Temporal dynamics of base excision / single-strand break repair protein complex assembly and disassembly are modulated by the PARP1/NAD+/SIRT6 axis

SUMMARYAssembly and disassembly of DNA repair protein complexes at sites of DNA damage is essential to maintain genomic integrity. We investigated factors coordinating assembly of the base excision repair (BER) proteins, DNA polymerase β (Polβ) and XRCC1, to DNA lesion sites, identifying a new role for Polβ in regulating XRCC1 disassembly from DNA repair complexes and conversely, demonstrating Polβ’s dependence on XRCC1 for complex assembly. RealPAR, a genetically-encoded probe for live cell imaging of poly(ADP-ribose) (PAR), reveals that Polβ and XRCC1 require PAR for repair complex assembly and PAR degradation for disassembly. We find that BER complex assembly is further modulated by attenuation / augmentation of NAD+ biosynthesis. Finally, SIRT6 does not regulate PARP1 activation but impairs XRCC1 recruitment, leading to diminished Polβ abundance at sites of DNA damage. These findings highlight coordinated yet independent roles for both PARP1 and SIRT6 and their regulation by NAD+ bioavailability to facilitate BER.

Download Full-text

Faculty Opinions recommendation of Temporal dynamics of base excision/single-strand break repair protein complex assembly/disassembly are modulated by the PARP/NAD+/SIRT6 axis.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.741104035.793590825 ◽

2021 ◽

Author(s):

Olga Lavrik

Keyword(s):

Protein Complex ◽

Temporal Dynamics ◽

Strand Break ◽

Single Strand ◽

Complex Assembly ◽

Single Strand Break ◽

Single Strand Break Repair ◽

Repair Protein ◽

Base Excision ◽

Protein Complex Assembly

Download Full-text

Motor Coordination in Space: An Analysis of the Effects of Microgravity on XRCCI DNA Repair Protein Function

10.21203/rs.3.pex-1267/v1 ◽

2020 ◽

Author(s):

Vishruth Nagam

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Protein Function ◽

Motor Coordination ◽

Strand Break ◽

Single Strand Break ◽

Single Strand Break Repair ◽

Repair Protein ◽

Mature Neurons ◽

Microgravity Conditions

Abstract While in space, astronauts have been known to face exposure to stressors that may increase susceptibility to DNA damage. If DNA repair proteins are defective or nonexistent, DNA mutations may accumulate, causing increasingly abnormal function as one ages [1]. The DNA single-strand break repair protein XRCC1 is important for cerebellar neurogenesis and interneuron development [2]. According to previous studies, a deficiency of XRCC1 can lead to an increase in DNA damage, in mature neurons, and ataxia (a progressive loss of motor coordination) [2]. I propose to address how XRCC1’s efficiency can change in microgravity conditions. This experiment’s relevance is underscored by the importance of motor coordination and physical fitness for astronauts; determining the potential effects of microgravity on XRCC1 is crucial for future space exploration.

Download Full-text

Dynamic Compartmentalization of Base Excision Repair Proteins in Response to Nuclear and Mitochondrial Oxidative Stress

Molecular and Cellular Biology ◽

10.1128/mcb.01357-08 ◽

2008 ◽

Vol 29 (3) ◽

pp. 794-807 ◽

Cited By ~ 31

Author(s):

Lyra M. Griffiths ◽

Dan Swartzlander ◽

Kellen L. Meadows ◽

Keith D. Wilkinson ◽

Anita H. Corbett ◽

...

Keyword(s):

Oxidative Stress ◽

Dna Damage ◽

Dna Repair ◽

Base Excision Repair ◽

Oxidative Dna Damage ◽

Excision Repair ◽

Intracellular Localization ◽

Genotoxic Stress ◽

Mitochondrial Oxidative Stress ◽

Base Excision

ABSTRACT DNAs harbored in both nuclei and mitochondria of eukaryotic cells are subject to continuous oxidative damage resulting from normal metabolic activities or environmental insults. Oxidative DNA damage is primarily reversed by the base excision repair (BER) pathway, initiated by N-glycosylase apurinic/apyrimidinic (AP) lyase proteins. To execute an appropriate repair response, BER components must be distributed to accommodate levels of genotoxic stress that may vary considerably between nuclei and mitochondria, depending on the growth state and stress environment of the cell. Numerous examples exist where cells respond to signals, resulting in relocalization of proteins involved in key biological transactions. To address whether such dynamic localization contributes to efficient organelle-specific DNA repair, we determined the intracellular localization of the Saccharomyces cerevisiae N-glycosylase/AP lyases, Ntg1 and Ntg2, in response to nuclear and mitochondrial oxidative stress. Fluorescence microscopy revealed that Ntg1 is differentially localized to nuclei and mitochondria, likely in response to the oxidative DNA damage status of the organelle. Sumoylation is associated with targeting of Ntg1 to nuclei containing oxidative DNA damage. These studies demonstrate that trafficking of DNA repair proteins to organelles containing high levels of oxidative DNA damage may be a central point for regulating BER in response to oxidative stress.

Download Full-text

Scanning Force Microscopy and Nanomangulation: Studies of Dna and Proteins Involved in Dna Repair

Microscopy and Microanalysis ◽

10.1017/s1431927600018341 ◽

1999 ◽

Vol 5 (S2) ◽

pp. 1004-1005

Author(s):

Dorothy Erie ◽

Glenn Ratcliff ◽

Martin Guthold ◽

Valerie Bullock ◽

Michelle Pliske ◽

...

Keyword(s):

Dna Repair ◽

Physical Properties ◽

Conformational Changes ◽

Excision Repair ◽

Protein Complexes ◽

Scanning Force Microscopy ◽

Force Microscopy ◽

Scanning Force ◽

Base Excision ◽

User Friendly

Repair of damaged or incorrectly matched DNA is essential to the survival of all organisms. Consequently cells have devised a plentitude of pathways for repair. We have been investigating the mechanisms of mismatch repair and base excision repair. Both of these repair processes involve a large number of proteins that interact with one another as well as with DNA. Our long-term goal is to assemble complexes that are fully functional for DNA repair and to image the process of DNA repair. In addition, we wish to i) determine the stoicheometry of binding of the protein complexes to each other and to DNA, ii) monitor conformational changes due to substrate binding, iii) measure physical properties of DNA and the complexes. To accomplish this end, we have endeavored to improve techniques for solution imaging as well as those for data analysis. In this presentation I will discuss data on the stoicheometry of binding in several protein complexes and data on the physical properties of DNA.To measure the physical properties of DNA, we utilize a nanoManipulator, a modified Scanning Force Microscope with a novel, user-friendly interface that allows easy and controlled manipulation of nanometer-sized samples.

Download Full-text

Databases and Bioinformatics Tools for the Study of DNA Repair

Molecular Biology International ◽

10.4061/2011/475718 ◽

2011 ◽

Vol 2011 ◽

pp. 1-9 ◽

Cited By ~ 1

Author(s):

Kaja Milanowska ◽

Kristian Rother ◽

Janusz M. Bujnicki

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Excision Repair ◽

Uv Light ◽

Dna Lesions ◽

Environmental Chemicals ◽

Nonhomologous End Joining ◽

End Joining ◽

Repair Mechanisms ◽

Base Excision

DNA is continuously exposed to many different damaging agents such as environmental chemicals, UV light, ionizing radiation, and reactive cellular metabolites. DNA lesions can result in different phenotypical consequences ranging from a number of diseases, including cancer, to cellular malfunction, cell death, or aging. To counteract the deleterious effects of DNA damage, cells have developed various repair systems, including biochemical pathways responsible for the removal of single-strand lesions such as base excision repair (BER) and nucleotide excision repair (NER) or specialized polymerases temporarily taking over lesion-arrested DNA polymerases during the S phase in translesion synthesis (TLS). There are also other mechanisms of DNA repair such as homologous recombination repair (HRR), nonhomologous end-joining repair (NHEJ), or DNA damage response system (DDR). This paper reviews bioinformatics resources specialized in disseminating information about DNA repair pathways, proteins involved in repair mechanisms, damaging agents, and DNA lesions.

Download Full-text

Control of genome stability by Slx protein complexes

Biochemical Society Transactions ◽

10.1042/bst0370495 ◽

2009 ◽

Vol 37 (3) ◽

pp. 495-510 ◽

Cited By ~ 26

Author(s):

John Rouse

Keyword(s):

Dna Damage ◽

Dna Replication ◽

Protein Interactions ◽

Cellular Response ◽

Genome Stability ◽

Molecular Mechanisms ◽

Excision Repair ◽

Protein Complexes ◽

Alkylating Agents ◽

Strand Break

The six Saccharomyces cerevisiae SLX genes were identified in a screen for factors required for the viability of cells lacking Sgs1, a member of the RecQ helicase family involved in processing stalled replisomes and in the maintenance of genome stability. The six SLX gene products form three distinct heterodimeric complexes, and all three have catalytic activity. Slx3–Slx2 (also known as Mus81–Mms4) and Slx1–Slx4 are both heterodimeric endonucleases with a marked specificity for branched replication fork-like DNA species, whereas Slx5–Slx8 is a SUMO (small ubiquitin-related modifier)-targeted E3 ubiquitin ligase. All three complexes play important, but distinct, roles in different aspects of the cellular response to DNA damage and perturbed DNA replication. Slx4 interacts physically not only with Slx1, but also with Rad1–Rad10 [XPF (xeroderma pigmentosum complementation group F)–ERCC1 (excision repair cross-complementing 1) in humans], another structure-specific endonuclease that participates in the repair of UV-induced DNA damage and in a subpathway of recombinational DNA DSB (double-strand break) repair. Curiously, Slx4 is essential for repair of DSBs by Rad1–Rad10, but is not required for repair of UV damage. Slx4 also promotes cellular resistance to DNA-alkylating agents that block the progression of replisomes during DNA replication, by facilitating the error-free mode of lesion bypass. This does not require Slx1 or Rad1–Rad10, and so Slx4 has several distinct roles in protecting genome stability. In the present article, I provide an overview of our current understanding of the cellular roles of the Slx proteins, paying particular attention to the advances that have been made in understanding the cellular roles of Slx4. In particular, protein–protein interactions and underlying molecular mechanisms are discussed and I draw attention to the many questions that have yet to be answered.

Download Full-text

The DNA repair protein NBS1 influences the base excision repair pathway

Carcinogenesis ◽

10.1093/carcin/bgp004 ◽

2009 ◽

Vol 30 (3) ◽

pp. 408-415 ◽

Cited By ~ 10

Author(s):

D. Sagan ◽

R. Muller ◽

C. Kroger ◽

A. Hematulin ◽

S. Mortl ◽

...

Keyword(s):

Dna Repair ◽

Base Excision Repair ◽

Excision Repair ◽

Repair Pathway ◽

Base Excision Repair Pathway ◽

Repair Protein ◽

Dna Repair Protein ◽

Base Excision

Download Full-text

DNA polymerase X from Deinococcus radiodurans implicated in bacterial tolerance to DNA damage is characterized as a short patch base excision repair polymerase

Microbiology ◽

10.1099/mic.0.029223-0 ◽

2009 ◽

Vol 155 (9) ◽

pp. 3005-3014 ◽

Cited By ~ 19

Author(s):

Nivedita P. Khairnar ◽

Hari S. Misra

Keyword(s):

Dna Damage ◽

Dna Polymerase ◽

Base Excision Repair ◽

Deinococcus Radiodurans ◽

Excision Repair ◽

Strand Break ◽

Klenow Fragment ◽

E Coli ◽

Base Excision

The Deinococcus radiodurans R1 genome encodes an X-family DNA repair polymerase homologous to eukaryotic DNA polymerase β. The recombinant deinococcal polymerase X (PolX) purified from transgenic Escherichia coli showed deoxynucleotidyltransferase activity. Unlike the Klenow fragment of E. coli, this enzyme showed short patch DNA synthesis activity on heteropolymeric DNA substrate. The recombinant enzyme showed 5′-deoxyribose phosphate (5′-dRP) lyase activity and base excision repair function in vitro, with the help of externally supplied glycosylase and AP endonuclease functions. A polX disruption mutant of D. radiodurans expressing 5′-dRP lyase and a truncated polymerase domain was comparatively less sensitive to γ-radiation than a polX deletion mutant. Both mutants showed higher sensitivity to hydrogen peroxide. Excision repair mutants of E. coli expressing this polymerase showed functional complementation of UV sensitivity. These results suggest the involvement of deinococcal polymerase X in DNA-damage tolerance of D. radiodurans, possibly by contributing to DNA double-strand break repair and base excision repair.

Download Full-text