scholarly journals SOS induction in mycobacteria: analysis of the DNA‐binding activity of a LexA‐like repressor and its role in DNA damage induction of the recA gene from Mycobacterium smegmatis

1997 ◽  
Vol 26 (4) ◽  
pp. 643-653 ◽  
Author(s):  
Steven I. Durbach ◽  
Susan J. Andersen ◽  
Valerie Mizrahi
Keyword(s):  
Dna Damage ◽  
Dna Binding ◽  
Mycobacterium Smegmatis ◽  
Binding Activity ◽  
Reca Gene ◽  
Dna Binding Activity ◽  
Sos Induction

scholarly journals Cu,Zn-superoxide dismutase deficiency in mice leads to organ-specific increase in oxidatively damaged DNA and NF-κB1 protein activity.

2010 ◽  
Vol 57 (4) ◽  
Author(s):  
Agnieszka Siomek ◽  
Kamil Brzoska ◽  
Barbara Sochanowicz ◽  
Daniel Gackowski ◽  
Rafal Rozalski ◽  
...  
Keyword(s):  
Dna Damage ◽  
Superoxide Dismutase ◽  
Dna Binding ◽  
Urinary Excretion ◽  
Oxidative Dna Damage ◽  
Binding Activity ◽  
Wild Type ◽  
Dna Binding Activity ◽  
Organ Specific ◽  
Damaged Dna

Earlier experimental studies have demonstrated that: i) Cu,Zn-superoxide dismutase deficiency leads to oxidative stress and carcinogenesis; ii) dysregulation of NF-κB pathway can mediate a wide variety of diseases, including cancer. Therefore, we decided, for the first time, to examine the level of oxidative DNA damage and the DNA binding activity of NF-κB proteins in SOD1 knockout, heterozygous and wild-type mice. Two kinds of biomarkers of oxidatively damaged DNA: urinary excretion of 8-oxodG and 8-oxoGua, and the level of oxidatively damaged DNA were analysed using HPLC-GC-MS and HPLC-EC. The DNA binding activity of p50 and p65 proteins in a nuclear extracts was assessed using NF-κB p50/p65 EZ-TFA transcription factor assay. These parameters were determined in the brain, liver, kidney and urine of SOD1 knockout, heterozygous and wild-type mice. The level of 8-oxodG in DNA was higher in the liver and kidney of knockout mice than in wild type. No differences were found in urinary excretion of 8-oxoGua and 8-oxodG between wild type and the SOD1-deficient animals. The activity of the p50 protein was higher in the kidneys, but surprisingly not in the livers of SOD1-deficient mice, whereas p65 activity did not show any variability. Our results indicate that in Cu,Zn-SOD-deficient animals the level of oxidative DNA damage and NF-κB1 activity are elevated in certain organs only, which may provide some explanation for organ-specific ROS-induced carcinogenesis.

scholarly journals Structure analysis of FAAP24 reveals single-stranded DNA-binding activity and domain functions in DNA damage response

Cell Research ◽  
2013 ◽  
Vol 23 (10) ◽  
pp. 1215-1228 ◽  
Author(s):  
Yucai Wang ◽  
Xiao Han ◽  
Fangming Wu ◽  
Justin W Leung ◽  
Megan G Lowery ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Binding ◽  
Structure Analysis ◽  
Dna Damage Response ◽  
Binding Activity ◽  
Single Stranded Dna ◽  
Dna Binding Activity ◽  
Damage Response

scholarly journals NAD+ Modulates p53 DNA Binding Specificity and Function

2004 ◽  
Vol 24 (22) ◽  
pp. 9958-9967 ◽  
Author(s):  
Kevin G. McLure ◽  
Masatoshi Takagi ◽  
Michael B. Kastan
Keyword(s):  
Dna Damage ◽  
Dna Binding ◽  
Binding Specificity ◽  
Binding Activity ◽  
Vitamin B ◽  
Cancer Therapies ◽  
Dna Binding Activity ◽  
Dna Binding Specificity

ABSTRACT DNA damage induces p53 DNA binding activity, which affects tumorigenesis, tumor responses to therapies, and the toxicities of cancer therapies (B. Vogelstein, D. Lane, and A. J. Levine, Nature 408:307-310, 2000; K. H. Vousden and X. Lu, Nat. Rev. Cancer 2:594-604, 2002). Both transcriptional and transcription-independent activities of p53 contribute to DNA damage-induced cell cycle arrest, apoptosis, and aneuploidy prevention (M. B. Kastan et al., Cell 71:587-597, 1992; K. H. Vousden and X. Lu, Nat. Rev. Cancer 2:594-604, 2002). Small-molecule manipulation of p53 DNA binding activity has been an elusive goal, but here we show that NAD+ binds to p53 tetramers, induces a conformational change, and modulates p53 DNA binding specificity in vitro. Niacinamide (vitamin B3) increases the rate of intracellular NAD+ synthesis, alters radiation-induced p53 DNA binding specificity, and modulates activation of a subset of p53 transcriptional targets. These effects are likely due to a direct effect of NAD+ on p53, as a molecule structurally related to part of NAD+, TDP, also inhibits p53 DNA binding, and the TDP precursor, thiamine (vitamin B1), inhibits intracellular p53 activity. Niacinamide and thiamine affect two p53-regulated cellular responses to ionizing radiation: rereplication and apoptosis. Thus, niacinamide and thiamine form a novel basis for the development of small molecules that affect p53 function in vivo, and these results suggest that changes in cellular energy metabolism may regulate p53.

scholarly journals The Role of UAF1 in the Fanconi Anemia Pathway Regulation of Homologous Recombination-Mediated Genome Maintenance

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1041-1041
Author(s):  
Fengshan Liang ◽  
Simonne Longerich ◽  
Caroline Tang ◽  
Olga Buzovestsky ◽  
Yong Xiong ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Dna Binding ◽  
Homologous Recombination ◽  
Complex Formation ◽  
Binding Activity ◽  
Genome Maintenance ◽  
Synaptic Complex ◽  
Dna Binding Activity ◽  
D Loop

Abstract Background: Fanconi anemia (FA), a cancer-prone genetic disease, is caused by defects in the FA-DNA repair pathway. In response to DNA interstrand crosslink (ICL)-induced DNA damage, FANCI-FANCD2 mono-ubiquitination licenses the execution of downstream DNA damage signaling and repair steps, including repair by homologous recombination (HR) that utilizes the recombinase RAD51 and its cohort of accessory factors. Timely deubiquitination of FANCD2 by the UAF1-USP1 deubiquitinating enzyme complex is also critically important for the FA pathway. As such, UAF1 depletion results in persistent FANCD2 ubiquitination and DNA damage hypersensitivity. UAF1 deficient cells are also impaired for DNA repair by homologous recombination. UAF1 physically associates with RAD51AP1, a protein that enhances the activity of the RAD51 recombinase. It remains to be defined how UAF1 regulates homologous recombination and genome stability. Methods: Highly purified proteins were used to define the DNA binding activity and protein interaction of UAF1. In vitroD-loop formation reaction and synaptic complex assembly assay were used to discover the function of UAF1 in RAD51 recombinase enhancement. HeLa and U2OS-DR-GFP cell lines with impaired UAF1-RAD51AP1 interaction or UAF1 DNA binding were generated to examine DNA-damage agent sensitivity and HR efficiency. Results: (1) UAF1 possesses a DNA binding activity capable of engaging ssDNA, dsDNA and has a preference for the D-loop DNA substrate. We further identified that the N-terminus but not C-terminal SLD domain of UAF1 binds DNA. (2) UAF1 forms a dimeric complex with RAD51AP1. Our results also revealed a trimeric complex of RAD51-RAD51AP1-UAF1, with RAD51AP1 providing a tethering function between the other two proteins. (3) The RAD51AP1-UAF1 interaction interface was defined showing a novel SIM motif in the middle portion of RAD51AP1and the SLD1-SLD2 domain of UAF1 mediate protein complex formation. Based on the domain mapping results, point mutants of RAD51AP1 and UAF1 that are specifically compromised for the formation of the RAD51AP1-UAF1 complex were generated. (4) UAF1 synergizes with RAD51AP1 in the RAD51-mediated D-loop reaction and that this functional synergy requires the RAD51AP1-UAF1 complex and also the DNA and RAD51 binding attributes of RAD51AP1. (5) RAD51AP1-UAF1 works in conjunction with the RAD51 presynaptic filament in the capture of the duplex DNA partner and in the assembly of the synaptic complex. (6) Human cell lines impaired for RAD51AP1-UAF1 complex formation are compromised for the ability to repair DNA damage and to execute HR. (7) DNA repair function of the RAD51AP1-UAF1 complex is likely USP1-independent. Conclusions: The physical interaction between UAF1 and RAD51AP1 is indispensable for functional synergy in vitro and, accordingly, for the biological function of UAF1 in HR and DNA damage repair. Our findings provide insights into a novel USP1-independent regulatory mechanism of UAF1 on homologous recombination-mediated genome maintenance. Disclosures No relevant conflicts of interest to declare.

scholarly journals UAF1 DNA Binding Activity Is Critical for RAD51-Mediated Homologous DNA Pairing

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 2497-2497
Author(s):  
Fengshan Liang ◽  
Adam S Miller ◽  
Carolilne Tang ◽  
Patrick Sung ◽  
Gary M. Kupfer
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Dna Binding ◽  
Complex Formation ◽  
Binding Activity ◽  
Loop Formation ◽  
Dna Binding Activity ◽  
D Loop ◽  
In Cells ◽  
Dna Pairing

Background: In the Fanconi anemia (FA) DNA repair pathway, DNA damage induces the mono-ubiquitination of the FANCI-FANCD2 (ID2) heterodimer by the FA core complex through its inherent E3 ligase activity. The timely deubiquitination of ID2 by USP1-UAF1 deubiquitinase complex is also critically important for the FA DNA repair. UAF1 has a DNA binding activity, which is required for FANCD2 deubiquitination. UAF1 also enhances RAD51-mediated homologous DNA pairing in a manner that is dependent on complex formation with RAD51AP1. UAF1 deficient cells are impaired for DNA repair by homologous recombination (HR).The biochemical and cellular functions of UAF1 DNA binding activity in HR remain elusive. Methods:UAF1 wild type and DNA binding mutant proteins were purified and used to define its biochemical properties in HR. In vitroD-loop formation and synaptic complex assembly assay were performed to discover the DNA binding of UAF1 in RAD51 recombinase enhancement. U2OS-DR-GFP cell lines with impaired UAF1 or RAD51AP1DNA binding were generated to examine HR efficiency and DNA damage resistance. Results:UAF1 preferentially binds an HR-intermediate-like DNA substrate (D-loop, Fig.1). The DNA binding deficient mutant of UAF1 is unable to stimulate RAD51AP1 promotion of RAD51-mediated D-loop (Fig. 2) and the ability to recruit homologous DNA to form the presynaptic complex formation in HR (Fig. 3). In cells, the UAF1 DNA-binding mutant is compromised for the ability to repair DNA damage and to implement HR (Fig. 4). Such activity correlates with the ability to confer resistance to DNA cross linking agents such as mitomycin C (Fig. 4). The DNA binding of UAF1 and RAD51AP1 have a coordinated role in HR-directed DNA damage repair (Fig. 5). Conclusions: UAF1 DNA binding activity is indispensable for its function in enhancing RAD51-mediated homologous DNA pairing within the context of the UAF1-RAD51AP1 complex. UAF1 DNA binding deficiency causes DNA damage sensitivity and impairs HR efficiency in cells. Translational Applicability:Our findings reveal a critical role of UAF1 DNA binding in DNA repair and genome maintenance. The identification of UAF1's role in repair will enable targeted efforts to improve molecular approaches for FA therapy. Disclosures No relevant conflicts of interest to declare.

scholarly journals Identification of an XRCC1 DNA binding activity essential for retention at sites of DNA damage

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Mac C. Y. Mok ◽  
Anna Campalans ◽  
Monica C. Pillon ◽  
Alba Guarné ◽  
J. Pablo Radicella ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Binding ◽  
Binding Activity ◽  
Dna Binding Activity

scholarly journals The DNA-binding activity of USP1-associated factor 1 is required for efficient RAD51-mediated homologous DNA pairing and homology-directed DNA repair

2020 ◽  
Vol 295 (24) ◽  
pp. 8186-8194 ◽  
Author(s):  
Fengshan Liang ◽  
Adam S. Miller ◽  
Caroline Tang ◽  
David Maranon ◽  
Elizabeth A. Williamson ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Dna Binding ◽  
Fanconi Anemia ◽  
Complementation Group ◽  
Binding Activity ◽  
Associated Factor ◽  
Dna Binding Activity ◽  
Damage Repair ◽  
Dna Pairing

USP1-associated factor 1 (UAF1) is an integral component of the RAD51-associated protein 1 (RAD51AP1)–UAF1-ubiquitin-specific peptidase 1 (USP1) trimeric deubiquitinase complex. This complex acts on DNA-bound, monoubiquitinated Fanconi anemia complementation group D2 (FANCD2) protein in the Fanconi anemia pathway of the DNA damage response. Moreover, RAD51AP1 and UAF1 cooperate to enhance homologous DNA pairing mediated by the recombinase RAD51 in DNA repair via the homologous recombination (HR) pathway. However, whereas the DNA-binding activity of RAD51AP1 has been shown to be important for RAD51-mediated homologous DNA pairing and HR-mediated DNA repair, the role of DNA binding by UAF1 in these processes is unclear. We have isolated mutant UAF1 variants that are impaired in DNA binding and tested them together with RAD51AP1 in RAD51-mediated HR. This biochemical analysis revealed that the DNA-binding activity of UAF1 is indispensable for enhanced RAD51 recombinase activity within the context of the UAF1–RAD51AP1 complex. In cells, DNA-binding deficiency of UAF1 increased DNA damage sensitivity and impaired HR efficiency, suggesting that UAF1 and RAD51AP1 have coordinated roles in DNA binding during HR and DNA damage repair. Our findings show that even though UAF1's DNA-binding activity is redundant with that of RAD51AP1 in FANCD2 deubiquitination, it is required for efficient HR-mediated chromosome damage repair.

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