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1.
DNA Repair (Amst) ; 141: 103728, 2024 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-39029374

RESUMO

Nucleotide excision repair (NER) clears genomes of DNA adducts formed by UV light, environmental agents, and antitumor drugs. Gene mutations that lead to defects in the core NER reaction cause the skin cancer-prone disease xeroderma pigmentosum. In NER, DNA lesions are excised within an oligonucleotide of 25-30 residues via a complex, multi-step reaction that is regulated by protein-protein interactions. These interactions were first characterized in the 1990s using pull-down, co-IP and yeast two-hybrid assays. More recently, high-resolution structures and detailed functional studies have started to yield detailed pictures of the progression along the NER reaction coordinate. In this review, we highlight how the study of interactions among proteins by structural and/or functional studies have provided insights into the mechanisms by which the NER machinery recognizes and excises DNA lesions. Furthermore, we identify reported, but poorly characterized or unsubstantiated interactions in need of further validation.

2.
Nanomedicine (Lond) ; 2024 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-38722243

RESUMO

Aim: To investigate whether medical devices coated with a synthesized nanocomposite of poly(methylmethacrylate-co-dimethyl acrylamide) (PMMDMA) and silver nanoparticles (AgNPs) could improve their antibiofilm and antimicrobial activities. We also investigated the nanocomposite's safety. Materials & methods: The nanocomposite was synthesized and characterized using analytical techniques. Medical devices coated with the nanocomposite were evaluated for bacterial adhesion and hemolytic activity in vitro. Results: The nanocomposite formation was demonstrated with the incorporation of AgNPs into the polymer matrix. The nanocomposite proved to be nonhemolytic and significantly inhibited bacterial biofilm formation. Conclusion: The PMMDMA-AgNPs nanocomposite was more effective in preventing biofilm formation than PMMDMA alone and is a promising strategy for coating medical devices and reducing mortality due to hospital-acquired infections.

3.
NAR Cancer ; 6(1): zcae013, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38500596

RESUMO

Nucleotide excision repair (NER) reduces efficacy of treatment with platinum (Pt)-based chemotherapy by removing Pt lesions from DNA. Previous study has identified that missense mutation or loss of the NER genes Excision Repair Cross Complementation Group 1 and 2 (ERCC1 and ERCC2) leads to improved patient outcomes after treatment with Pt-based chemotherapies. Although most NER gene alterations found in patient tumors are missense mutations, the impact of mutations in the remaining nearly 20 NER genes is unknown. Towards this goal, we previously developed a machine learning strategy to predict genetic variants in an essential NER protein, Xeroderma Pigmentosum Complementation Group A (XPA), that disrupt repair. In this study, we report in-depth analyses of a subset of the predicted variants, including in vitro analyses of purified recombinant protein and cell-based assays to test Pt agent sensitivity in cells and determine mechanisms of NER dysfunction. The most NER deficient variant Y148D had reduced protein stability, weaker DNA binding, disrupted recruitment to damage, and degradation. Our findings demonstrate that tumor mutations in XPA impact cell survival after cisplatin treatment and provide valuable mechanistic insights to improve variant effect prediction. Broadly, these findings suggest XPA tumor variants should be considered when predicting chemotherapy response.

4.
Nat Commun ; 15(1): 1388, 2024 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-38360910

RESUMO

Most genotoxic anticancer agents fail in tumors with intact DNA repair. Therefore, trabectedin, anagent more toxic to cells with active DNA repair, specifically transcription-coupled nucleotide excision repair (TC-NER), provides therapeutic opportunities. To unlock the potential of trabectedin and inform its application in precision oncology, an understanding of the mechanism of the drug's TC-NER-dependent toxicity is needed. Here, we determine that abortive TC-NER of trabectedin-DNA adducts forms persistent single-strand breaks (SSBs) as the adducts block the second of the two sequential NER incisions. We map the 3'-hydroxyl groups of SSBs originating from the first NER incision at trabectedin lesions, recording TC-NER on a genome-wide scale. Trabectedin-induced SSBs primarily occur in transcribed strands of active genes and peak near transcription start sites. Frequent SSBs are also found outside gene bodies, connecting TC-NER to divergent transcription from promoters. This work advances the use of trabectedin for precision oncology and for studying TC-NER and transcription.


Assuntos
Reparo por Excisão , Neoplasias , Humanos , Trabectedina , Transcrição Gênica , Medicina de Precisão , Reparo do DNA , Dano ao DNA , DNA/genética , Nucleotídeos , Quebras de DNA
5.
NAR Cancer ; 5(4): zcad057, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-38058548

RESUMO

The therapeutic efficacy of cisplatin and oxaliplatin depends on the balance between the DNA damage induction and the DNA damage response of tumor cells. Based on clinical evidence, oxaliplatin is administered to cisplatin-unresponsive cancers, but the underlying molecular causes for this tumor specificity are not clear. Hence, stratification of patients based on DNA repair profiling is not sufficiently utilized for treatment selection. Using a combination of genetic, transcriptomics and imaging approaches, we identified factors that promote global genome nucleotide excision repair (GG-NER) of DNA-platinum adducts induced by oxaliplatin, but not by cisplatin. We show that oxaliplatin-DNA lesions are a poor substrate for GG-NER initiating factor XPC and that DDB2 and HMGA2 are required for efficient binding of XPC to oxaliplatin lesions and subsequent GG-NER initiation. Loss of DDB2 and HMGA2 therefore leads to hypersensitivity to oxaliplatin but not to cisplatin. As a result, low DDB2 levels in different colon cancer cells are associated with GG-NER deficiency and oxaliplatin hypersensitivity. Finally, we show that colon cancer patients with low DDB2 levels have a better prognosis after oxaliplatin treatment than patients with high DDB2 expression. We therefore propose that DDB2 is a promising predictive marker of oxaliplatin treatment efficiency in colon cancer.

6.
iScience ; 26(7): 107219, 2023 Jul 21.
Artigo em Inglês | MEDLINE | ID: mdl-37529320

RESUMO

The vast spectrum of clinical features of COVID-19 keeps challenging scientists and clinicians. Low resistance to infection might result in long-term viral persistence, but the underlying mechanisms remain unclear. Here, we studied the immune response of immunocompetent COVID-19 patients with prolonged SARS-CoV-2 infection by immunophenotyping, cytokine and serological analysis. Despite viral loads and symptoms comparable to regular mildly symptomatic patients, long-term carriers displayed weaker systemic IFN-I responses and fewer circulating pDCs and NK cells at disease onset. Type 1 cytokines remained low, while type-3 cytokines were in turn enhanced. Of interest, we observed no defects in antigen-specific cytotoxic T cell responses, and circulating antibodies displayed higher affinity against different variants of SARS-CoV-2 Spike protein in these patients. The identification of distinct immune responses in long-term carriers adds up to our understanding of essential host protective mechanisms to ensure tissue damage control despite prolonged viral infection.

7.
NAR Cancer ; 5(3): zcad042, 2023 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-37554969

RESUMO

Targeting BRCA1- and BRCA2-deficient tumors through synthetic lethality using poly(ADP-ribose) polymerase inhibitors (PARPi) has emerged as a successful strategy for cancer therapy. PARPi monotherapy has shown excellent efficacy and safety profiles in clinical practice but is limited by the need for tumor genome mutations in BRCA or other homologous recombination genes as well as the rapid emergence of resistance. In this study, we identified 2-chloro-N,N-diethylethanamine hydrochloride (CDEAH) as a small molecule that selectively kills PARP1- and xeroderma pigmentosum A-deficient cells. CDEAH is a monofunctional alkylating agent that preferentially alkylates guanine nucleobases, forming DNA adducts that can be removed from DNA by either a PARP1-dependent base excision repair or nucleotide excision repair. Treatment of PARP1-deficient cells leads to the formation of strand breaks, an accumulation of cells in S phase and activation of the DNA damage response. Furthermore, CDEAH selectively inhibits PARP1-deficient xenograft tumor growth compared to isogenic PARP1-proficient tumors. Collectively, we report the discovery of an alkylating agent inducing DNA damage that requires PARP1 activity for repair and acts synergistically with PARPi.

8.
bioRxiv ; 2023 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-37425789

RESUMO

Nucleotide excision repair (NER) neutralizes treatment with platinum (Pt)-based chemotherapy by removing Pt lesions from DNA. Previous study has identified that missense mutation or loss of either of the NER genes Excision Repair Cross Complementation Group 1 and 2 ( ERCC1 and ERCC2 ) leads to improved patient outcomes after treatment with Pt-based chemotherapies. Although most NER gene alterations found in patient tumors are missense mutations, the impact of such mutations in the remaining nearly 20 NER genes is unknown. Towards this goal, we previously developed a machine learning strategy to predict genetic variants in an essential NER scaffold protein, Xeroderma Pigmentosum Complementation Group A (XPA), that disrupt repair activity on a UV-damaged substrate. In this study, we report in-depth analyses of a subset of the predicted NER-deficient XPA variants, including in vitro analyses of purified recombinant protein and cell-based assays to test Pt agent sensitivity in cells and determine mechanisms of NER dysfunction. The most NER deficient variant Y148D had reduced protein stability, weaker DNA binding, disrupted recruitment to damage, and degradation resulting from tumor missense mutation. Our findings demonstrate that tumor mutations in XPA impact cell survival after cisplatin treatment and provide valuable mechanistic insights to further improve variant effect prediction efforts. More broadly, these findings suggest XPA tumor variants should be considered when predicting patient response to Pt-based chemotherapy. Significance: A destabilized, readily degraded tumor variant identified in the NER scaffold protein XPA sensitizes cells to cisplatin, suggesting that XPA variants can be used to predict response to chemotherapy.

9.
Chem Res Toxicol ; 36(6): 822-837, 2023 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-37208809

RESUMO

Cisplatin (CP) is a common antitumor drug that is used to treat many solid tumors. The activity of CP is attributed to the formation of DNA-DNA cross-links, which consist of 1,2-intra-, 1,3-intra-, and interstrand cross-links. To better understand how each intrastrand cross-link contributes to the activity of CP, we have developed comprehensive ultraperformance liquid chromatography-selective ion monitoring (UPLC-SIM) assays to quantify 1,2-GG-, 1,2-AG-, 1,3-GCG-, and 1,3-GTG-intrastrand cross-links. The limit of quantitation for the developed assays ranged from 5 to 50 fmol or as low as 6 cross-links per 108 nucleotides. To demonstrate the utility of the UPLC-SIM assays, we first performed in vitro cross-link formation kinetics experiments. We confirmed that the 1,2-GG-intrastrand cross-links were the most abundant intrastrand cross-link and formed at a faster rate compared to 1,2-AG- and 1,3-intrastrand cross-links. Furthermore, we investigated the repair kinetics of intrastrand cross-links in CP-treated wild-type and nucleotide excision repair (NER)-deficient U2OS cells. We observed a slow decrease of both 1,2- and 1,3-intrastrand cross-links in wild-type cells and no evidence of direct repair in the NER-deficient cells. Taken together, we have demonstrated that our assays are capable of accurately quantifying intrastrand cross-links in CP-treated samples and can be utilized to better understand the activity of CP.


Assuntos
Cisplatino , Adutos de DNA , Cisplatino/farmacologia , DNA/química , Cromatografia Líquida , Espectrometria de Massas , Reparo do DNA , Reagentes de Ligações Cruzadas/química
10.
Cureus ; 15(2): e34907, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36938295

RESUMO

The prevalence of overweight and obesity in Jamaica has been steadily increasing over the past decade and is now a significant health issue. This paper focuses on the trends in the prevalence of overweight and obesity in Jamaica from 2000 to 2016. Overweight and obesity prevalence in adults increased from 43.8% in 2000 to 55.5% in 2016, from 34.2% in 2000 to 47.4% in 2016in adult males, and from 53.0% in 2000 to 63.6% in 2016 in adult females. In children/adolescents aged 10 to 19 years, the prevalence of obesity has doubled between 2000 and 2016. The data shows that the prevalence of overweight and obesity in children/adolescents increased from 5% in 2000 to 11.4% in 2016, from 4.4% in 2000 to 11.0% in 2016 in boys, and from 5.5% in 2000 to 11.9% in 2016 in girls.

11.
Proc Natl Acad Sci U S A ; 120(11): e2208860120, 2023 03 14.
Artigo em Inglês | MEDLINE | ID: mdl-36893274

RESUMO

XPA is a central scaffold protein that coordinates the assembly of repair complexes in the global genome (GG-NER) and transcription-coupled nucleotide excision repair (TC-NER) subpathways. Inactivating mutations in XPA cause xeroderma pigmentosum (XP), which is characterized by extreme UV sensitivity and a highly elevated skin cancer risk. Here, we describe two Dutch siblings in their late forties carrying a homozygous H244R substitution in the C-terminus of XPA. They present with mild cutaneous manifestations of XP without skin cancer but suffer from marked neurological features, including cerebellar ataxia. We show that the mutant XPA protein has a severely weakened interaction with the transcription factor IIH (TFIIH) complex leading to an impaired association of the mutant XPA and the downstream endonuclease ERCC1-XPF with NER complexes. Despite these defects, the patient-derived fibroblasts and reconstituted knockout cells carrying the XPA-H244R substitution show intermediate UV sensitivity and considerable levels of residual GG-NER (~50%), in line with the intrinsic properties and activities of the purified protein. By contrast, XPA-H244R cells are exquisitely sensitive to transcription-blocking DNA damage, show no detectable recovery of transcription after UV irradiation, and display a severe deficiency in TC-NER-associated unscheduled DNA synthesis. Our characterization of a new case of XPA deficiency that interferes with TFIIH binding and primarily affects the transcription-coupled subpathway of nucleotide excision repair, provides an explanation of the dominant neurological features in these patients, and reveals a specific role for the C-terminus of XPA in TC-NER.


Assuntos
Neoplasias Cutâneas , Xeroderma Pigmentoso , Humanos , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Alelos , Proteína de Xeroderma Pigmentoso Grupo A/genética , Proteína de Xeroderma Pigmentoso Grupo A/metabolismo , Reparo do DNA/genética , Dano ao DNA/genética , Xeroderma Pigmentoso/genética , Xeroderma Pigmentoso/metabolismo , Neoplasias Cutâneas/genética , Fator de Transcrição TFIIH/genética , Fator de Transcrição TFIIH/metabolismo
12.
ACS Infect Dis ; 9(3): 423-449, 2023 03 10.
Artigo em Inglês | MEDLINE | ID: mdl-36795604

RESUMO

Leishmaniasis is an infectious disease responsible for a huge rate of morbidity and mortality in humans. Chemotherapy consists of the use of pentavalent antimonial, amphotericin B, pentamidine, miltefosine, and paromomycin. However, these drugs are associated with some drawbacks such as high toxicity, administration by parenteral route, and most seriously the resistance of some strains of the parasite to them. Several strategies have been used to increase the therapeutic index and reduce the toxic effects of these drugs. Among them, the use of nanosystems that have great potential as a site-specific drug delivery system stands out. This review aims to compile results from studies that were carried out using first- and second-line antileishmanial drug-carrying nanosystems. The articles referred to here were published between 2011 and 2021. This study shows the promise of effective applicability of drug-carrying nanosystems in the field of antileishmanial therapeutics, with the perspective of providing better patient adherence to treatment, increased therapeutic efficacy, reduced toxicity of conventional drugs, as well as the potential to efficiently improve the treatment of leishmaniasis.


Assuntos
Antiprotozoários , Leishmaniose , Humanos , Preparações Farmacêuticas , Antiprotozoários/farmacologia , Antiprotozoários/uso terapêutico , Leishmaniose/tratamento farmacológico , Pentamidina/farmacologia , Pentamidina/uso terapêutico , Paromomicina/farmacologia
13.
NAR Cancer ; 5(1): zcac044, 2023 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-36683914

RESUMO

Unrepaired oxidatively-stressed replication forks can lead to chromosomal instability and neoplastic transformation or cell death. To meet these challenges cells have evolved a robust mechanism to repair oxidative genomic DNA damage through the base excision repair (BER) pathway, but less is known about repair of oxidative damage at replication forks. We found that depletion or genetic deletion of EEPD1 decreases clonogenic cell survival after oxidative DNA damage. We demonstrate that EEPD1 is recruited to replication forks stressed by oxidative damage induced by H2O2 and that EEPD1 promotes replication fork repair and restart and decreases chromosomal abnormalities after such damage. EEPD1 binds to abasic DNA structures and promotes resolution of genomic abasic sites after oxidative stress. We further observed that restoration of expression of EEPD1 via expression vector transfection restores cell survival and suppresses chromosomal abnormalities induced by oxidative stress in EEPD1-depleted cells. Consistent with this, we found that EEPD1 preserves replication fork integrity by preventing oxidatively-stressed unrepaired fork fusion, thereby decreasing chromosome instability and mitotic abnormalities. Our results indicate a novel role for EEPD1 in replication fork preservation and maintenance of chromosomal stability during oxidative stress.

14.
Nucleic Acids Res ; 51(2): 631-649, 2023 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-36594163

RESUMO

TRAIP is a key factor involved in the DNA damage response (DDR), homologous recombination (HR) and DNA interstrand crosslink (ICL) repair. However, the exact functions of TRAIP in these processes in mammalian cells are not fully understood. Here we identify the zinc finger protein 212, ZNF212, as a novel binding partner for TRAIP and find that ZNF212 colocalizes with sites of DNA damage. The recruitment of TRAIP or ZNF212 to sites of DNA damage is mutually interdependent. We show that depletion of ZNF212 causes defects in the DDR and HR-mediated repair in a manner epistatic to TRAIP. In addition, an epistatic analysis of Zfp212, the mouse homolog of human ZNF212, in mouse embryonic stem cells (mESCs), shows that it appears to act upstream of both the Neil3 and Fanconi anemia (FA) pathways of ICLs repair. We find that human ZNF212 interacted directly with NEIL3 and promotes its recruitment to ICL lesions. Collectively, our findings identify ZNF212 as a new factor involved in the DDR, HR-mediated repair and ICL repair though direct interaction with TRAIP.


Assuntos
Reparo do DNA , Anemia de Fanconi , Animais , Camundongos , Humanos , Reparo do DNA/genética , Dano ao DNA , Replicação do DNA , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Genômica , Anemia de Fanconi/genética , Mamíferos/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Proteínas do Tecido Nervoso/genética
15.
Mil Med ; 188(1-2): e198-e204, 2023 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-34027970

RESUMO

INTRODUCTION: Following suspected sonic attacks on U.S. Embassies, a subset of individuals presented with a unique cluster of symptoms believed to have resulted from exposure to directed energy. Directed energy has been described as exposure to a unique sound/pressure phenomenon such as infrasonic or ultrasonic acoustic or electromagnetic energy. The Joint Force does not have an established protocol to guide vestibular physical therapy for individuals exposed to directed energy. Therefore, we have provided evidence-based guidance for the treatment of oculomotor- and vestibular-related impairments from similar populations. MATERIALS AND METHODS: Published evidence was used to inform suggestions for clinical best practice. We offer resources for the management of non-oculomotor- and non-vestibular-related impairments, before discussing physical therapy interventions for dizziness and imbalance. RESULTS: The physical therapist should design a treatment program that addresses the individual's health condition(s), body structure and function impairments, activity limitations, and participation restrictions after suspected directed energy exposure. This treatment program may include static standing, compliant surface standing, weight shifting, modified center of gravity, gait, and gaze stabilization or vestibular-ocular reflex training. Habituation may also be prescribed. Interventions were selected that require little to no specialized equipment, as such equipment may not be available in all settings (i.e., operational environments). CONCLUSIONS: Evidence-based guidance for prescribing a comprehensive vestibular physical therapy regimen for individuals exposed to directed energy may aid in their rehabilitation and return to duty. This standardized approach can help physical therapists to treat complaints that do not match any previously known medical conditions but resemble brain injury or vestibular pathology.


Assuntos
Lesões Encefálicas , Doenças Vestibulares , Humanos , Doenças Vestibulares/terapia , Tontura , Modalidades de Fisioterapia , Vertigem
16.
Proc Natl Acad Sci U S A ; 119(34): e2207408119, 2022 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-35969784

RESUMO

The xeroderma pigmentosum protein A (XPA) and replication protein A (RPA) proteins fulfill essential roles in the assembly of the preincision complex in the nucleotide excision repair (NER) pathway. We have previously characterized the two interaction sites, one between the XPA N-terminal (XPA-N) disordered domain and the RPA32 C-terminal domain (RPA32C), and the other with the XPA DNA binding domain (DBD) and the RPA70AB DBDs. Here, we show that XPA mutations that inhibit the physical interaction in either site reduce NER activity in biochemical and cellular systems. Combining mutations in the two sites leads to an additive inhibition of NER, implying that they fulfill distinct roles. Our data suggest a model in which the interaction between XPA-N and RPA32C is important for the initial association of XPA with NER complexes, while the interaction between XPA DBD and RPA70AB is needed for structural organization of the complex to license the dual incision reaction. Integrative structural models of complexes of XPA and RPA bound to single-stranded/double-stranded DNA (ss/dsDNA) junction substrates that mimic the NER bubble reveal key features of the architecture of XPA and RPA in the preincision complex. Most critical among these is that the shape of the NER bubble is far from colinear as depicted in current models, but rather the two strands of unwound DNA must assume a U-shape with the two ss/dsDNA junctions localized in close proximity. Our data suggest that the interaction between XPA and RPA70 is key for the organization of the NER preincision complex.


Assuntos
Reparo do DNA , Proteína de Replicação A , Proteína de Xeroderma Pigmentoso Grupo A , DNA/metabolismo , Dano ao DNA , Ligação Proteica , Domínios Proteicos , Proteína de Replicação A/genética , Proteína de Replicação A/metabolismo , Proteína de Xeroderma Pigmentoso Grupo A/genética , Proteína de Xeroderma Pigmentoso Grupo A/metabolismo
17.
Nat Commun ; 13(1): 4762, 2022 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-35963869

RESUMO

Cells employ global genome nucleotide excision repair (GGR) to eliminate a broad spectrum of DNA lesions, including those induced by UV light. The lesion-recognition factor XPC initiates repair of helix-destabilizing DNA lesions, but binds poorly to lesions such as CPDs that do not destabilize DNA. How difficult-to-repair lesions are detected in chromatin is unknown. Here, we identify the poly-(ADP-ribose) polymerases PARP1 and PARP2 as constitutive interactors of XPC. Their interaction results in the XPC-stimulated synthesis of poly-(ADP-ribose) (PAR) by PARP1 at UV lesions, which in turn enables the recruitment and activation of the PAR-regulated chromatin remodeler ALC1. PARP2, on the other hand, modulates the retention of ALC1 at DNA damage sites. Notably, ALC1 mediates chromatin expansion at UV-induced DNA lesions, leading to the timely clearing of CPD lesions. Thus, we reveal how chromatin containing difficult-to-repair DNA lesions is primed for repair, providing insight into mechanisms of chromatin plasticity during GGR.


Assuntos
Cromatina , Inibidores de Poli(ADP-Ribose) Polimerases , Cromatina/genética , DNA/genética , DNA/metabolismo , Dano ao DNA , Reparo do DNA , Proteínas de Ligação a DNA/metabolismo , Poli Adenosina Difosfato Ribose/metabolismo
18.
ACS Chem Biol ; 17(7): 1672-1676, 2022 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-35700389

RESUMO

2,6-Diaminopurine (Z) is a naturally occurring adenine (A) analog that bacteriophages employ in place of A in their genetic alphabet. Recent discoveries of biogenesis pathways of Z in bacteriophages have stimulated substantial research interest in this DNA modification. Here, we systematically examined the effects of Z on the efficiency and fidelity of DNA transcription. Our results showed that Z exhibited no mutagenic yet substantial inhibitory effects on transcription mediated by purified T7 RNA polymerase and by human RNA polymerase II in HeLa nuclear extracts and in human cells. A structurally related adenine analog, 2-aminopurine (2AP), strongly blocked T7 RNA polymerase but did not impede human RNA polymerase II in vitro or in human cells, where no mutant transcript could be detected. The lack of mutagenic consequence and the presence of a strong blockage effect of Z on transcription suggest a role of Z in transcriptional regulation. Z is also subjected to removal by transcription-coupled nucleotide-excision repair (TC-NER), but not global-genome NER in human cells. Our findings provide new insight into the effects of Z on transcription and its potential biological functions.


Assuntos
2-Aminopurina , RNA Polimerase II , 2-Aminopurina/análogos & derivados , 2-Aminopurina/farmacologia , DNA , Reparo do DNA , Humanos , RNA Polimerase II/metabolismo , Transcrição Gênica
19.
Mol Cell ; 82(7): 1343-1358.e8, 2022 04 07.
Artigo em Inglês | MEDLINE | ID: mdl-35271816

RESUMO

Nucleotide excision repair (NER) counteracts the onset of cancer and aging by removing helix-distorting DNA lesions via a "cut-and-patch"-type reaction. The regulatory mechanisms that drive NER through its successive damage recognition, verification, incision, and gap restoration reaction steps remain elusive. Here, we show that the RAD5-related translocase HLTF facilitates repair through active eviction of incised damaged DNA together with associated repair proteins. Our data show a dual-incision-dependent recruitment of HLTF to the NER incision complex, which is mediated by HLTF's HIRAN domain that binds 3'-OH single-stranded DNA ends. HLTF's translocase motor subsequently promotes the dissociation of the stably damage-bound incision complex together with the incised oligonucleotide, allowing for an efficient PCNA loading and initiation of repair synthesis. Our findings uncover HLTF as an important NER factor that actively evicts DNA damage, thereby providing additional quality control by coordinating the transition between the excision and DNA synthesis steps to safeguard genome integrity.


Assuntos
Reparo do DNA , Proteínas de Ligação a DNA , DNA/genética , DNA/metabolismo , Dano ao DNA , Replicação do DNA , Proteínas de Ligação a DNA/genética
20.
Knee Surg Sports Traumatol Arthrosc ; 30(6): 1915-1926, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35258647

RESUMO

PURPOSE: Orthopedic literature remains divided on the utility of biologic augmentation to optimize outcomes after isolated meniscal repair. The aim of this systematic review is to analyze the clinical outcomes and re-operation rates of biologically augmented meniscal repairs. METHODS: PubMed, CINAHL, Cochrane, and EMBASE databases were queried in October 2020 for published literature on isolated meniscal repair with biological augmentation. Studies were assessed for quality and risk of bias by two appraisal tools. Patient demographics, meniscal tear characteristics, surgical procedure, augmentation type, post-operative rehabilitation, patient reported outcome measures, and length of follow-up were recorded, reviewed, and analyzed by two independent reviewers. RESULTS: Of 3794 articles, 18 met inclusion criteria and yielded 537 patients who underwent biologic augmentation of meniscal repair. The biologically augmented repair rates were 5.8-27.0% with PRP augmentation, 0.0-28.5% with fibrin clot augmentation, 0.0-12.9% with marrow stimulation, and 0.0% with stem cell augmentation. One of seven studies showed lower revision rates with augmented meniscal repair compared to standard repair techniques, whereas five of seven found no benefit. Three of ten studies found significant functional improvement of biologically augmented repair versus standard repair techniques and six of ten studies found no difference. There was significant heterogeneity in methods for biologic preparation, delivery, and post-operative rehabilitation protocols. CONCLUSION: Patients reported significant improvements in functional outcomes scores after repair with biological augmentation, though the benefit over standard repair controls is questionable. Revision rates after biologically augmented meniscal repair also appear similar to standard repair techniques. Clinicians should bear this in mind when considering biologic augmentation in the setting of meniscal repair. LEVEL OF EVIDENCE: IV.


Assuntos
Produtos Biológicos , Traumatismos do Joelho , Lesões do Menisco Tibial , Artroscopia/métodos , Humanos , Traumatismos do Joelho/cirurgia , Meniscos Tibiais/cirurgia , Lesões do Menisco Tibial/cirurgia
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