversity, bacteriophage-encoded DNA polymerases can also be useful tools in genetic engineering and biotechnology.

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