scholarly journals Mammalian DNA Polymerase Kappa Activity and Specificity

Molecules ◽  
2019 ◽  
Vol 24 (15) ◽  
pp. 2805 ◽  
Author(s):  
Hannah R. Stern ◽  
Jana Sefcikova ◽  
Victoria E. Chaparro ◽  
Penny J. Beuning
Keyword(s):  
Dna Polymerase ◽  
Genome Instability ◽  
Fragile Sites ◽  
Chemotherapy Response ◽  
Nucleotide Polymorphisms ◽  
Lesion Bypass ◽  
Base Pairs ◽  
Domains Of Life ◽  
Copy Dna ◽  
Y Family

DNA polymerase (pol) kappa is a Y-family translesion DNA polymerase conserved throughout all domains of life. Pol kappa is special6 ized for the ability to copy DNA containing minor groove DNA adducts, especially N2-dG adducts, as well as to extend primer termini containing DNA damage or mismatched base pairs. Pol kappa generally cannot copy DNA containing major groove modifications or UV-induced photoproducts. Pol kappa can also copy structured or non-B-form DNA, such as microsatellite DNA, common fragile sites, and DNA containing G quadruplexes. Thus, pol kappa has roles both in maintaining and compromising genomic integrity. The expression of pol kappa is altered in several different cancer types, which can lead to genome instability. In addition, many cancer-associated single-nucleotide polymorphisms have been reported in the POLK gene, some of which are associated with poor survival and altered chemotherapy response. Because of this, identifying inhibitors of pol kappa is an active area of research. This review will address these activities of pol kappa, with a focus on lesion bypass and cellular mutagenesis.

scholarly journals Mechanism of double-base lesion bypass catalyzed by a Y-family DNA polymerase

2008 ◽  
Vol 36 (12) ◽  
pp. 3867-3878 ◽  
Author(s):  
Jessica A. Brown ◽  
Sean A. Newmister ◽  
Kevin A. Fiala ◽  
Zucai Suo
Keyword(s):  
Dna Polymerase ◽  
Lesion Bypass ◽  
Double Base ◽  
Base Lesion ◽  
Y Family

scholarly journals Multisite SUMOylation restrains DNA polymerase η interactions with DNA damage sites

2020 ◽  
Vol 295 (25) ◽  
pp. 8350-8362 ◽  
Author(s):  
Claire Guérillon ◽  
Stine Smedegaard ◽  
Ivo A. Hendriks ◽  
Michael L. Nielsen ◽  
Niels Mailand
Keyword(s):  
Dna Damage ◽  
Dna Polymerase ◽  
Feedback Inhibition ◽  
Dna Lesions ◽  
Nuclear Antigen ◽  
Inhibition Mechanism ◽  
Proteomic Profiling ◽  
Lesion Bypass ◽  
Y Family ◽  
Damaged Dna

Translesion DNA synthesis (TLS) mediated by low-fidelity DNA polymerases is an essential cellular mechanism for bypassing DNA lesions that obstruct DNA replication progression. However, the access of TLS polymerases to the replication machinery must be kept tightly in check to avoid excessive mutagenesis. Recruitment of DNA polymerase η (Pol η) and other Y-family TLS polymerases to damaged DNA relies on proliferating cell nuclear antigen (PCNA) monoubiquitylation and is regulated at several levels. Using a microscopy-based RNAi screen, here we identified an important role of the SUMO modification pathway in limiting Pol η interactions with DNA damage sites in human cells. We found that Pol η undergoes DNA damage- and protein inhibitor of activated STAT 1 (PIAS1)-dependent polySUMOylation upon its association with monoubiquitylated PCNA, rendering it susceptible to extraction from DNA damage sites by SUMO-targeted ubiquitin ligase (STUbL) activity. Using proteomic profiling, we demonstrate that Pol η is targeted for multisite SUMOylation, and that collectively these SUMO modifications are essential for PIAS1- and STUbL-mediated displacement of Pol η from DNA damage sites. These findings suggest that a SUMO-driven feedback inhibition mechanism is an intrinsic feature of TLS-mediated lesion bypass functioning to curtail the interaction of Pol η with PCNA at damaged DNA to prevent harmful mutagenesis.

scholarly journals Escherichia coli DNA Polymerase III Can Replicate Efficiently past a T-T cis-syn Cyclobutane Dimer if DNA Polymerase V and the 3′ to 5′ Exonuclease Proofreading Function Encoded by dnaQ Are Inactivated

2002 ◽  
Vol 184 (10) ◽  
pp. 2674-2681 ◽  
Author(s):  
Angela Borden ◽  
Paul I. O'Grady ◽  
Dominique Vandewiele ◽  
Antonio R. Fernández de Henestrosa ◽  
Christopher W. Lawrence ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dna Polymerase ◽  
Wild Type ◽  
Lesion Bypass ◽  
Base Pairs ◽  
Pol V ◽  
Pol I ◽  
Cyclobutane Dimer ◽  
Dna Polymerase V ◽  
Pol Iii

ABSTRACT Although very little replication past a T-T cis-syn cyclobutane dimer normally takes place in Escherichia coli in the absence of DNA polymerase V (Pol V), we previously observed as much as half of the wild-type bypass frequency in Pol V-deficient (ΔumuDC) strains if the 3′ to 5′ exonuclease proofreading activity of the Pol III ε subunit was also disabled by mutD5. This observation might be explained in at least two ways. In the absence of Pol V, wild-type Pol III might bind preferentially to the blocked primer terminus but be incapable of bypass, whereas the proofreading-deficient enzyme might dissociate more readily, providing access to bypass polymerases. Alternatively, even though wild-type Pol III is generally regarded as being incapable of lesion bypass, proofreading-impaired Pol III might itself perform this function. We have investigated this issue by examining dimer bypass frequencies in ΔumuDC mutD5 strains that were also deficient for Pol I, Pol II, and Pol IV, both singly and in all combinations. Dimer bypass frequencies were not decreased in any of these strains and indeed in some were increased to levels approaching those found in strains containing Pol V. Efficient dimer bypass was, however, entirely dependent on the proofreading deficiency imparted by mutD5, indicating the surprising conclusion that bypass was probably performed by the mutD5 Pol III enzyme itself. This mutant polymerase does not replicate past the much more distorted T-T (6-4) photoadduct, however, suggesting that it may only replicate past lesions, like the T-T dimer, that form base pairs normally.

scholarly journals Mechanism of Abasic Lesion Bypass Catalyzed by a Y-family DNA Polymerase

2007 ◽  
Vol 282 (11) ◽  
pp. 8188-8198 ◽  
Author(s):  
Kevin A. Fiala ◽  
Cameron D. Hypes ◽  
Zucai Suo
Keyword(s):  
Dna Polymerase ◽  
Lesion Bypass ◽  
Y Family

scholarly journals Complex Formation with Rev1 Enhances the Proficiency of Saccharomyces cerevisiae DNA Polymerase ζ for Mismatch Extension and for Extension Opposite from DNA Lesions

2006 ◽  
Vol 26 (24) ◽  
pp. 9555-9563 ◽  
Author(s):  
Narottam Acharya ◽  
Robert E. Johnson ◽  
Satya Prakash ◽  
Louise Prakash
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Polymerase ◽  
Dna Lesions ◽  
Synthetic Activity ◽  
Lesion Bypass ◽  
Dna Lesion ◽  
C Terminus ◽  
Accessory Subunit ◽  
Induced Mutagenesis ◽  
Y Family

ABSTRACT Rev1, a Y family DNA polymerase (Pol) functions together with Polζ, a B family Pol comprised of the Rev3 catalytic subunit and Rev7 accessory subunit, in promoting translesion DNA synthesis (TLS). Extensive genetic studies with Saccharomyces cerevisiae have indicated a requirement of both Polζ and Rev1 for damage-induced mutagenesis, implicating their involvement in mutagenic TLS. Polζ is specifically adapted to promote the extension step of lesion bypass, as it proficiently extends primer termini opposite DNA lesions, and it is also a proficient extender of mismatched primer termini on undamaged DNAs. Since TLS through UV-induced lesions and various other DNA lesions does not depend upon the DNA-synthetic activity of Rev1, Rev1 must contribute to Polζ-dependent TLS in a nonenzymatic way. Here, we provide evidence for the physical association of Rev1 with Polζ and show that this binding is mediated through the C terminus of Rev1 and the polymerase domain of Rev3. Importantly, a rev1 mutant that lacks the C-terminal 72 residues which inactivate interaction with Rev3 exhibits the same high degree of UV sensitivity and defectiveness in UV-induced mutagenesis as that conferred by the rev1Δ mutation. We propose that Rev1 binding to Polζ is indispensable for the targeting of Polζ to the replication fork stalled at a DNA lesion. In addition to this structural role, Rev1 binding enhances the proficiency of Polζ for the extension of mismatched primer termini on undamaged DNAs and for the extension of primer termini opposite DNA lesions.

scholarly journals Complex Formation of Yeast Rev1 and Rev7 Proteins: a Novel Role for the Polymerase-Associated Domain

2005 ◽  
Vol 25 (21) ◽  
pp. 9734-9740 ◽  
Author(s):  
Narottam Acharya ◽  
Lajos Haracska ◽  
Robert E. Johnson ◽  
Ildiko Unk ◽  
Satya Prakash ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Protein Interactions ◽  
Synthetic Activity ◽  
Stable Complex ◽  
Abasic Site ◽  
Protein Protein Interactions ◽  
Lesion Bypass ◽  
Accessory Subunits ◽  
High Degree ◽  
Y Family

ABSTRACT The Rev1 protein of Saccharomyces cerevisiae functions in translesion synthesis (TLS) together with DNA polymerase (Pol) ζ, which is comprised of the Rev3 catalytic and the Rev7 accessory subunits. Rev1, a member of the Y family of Pols, differs from other members in its high degree of specificity for incorporating a C opposite template G as well as opposite an abasic site. Although Rev1 is indispensable for Polζ-dependent TLS, its DNA synthetic activity is not required for many of the Polζ-dependent lesion bypass events. This observation has suggested a structural role for Rev1 in this process. Here we show that in yeast, Rev1 forms a stable complex with Rev7, and the two proteins copurify. Importantly, the polymerase-associated domain (PAD) of Rev1 mediates its binding to Rev7. These observations reveal a novel role for the PAD region of Rev1 in protein-protein interactions, and they raise the possibility of a similar involvement of the PAD of other Y family Pols in protein-protein interactions. We discuss the possible roles of Rev1 versus the Rev1-Rev7 complex in TLS.

scholarly journals Accuracy, lesion bypass, strand displacement and translocation by DNA polymerases

2004 ◽  
Vol 359 (1441) ◽  
pp. 17-23 ◽  
Author(s):  
Thomas A. Steitz ◽  
Y. Whitney Yin
Keyword(s):  
Conformational Change ◽  
Dna Polymerase ◽  
Active Site ◽  
Dna Polymerases ◽  
Lesion Bypass ◽  
Dna Structures ◽  
Template Strand ◽  
Pol I ◽  
Deoxynucleoside Triphosphate ◽  
Copy Dna

The structures of DNA polymerases from different families show common features and significant differences that shed light on the ability of these enzymes to accurately copy DNA and translocate. The structure of a B family DNA polymerase from phage RB69 exhibits an active–site closing conformational change in the fingers domain upon forming a ternary complex with primer template in deoxynucleoside triphosphate. The rotation of the fingers domain α–helices by 60° upon dNTP binding is analogous to the changes seen in other families of polymerases. When the 3' terminus is bound to the editing 3' exonuclease active site, the orientation of the DNA helix axis changes by 40° and the thumb domain re–orients with the DNA. Structures of substrate and product complexes of T7 RNA polymerase, a structural homologue of T7 DNA polymerase, show that family polymerases use the rotation conformational change of the fingers domain to translocate down the DNA. The fingers opening rotation that results in translocation is powered by the release of the product pyrophosphate and also enables the Pol I family polymerases to function as a helicase in displacing the downstream non–template strand from the template strand.

scholarly journals Cryo-EM structure of Pol κ−DNA−PCNA holoenzyme and implications for polymerase switching in DNA lesion bypass

2020 ◽  
Author(s):  
Claudia Lancey ◽  
Muhammad Tehseen ◽  
Masateru Takahashi ◽  
Mohamed A. Sobhy ◽  
Timothy J. Ragan ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Active Site ◽  
Genome Instability ◽  
Structural Basis ◽  
Conformational Sampling ◽  
Lesion Bypass ◽  
Catalytic Core ◽  
Dna Lesion ◽  
Thumb Domain ◽  
C Terminus

Replacement of the stalled replicative polymerase (Pol δ) at a DNA lesion by the error-prone DNA polymerase κ (Pol κ) restarts synthesis past the lesion to prevent genome instability. The switching from Pol δ to Pol κ is mediated by the processivity clamp PCNA but the structural basis of this mechanism is unknown. We determined the Cryo-EM structures of human Pol κ–DNA–PCNA complex and of a stalled Pol δ–DNA–PCNA complex at 3.9 and 4.7 Å resolution, respectively. In Pol κ complex, the C-terminus of the PAD domain docks the catalytic core to one PCNA protomer in an angled orientation, bending the DNA exiting Pol κ active site through PCNA. In Pol δ complex, the DNA is disengaged from the active site but is retained by the thumb domain. We present a model for polymerase switching facilitated by Pol κ recruitment to PCNA and Pol κ conformational sampling to seize the DNA from stalled Pol δ assisted by PCNA tilting.

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