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1.
Cell Mol Biol (Noisy-le-grand) ; 70(8): 104-109, 2024 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-39262256

RESUMO

This study investigated the role of Poly (ADP-ribose) Polymerase (PARP) in myocardial ischemia-reperfusion injury (MIRI) in elderly mice. It involves 30 elderly male KM mice divided into three groups: Sham, MIRI, and DPQ, where the MIRI and DPQ groups undergo myocardial ischemia-reperfusion with the DPQ group also receiving DPQ for PARP-1 inhibition. Over three weeks, assessments include histological analysis of myocardial lesions, left ventricular ejection fraction (LVEF) measurements, and evaluations of serum cardiac enzymes and inflammatory markers. This approach aims to understand the protective effects of DPQ in MIRI, focusing on its impact on cardiac health and inflammation via the JAK2/STAT3 pathway. The findings suggest that PARP activation exacerbates cardiac dysfunction and inflammation in MIRI by possibly modulating the JAK2/STAT3 signaling pathway. Inhibition of PARP-1 with DPQ mitigates these effects, as indicated by reduced myocardial lesions and inflammatory infiltration, improved LVEF, and altered levels of inflammatory markers and signaling molecules. However, the differences in STAT3 and p-STAT3 protein expression between the DPQ and MIRI groups were not statistically significant, suggesting that while PARP inhibition affects many aspects of MIRI pathology, its impact on the JAK2/STAT3 pathway may not fully explain the observed benefits. This study contributes to our understanding of the complex mechanisms underlying myocardial ischemia-reperfusion injury, particularly in the context of aging. It highlights the potential of PARP inhibition as a therapeutic strategy to attenuate cardiac dysfunction and inflammation in MIRI, though further research is necessary to fully elucidate the underlying molecular pathways and to explore the clinical relevance of these findings in humans.


Assuntos
Janus Quinase 2 , Traumatismo por Reperfusão Miocárdica , Miocárdio , Fator de Transcrição STAT3 , Transdução de Sinais , Animais , Traumatismo por Reperfusão Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/patologia , Masculino , Camundongos , Janus Quinase 2/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Miocárdio/patologia , Miocárdio/metabolismo , Poli(ADP-Ribose) Polimerases/metabolismo , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Envelhecimento , Poli(ADP-Ribose) Polimerase-1/metabolismo , Inflamação/patologia , Inflamação/metabolismo , Função Ventricular Esquerda/efeitos dos fármacos
2.
J Nutr Sci Vitaminol (Tokyo) ; 70(4): 295-304, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39218690

RESUMO

Cellular NAD+ is continuously degraded and synthesized under resting conditions. In mammals, NAD+ synthesis is primarily initiated from nicotinamide (Nam) by Nam phosphoribosyltransferase, whereas poly(ADP-ribose) polymerase 1 (PARP1) and 2 (PARP2), sirtuin1 (SIRT1), CD38, and sterile alpha and TIR motif containing 1 (SARM1) are involved in NAD+ breakdown. Using flux analysis with 2H-labeled Nam, we found that when mammalian cells were cultured in the absence of Nam, cellular NAD+ levels were maintained and NAD+ breakdown was completely suppressed. In the presence of Nam, the rate of NAD+ breakdown (RB) did not significantly change upon PARP1, PARP2, SIRT1, or SARM1 deletion, whereas stable expression of CD38 did not increase RB. However, RB in PARP1-deleted cells was much higher compared with that in wild-type cells, in which PARP1 activity was blocked with a selective inhibitor. In contrast, RB in CD38-overexpressing cells in the presence of a specific CD38 inhibitor was much lower compared with that in control cells. The results indicate that PARP1 deletion upregulates the activity of other NADases, whereas CD38 expression downregulates the activity of endogenous NADases, including PARP1 and PARP2. The rate of cellular NAD+ breakdown and the resulting NAD+ concentration may be maintained at a constant level, despite changes in the NAD+-degrading enzyme expression, through the compensatory regulation of NADase activity.


Assuntos
ADP-Ribosil Ciclase 1 , NAD , Poli(ADP-Ribose) Polimerase-1 , Sirtuína 1 , NAD/metabolismo , ADP-Ribosil Ciclase 1/metabolismo , ADP-Ribosil Ciclase 1/genética , Animais , Poli(ADP-Ribose) Polimerase-1/metabolismo , Sirtuína 1/metabolismo , Sirtuína 1/genética , Niacinamida/farmacologia , Niacinamida/metabolismo , Camundongos , Poli(ADP-Ribose) Polimerases/metabolismo , Humanos , Nicotinamida Fosforribosiltransferase/metabolismo , Nicotinamida Fosforribosiltransferase/genética , Deleção de Genes
3.
Chem Biol Interact ; 401: 111186, 2024 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-39116916

RESUMO

Studies on the molecular mechanisms of heavy metal toxicity in invertebrate reproduction are limited. Given that PARP-catalysed ADP-ribosylation is also involved in counteracting heavy metal toxicity and maintaining genomic integrity, and that PARylation is implicated in chromatin remodelling but its role in sperm chromatin remains to be elucidated, we investigated the effects of chromium(VI) at 1, 10 and 100 nM on the reproductive health of Mytilus galloprovincialis. The damage to the gonads was assessed by morphological analyses and the damage indices PARP and É£H2A.X were measured. Changes in the binding of protamine-like (PL) to DNA and the possibility of poly(ADP-ribosyl)ation of PL proteins were also investigated. Gonadal chromium accumulation and morphological damage were found, especially when the mussels were exposed to the highest dose of chromium(VI). In addition, the maximum expression of gonadal É£H2A.X and PARP were obtained at 100 and 10 nM Cr(VI), respectively. Interestingly, for the first time in all exposed conditions, poly(ADP)-ribosylation was detected on PL-II, which, together with PL-III and PL-IV, are the major nuclear basic proteins of Mytilus galloprovincialis sperm chromatin. Since PL-II is involved in the final high level of sperm chromatin compaction, this post-translational modification altered the binding of the PL protein to DNA, favouring the action of micrococcal nuclease on sperm chromatin. This study provides new insights into the effects of chromium(VI) on Mytilus galloprovincialis reproductive system and proposes a molecular mechanism hypothesis describing the toxic effects of this metal on PL-DNA binding, sperm chromatin and gonads.


Assuntos
Cromo , Mytilus , Protaminas , Animais , Mytilus/efeitos dos fármacos , Mytilus/metabolismo , Masculino , Cromo/toxicidade , Protaminas/metabolismo , Poli ADP Ribosilação/efeitos dos fármacos , Poli(ADP-Ribose) Polimerases/metabolismo , Histonas/metabolismo , Gônadas/efeitos dos fármacos , Gônadas/metabolismo , Espermatozoides/efeitos dos fármacos , Espermatozoides/metabolismo , Reprodução/efeitos dos fármacos , DNA/metabolismo , DNA/efeitos dos fármacos
4.
Biol Pharm Bull ; 47(8): 1437-1446, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39143009

RESUMO

Bisdemethoxycurcumin (BDMC) is one of major forms of curcuminoids found in the rhizomes of turmeric. Docetaxel (DTX) is the standard of care for men diagnosed with androgen-independent prostate cancers. Here we report for the first time that BDMC could reinforce the effect of DTX against prostate cancer in vitro and in vivo. In vitro study, PC3 and LNCaP cells were cultured and treated with BDMC and DTX alone or in combination. The effects on cell viability were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Apoptosis was assessed by annexin V/propidium iodide (PI) staining, while cell cycle was assessed by PI staining. Bax, Bcl-2, caspase, poly(ADP-ribose)polymerase (PARP), cyclin B1 and CDK1 expression were assayed by Western blot. We found that a combination treatment of BDMC (10 µM) with DTX (10 nM) was more effective in the inhibition of PC3 and LNCaP cell growth and induction of apoptosis as well as G2/M arrest, which is accompanied with the significant inhibition of Bcl-2, cyclin B1, CDK1 expression and significant increase of Bax, cleaved caspase-9, cleaved caspase-3 and cleaved PARP, than those by treatment of BDMC or DTX alone. Moreover, in vivo evaluation further demonstrated the superior anticancer efficacy of BDMC and DTX compared to DTX alone in a murine prostate cancer model. These results suggest that BDMC can be an attractive therapeutic candidate in enhancing the efficacy of DTX in prostate cancer treatment.


Assuntos
Antineoplásicos , Apoptose , Diarileptanoides , Docetaxel , Neoplasias da Próstata , Masculino , Diarileptanoides/farmacologia , Diarileptanoides/uso terapêutico , Humanos , Animais , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Neoplasias da Próstata/metabolismo , Docetaxel/farmacologia , Docetaxel/uso terapêutico , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Sinergismo Farmacológico , Ciclina B1/metabolismo , Camundongos Nus , Ensaios Antitumorais Modelo de Xenoenxerto , Camundongos , Curcumina/análogos & derivados , Curcumina/farmacologia , Curcumina/uso terapêutico , Sobrevivência Celular/efeitos dos fármacos , Taxoides/farmacologia , Taxoides/uso terapêutico , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Camundongos Endogâmicos BALB C , Poli(ADP-Ribose) Polimerases/metabolismo , Proteína Quinase CDC2/metabolismo
5.
Biochem J ; 481(17): 1097-1123, 2024 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-39178157

RESUMO

ADP-ribosylation is a prominent and versatile post-translational modification, which regulates a diverse set of cellular processes. Poly-ADP-ribose (PAR) is synthesised by the poly-ADP-ribosyltransferases PARP1, PARP2, tankyrase (TNKS), and tankyrase 2 (TNKS2), all of which are linked to human disease. PARP1/2 inhibitors have entered the clinic to target cancers with deficiencies in DNA damage repair. Conversely, tankyrase inhibitors have continued to face obstacles on their way to clinical use, largely owing to our limited knowledge of their molecular impacts on tankyrase and effector pathways, and linked concerns around their tolerability. Whilst detailed structure-function studies have revealed a comprehensive picture of PARP1/2 regulation, our mechanistic understanding of the tankyrases lags behind, and thereby our appreciation of the molecular consequences of tankyrase inhibition. Despite large differences in their architecture and cellular contexts, recent structure-function work has revealed striking parallels in the regulatory principles that govern these enzymes. This includes low basal activity, activation by intra- or inter-molecular assembly, negative feedback regulation by auto-PARylation, and allosteric communication. Here we compare these poly-ADP-ribosyltransferases and point towards emerging parallels and open questions, whose pursuit will inform future drug development efforts.


Assuntos
Poli(ADP-Ribose) Polimerase-1 , Tanquirases , Tanquirases/metabolismo , Tanquirases/antagonistas & inibidores , Tanquirases/genética , Tanquirases/química , Humanos , Poli(ADP-Ribose) Polimerase-1/metabolismo , Poli(ADP-Ribose) Polimerase-1/antagonistas & inibidores , Poli(ADP-Ribose) Polimerase-1/genética , Poli(ADP-Ribose) Polimerases/metabolismo , Poli(ADP-Ribose) Polimerases/química , Poli(ADP-Ribose) Polimerases/genética , Animais , Processamento de Proteína Pós-Traducional , Reparo do DNA , ADP-Ribosilação , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Poli ADP Ribosilação/genética
6.
Nutrients ; 16(16)2024 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-39203802

RESUMO

Dendritic cells (DCs) are crucial in initiating and shaping both innate and adaptive immune responses. Clinical studies and experimental models have highlighted their significant involvement in various autoimmune diseases, positioning them as promising therapeutic targets. Nicotinamide (NAM), a form of vitamin B3, with its anti-inflammatory properties, has been suggested, while the involvement of NAM in DCs regulation remains elusive. Here, through analyzing publicly available databases, we observe substantial alterations in NAM levels and NAM metabolic pathways during DCs activation. Furthermore, we discover that NAM, but not Nicotinamide Mononucleotide (NMN), significantly inhibits DCs over-activation in vitro and in vivo. The suppression of DCs hyperactivation effectively alleviates symptoms of psoriasis. Mechanistically, NAM impairs DCs activation through a Poly (ADP-ribose) polymerases (PARPs)-NF-κB dependent manner. Notably, phosphoribosyl transferase (NAMPT) and PARPs are significantly upregulated in lipopolysaccharide (LPS)-stimulated DCs and psoriasis patients; elevated NAMPT and PARPs expression in psoriasis patients correlates with higher psoriasis area and severity index (PASI) scores. In summary, our findings underscore the pivotal role of NAM in modulating DCs functions and autoimmune disorders. Targeting the NAMPT-PARP axis emerges as a promising therapeutic approach for DC-related diseases.


Assuntos
Doenças Autoimunes , Células Dendríticas , Niacinamida , Nicotinamida Fosforribosiltransferase , Poli(ADP-Ribose) Polimerases , Psoríase , Transdução de Sinais , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/metabolismo , Células Dendríticas/imunologia , Niacinamida/farmacologia , Humanos , Transdução de Sinais/efeitos dos fármacos , Animais , Psoríase/tratamento farmacológico , Psoríase/imunologia , Psoríase/metabolismo , Doenças Autoimunes/tratamento farmacológico , Nicotinamida Fosforribosiltransferase/metabolismo , Camundongos , Poli(ADP-Ribose) Polimerases/metabolismo , NF-kappa B/metabolismo , Camundongos Endogâmicos C57BL , Lipopolissacarídeos
7.
Expert Rev Mol Med ; 26: e17, 2024 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-39189367

RESUMO

ADP-ribosyltransferases of the PARP family encompass a group of enzymes with variegated regulatory functions in cells, ranging from DNA damage repair to the control of cell-cycle progression and immune response. Over the years, this knowledge has led to the use of PARP1/2 inhibitors as mainstay pharmaceutical strategies for the treatment of ovarian, pancreatic, prostate and breast cancers, holding mutations in genes encoding for proteins involved in the DNA repair mechanisms (synthetic lethality). Meanwhile, the last decade has witnessed significant progress in comprehending cellular pathways regulated by mono-ADP-ribosylation, with a huge effort in the development of novel selective compounds to inhibit those PARPs endowed with mono-ADP-ribosylation activity. This review focuses on the progress achieved in the cancer field, delving into most recent findings regarding the role of a subset of enzymes - the interferon-stimulated PARPs - in cancer progression.


Assuntos
ADP-Ribosilação , Interferons , Neoplasias , Inibidores de Poli(ADP-Ribose) Polimerases , Poli(ADP-Ribose) Polimerases , Transdução de Sinais , Humanos , Neoplasias/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Transdução de Sinais/efeitos dos fármacos , Interferons/metabolismo , Poli(ADP-Ribose) Polimerases/metabolismo , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Animais , Reparo do DNA
8.
Int J Mol Sci ; 25(16)2024 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-39201718

RESUMO

Poly (ADP-Ribose) Polymerase (PARP) inhibitors have changed the outcomes and therapeutic strategy for several cancer types. As a targeted therapeutic mainly for patients with BRCA1/2 mutations, PARP inhibitors have commonly been exploited for their capacity to prevent DNA repair. In this review, we discuss the multifaceted roles of PARP-1 and PARP-2 beyond DNA repair, including the impact of PARP-1 on chemokine signalling, immune modulation, and transcriptional regulation of gene expression, particularly in the contexts of angiogenesis and epithelial-to-mesenchymal transition (EMT). We evaluate the pre-clinical role of PARP inhibitors, either as single-agent or combination therapies, to block the metastatic process. Efficacy of PARP inhibitors was demonstrated via DNA repair-dependent and independent mechanisms, including DNA damage, cell migration, invasion, initial colonization at the metastatic site, osteoclastogenesis, and micrometastasis formation. Finally, we summarize the recent clinical advancements of PARP inhibitors in the prevention and progression of distant metastases, with a particular focus on specific metastatic sites and PARP-1 selective inhibitors. Overall, PARP inhibitors have demonstrated great potential in inhibiting the metastatic process, pointing the way for greater use in early cancer settings.


Assuntos
Metástase Neoplásica , Neoplasias , Poli(ADP-Ribose) Polimerase-1 , Inibidores de Poli(ADP-Ribose) Polimerases , Poli(ADP-Ribose) Polimerases , Humanos , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Poli(ADP-Ribose) Polimerase-1/metabolismo , Poli(ADP-Ribose) Polimerase-1/antagonistas & inibidores , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Neoplasias/metabolismo , Neoplasias/genética , Poli(ADP-Ribose) Polimerases/metabolismo , Animais , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos
9.
Biomol Biomed ; 24(5): 1199-1215, 2024 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-39213416

RESUMO

Cardiac arrhythmias are often linked to the overactivity of cardiac fibroblasts (CFs). Investigating the impact of poly (ADP-ribose) polymerase 9 (PARP9) on Angiotensin II (Ang II)-induced fibroblast activation and the therapeutic effects of pirfenidone (PFD) offers valuable insights into cardiac arrhythmias. This study utilized weighted gene co-expression network analysis (WGCNA), differential gene expression (DEG) analysis, protein-protein interaction (PPI), and receiver operating characteristic (ROC) analysis on the GSE42955 dataset to identify the hub gene with a significant diagnostic value. The ImmuCellAI tool revealed an association between PARP9 and immune cell infiltration. Our in vitro assessments focused on the influence of PFD on myofibroblast differentiation, transforming growth factor-beta (TGF-ß) expression, and Ang II-induced proliferation and migration in CFs. Additionally, we explored the impact on fibrosis markers and the TGF-ß/Smad signaling pathway in the context of PARP9 overexpression. Analysis of the GSE42955 dataset revealed PARP9 as a central gene with high clinical diagnostic value, linked to seven types of immune cells. The in vitro studies demonstrated that PFD significantly mitigates Ang II-induced CF proliferation, migration, and fibrosis. It also reduces Ang II-induced PARP9 expression and decreases fibrosis markers, including TGF-ß, collagen I, collagen III, and α-SMA. Notably, PARP9 overexpression can partially counteract PFD's inhibitory effects on CFs and modify the expression of fibronectin, CTGF, α-SMA, collagen I, collagen III, MMP2, MMP9, TGF-ß, and p-Smad2/3 in the TGF-ß/Smad signaling pathway. In summary, our findings suggest that PFD effectively counteracts the adverse effects of Ang II-induced CF proliferation and fibrosis, and modulates the TGF-ß/Smad signaling pathway and PARP9 expression. This identifies a potential therapeutic approach for managing myocardial fibrosis.


Assuntos
Angiotensina II , Miocárdio , Piridonas , Transdução de Sinais , Proteínas Smad , Fator de Crescimento Transformador beta , Piridonas/farmacologia , Piridonas/uso terapêutico , Fator de Crescimento Transformador beta/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/metabolismo , Miocárdio/metabolismo , Miocárdio/patologia , Angiotensina II/farmacologia , Angiotensina II/metabolismo , Humanos , Poli(ADP-Ribose) Polimerases/metabolismo , Poli(ADP-Ribose) Polimerases/genética , Fibrose/metabolismo , Animais , Proliferação de Células/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Movimento Celular/efeitos dos fármacos , Miofibroblastos/efeitos dos fármacos , Miofibroblastos/metabolismo , Miofibroblastos/patologia , Diferenciação Celular/efeitos dos fármacos , Ratos
10.
Protein Expr Purif ; 224: 106580, 2024 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-39154924

RESUMO

Poly-ADP-ribose polymerase-14 (PARP14) can modify proteins and nucleic acids by the reversible addition of a single ADP-ribose molecule. Aberrant PARP14 functions have been related to cancer and inflammation, and its domains are involved in processes related to viral infection. Previous research indicates that PARP14 functions might be mediated via a multitude of target proteins. In vitro studies of this large multidomain enzyme have been complicated by difficulties to obtain biochemical quantities of pure protein. Here we present a strategy that allows bacterial expression and purification of a functional multidomain construct of PARP14. We substituted an internal KH domain and its neighboring unstructured region with a SUMO domain to obtain a protein construct that encompasses three macrodomains, a WWE domain, and a PARP catalytic domain. We show that the resulting construct retains both ADP-ribosyltransferase and de-MARylase activities. This construct will be useful in structural and functional studies of PARP14.


Assuntos
Escherichia coli , Poli(ADP-Ribose) Polimerases , Poli(ADP-Ribose) Polimerases/química , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Humanos , Domínios Proteicos , Proteínas Recombinantes/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/biossíntese , Expressão Gênica , Clonagem Molecular
11.
G3 (Bethesda) ; 14(9)2024 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-39056235

RESUMO

Poly(ADP-ribose) polymerase 1 (PARP1) is a highly conserved nuclear protein in multicellular organisms that by modulating chromatin opening facilitates gene expression during development. All reported Parp1 null knockout mouse strains are viable with no developmental anomalies. It was believed that functional redundancy with other PARP family members, mainly PARP2, explains such a controversy. However, while PARP2 has similar catalytic domain to PARP1, it lacks other domains, making the absence of developmental problems in Parp1 mice knockouts unlikely. Contrary to prior assumptions, in our analysis of the best-investigated Parp1 knockout mouse strain, we identified persistent mRNA expression, albeit at reduced levels. Transcript analysis revealed an alternatively spliced Parp1 variant lacking exon 2. Subsequent protein analysis confirmed the existence of a truncated PARP1 protein in knockout mice. The decreased level of poly(ADP-ribose) (pADPr) was detected in Parp1 knockout embryonic stem (ES) cells with western blotting analysis, but immunofluorescence staining did not detect any difference in distribution or level of pADPr in nuclei of knockout ES cells. pADPr level in double Parp1 Parg mutant ES cells greatly exceeded its amount in normal and even in hypomorph Parg mutant ES cells, suggesting the presence of functionally active PARP1. Therefore, our findings challenge the conventional understanding of PARP1 depletion effects.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Poli(ADP-Ribose) Polimerase-1 , Animais , Camundongos , Processamento Alternativo , Células-Tronco Embrionárias/metabolismo , Loci Gênicos , Camundongos Knockout , Poli(ADP-Ribose) Polimerase-1/genética , Poli(ADP-Ribose) Polimerase-1/metabolismo , Poli Adenosina Difosfato Ribose/metabolismo , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo
12.
Nat Commun ; 15(1): 5822, 2024 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-38987289

RESUMO

DNA polymerase theta (Polθ)-mediated end-joining (TMEJ) repairs DNA double-strand breaks and confers resistance to genotoxic agents. How Polθ is regulated at the molecular level to exert TMEJ remains poorly characterized. We find that Polθ interacts with and is PARylated by PARP1 in a HPF1-independent manner. PARP1 recruits Polθ to the vicinity of DNA damage via PARylation dependent liquid demixing, however, PARylated Polθ cannot perform TMEJ due to its inability to bind DNA. PARG-mediated de-PARylation of Polθ reactivates its DNA binding and end-joining activities. Consistent with this, PARG is essential for TMEJ and the temporal recruitment of PARG to DNA damage corresponds with TMEJ activation and dissipation of PARP1 and PAR. In conclusion, we show a two-step spatiotemporal mechanism of TMEJ regulation. First, PARP1 PARylates Polθ and facilitates its recruitment to DNA damage sites in an inactivated state. PARG subsequently activates TMEJ by removing repressive PAR marks on Polθ.


Assuntos
Quebras de DNA de Cadeia Dupla , Reparo do DNA por Junção de Extremidades , DNA Polimerase teta , DNA Polimerase Dirigida por DNA , Poli(ADP-Ribose) Polimerase-1 , Humanos , Poli(ADP-Ribose) Polimerase-1/metabolismo , Poli(ADP-Ribose) Polimerase-1/genética , DNA Polimerase Dirigida por DNA/metabolismo , Poli Adenosina Difosfato Ribose/metabolismo , Dano ao DNA , Animais , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , DNA/metabolismo , DNA/genética , Células HEK293 , Poli ADP Ribosilação , Poli(ADP-Ribose) Polimerases/metabolismo , Poli(ADP-Ribose) Polimerases/genética , Proteínas de Transporte , Glicosídeo Hidrolases , Proteínas Nucleares
14.
FASEB J ; 38(13): e23775, 2024 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-38967223

RESUMO

Inflammatory bowel disease (IBD) is a chronic disease of the gastrointestinal tract affecting millions of people. Here, we investigated the expression and functions of poly(ADP-ribose) polymerase 14 (Parp14), an important regulatory protein in immune cells, with an IBD patient cohort as well as two mouse colitis models, that is, IBD-mimicking oral dextran sulfate sodium (DSS) exposure and oral Salmonella infection. Parp14 was expressed in the human colon by cells in the lamina propria, but, in particular, by the epithelial cells with a granular staining pattern in the cytosol. The same expression pattern was evidenced in both mouse models. Parp14-deficiency caused increased rectal bleeding as well as stronger epithelial erosion, Goblet cell loss, and immune cell infiltration in DSS-exposed mice. The absence of Parp14 did not affect the mouse colon bacterial microbiota. Also, the colon leukocyte populations of Parp14-deficient mice were normal. In contrast, bulk tissue RNA-Seq demonstrated that the colon transcriptomes of Parp14-deficient mice were dominated by abnormalities in inflammation and infection responses both prior and after the DSS exposure. Overall, the data indicate that Parp14 has an important role in the maintenance of colon epithelial barrier integrity. The prognostic and predictive biomarker potential of Parp14 in IBD merits further investigation.


Assuntos
Colite , Sulfato de Dextrana , Camundongos Endogâmicos C57BL , Poli(ADP-Ribose) Polimerases , Animais , Feminino , Humanos , Masculino , Camundongos , Colite/genética , Colite/induzido quimicamente , Colite/patologia , Colo/patologia , Colo/metabolismo , Sulfato de Dextrana/toxicidade , Modelos Animais de Doenças , Microbioma Gastrointestinal , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/patologia , Doenças Inflamatórias Intestinais/metabolismo , Camundongos Knockout , Poli(ADP-Ribose) Polimerases/metabolismo , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/deficiência
15.
Subcell Biochem ; 104: 531-548, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38963499

RESUMO

Vault ribonucleoprotein particles are naturally designed nanocages, widely found in the eukaryotic kingdom. Vaults consist of 78 copies of the major vault protein (MVP) that are organized in 2 symmetrical cup-shaped halves, of an approximate size of 70x40x40 nm, leaving a huge internal cavity which accommodates the vault poly(ADP-ribose) polymerase (vPARP), the telomerase-associated protein-1 (TEP1) and some small untranslated RNAs. Diverse hypotheses have been developed on possible functions of vaults, based on their unique capsular structure, their rapid movements and the distinct subcellular localization of the particles, implicating transport of cargo, but they are all pending confirmation. Vault particles also possess many attributes that can be exploited in nanobiotechnology, particularly in the creation of vehicles for the delivery of multiple molecular cargoes. Here we review what is known about the structure and dynamics of the vault complex and discuss a possible mechanism for the vault opening process. The recent findings in the characterization of the vaults in cells and in its natural microenvironment will be also discussed.


Assuntos
Partículas de Ribonucleoproteínas em Forma de Abóbada , Partículas de Ribonucleoproteínas em Forma de Abóbada/metabolismo , Partículas de Ribonucleoproteínas em Forma de Abóbada/química , Partículas de Ribonucleoproteínas em Forma de Abóbada/genética , Humanos , Animais , Poli(ADP-Ribose) Polimerases/metabolismo , Poli(ADP-Ribose) Polimerases/química
16.
Nat Commun ; 15(1): 6197, 2024 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-39043663

RESUMO

Replication stress compromises genomic integrity. Fork blocking lesions such as those induced by cisplatin and other chemotherapeutic agents arrest replication forks. Repriming downstream of these lesions represents an important mechanism of replication restart, however the single stranded DNA (ssDNA) gaps left behind, unless efficiently filled, can serve as entry point for nucleases. Nascent strand gaps can be repaired by BRCA-mediated homology repair. Alternatively, gaps can also be filled by translesion synthesis (TLS) polymerases. How these events are regulated is still not clear. Here, we show that PARP10, a poorly-characterized mono-ADP-ribosyltransferase, is recruited to nascent strand gaps to promote their repair. PARP10 interacts with the ubiquitin ligase RAD18 and recruits it to these structures, resulting in the ubiquitination of the replication factor PCNA. PCNA ubiquitination, in turn, recruits the TLS polymerase REV1 for gap filling. We show that PARP10 recruitment to gaps and the subsequent REV1-mediated gap filling requires both the catalytic activity of PARP10, and its ability to interact with PCNA. We moreover show that PARP10 is hyperactive in BRCA-deficient cells, and its inactivation potentiates gap accumulations and cytotoxicity in these cells. Our work uncovers PARP10 as a regulator of ssDNA gap filling, which promotes genomic stability in BRCA-deficient cells.


Assuntos
Reparo do DNA , Replicação do DNA , DNA de Cadeia Simples , Proteínas de Ligação a DNA , Poli(ADP-Ribose) Polimerases , Antígeno Nuclear de Célula em Proliferação , Ubiquitina-Proteína Ligases , Ubiquitinação , Humanos , Antígeno Nuclear de Célula em Proliferação/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , DNA de Cadeia Simples/metabolismo , DNA de Cadeia Simples/genética , Poli(ADP-Ribose) Polimerases/metabolismo , Poli(ADP-Ribose) Polimerases/genética , Dano ao DNA , Proteína BRCA2/metabolismo , Proteína BRCA2/genética , Nucleotidiltransferases/metabolismo , Nucleotidiltransferases/genética , Proteína BRCA1/metabolismo , Proteína BRCA1/genética , Linhagem Celular Tumoral , Células HEK293 , Síntese de DNA Translesão , DNA Polimerase Dirigida por DNA , Proteínas Proto-Oncogênicas
17.
Cell Rep Med ; 5(7): 101649, 2024 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-39019005

RESUMO

Tumor-infiltrating regulatory T cells (TI-Tregs) elicit immunosuppressive effects in the tumor microenvironment (TME) leading to accelerated tumor growth and resistance to immunotherapies against solid tumors. Here, we demonstrate that poly-(ADP-ribose)-polymerase-11 (PARP11) is an essential regulator of immunosuppressive activities of TI-Tregs. Expression of PARP11 correlates with TI-Treg cell numbers and poor responses to immune checkpoint blockade (ICB) in human patients with cancer. Tumor-derived factors including adenosine and prostaglandin E2 induce PARP11 in TI-Tregs. Knockout of PARP11 in the cells of the TME or treatment of tumor-bearing mice with selective PARP11 inhibitor ITK7 inactivates TI-Tregs and reinvigorates anti-tumor immune responses. Accordingly, ITK7 decelerates tumor growth and significantly increases the efficacy of anti-tumor immunotherapies including ICB and adoptive transfer of chimeric antigen receptor (CAR) T cells. These results characterize PARP11 as a key driver of TI-Treg activities and a major regulator of immunosuppressive TME and argue for targeting PARP11 to augment anti-cancer immunotherapies.


Assuntos
Imunoterapia , Poli(ADP-Ribose) Polimerases , Linfócitos T Reguladores , Microambiente Tumoral , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Animais , Humanos , Camundongos , Microambiente Tumoral/imunologia , Microambiente Tumoral/efeitos dos fármacos , Imunoterapia/métodos , Poli(ADP-Ribose) Polimerases/metabolismo , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linhagem Celular Tumoral , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Camundongos Endogâmicos C57BL , Neoplasias/imunologia , Neoplasias/terapia
18.
Biochem Biophys Res Commun ; 731: 150371, 2024 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-39004065

RESUMO

Vascular endothelial cytoskeletal disruption leads to increased vascular permeability and is involved in the pathogenesis and progression of various diseases. Oxidative stress can increase vascular permeability by weakening endothelial cell-to-cell junctions and decrease intracellular nicotinamide adenine dinucleotide (NAD+) levels. However, it remains unclear how intracellular NAD+ variations caused by oxidative stress alter the vascular endothelial cytoskeletal organization. In this study, we demonstrated that oxidative stress activates poly (ADP-ribose [ADPr]) polymerase (PARP), which consume large amounts of intracellular NAD+, leading to cytoskeletal disruption in vascular endothelial cells. We found that hydrogen peroxide (H2O2) could transiently disrupt the cytoskeleton and reduce intracellular total NAD levels in human umbilical vein endothelial cells (HUVECs). H2O2 stimulation led to rapid increase in ADPr protein levels in HUVECs. Pharmaceutical PARP inhibition counteracted H2O2-induced total NAD depletion and cytoskeletal disruption, suggesting that NAD+ consumption by PARP induced cytoskeletal disruption. Additionally, supplementation with nicotinamide mononucleotide (NMN), the NAD+ precursor, prevented both intracellular total NAD depletion and cytoskeletal disruption induced by H2O2 in HUVECs. Inhibition of the NAD+ salvage pathway by FK866, a nicotinamide phosphoribosyltransferase inhibitor, maintained H2O2-induced cytoskeletal disruption, suggesting that intracellular NAD+ plays a crucial role in recovery from cytoskeletal disruption. Our findings provide further insights into the potential application of PARP inhibition and NMN supplementation for the treatment and prevention of diseases involving vascular hyperpermeability.


Assuntos
Citoesqueleto , Células Endoteliais da Veia Umbilical Humana , Peróxido de Hidrogênio , NAD , Estresse Oxidativo , Poli(ADP-Ribose) Polimerases , Humanos , Citoesqueleto/metabolismo , Citoesqueleto/efeitos dos fármacos , NAD/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Poli(ADP-Ribose) Polimerases/metabolismo , Peróxido de Hidrogênio/farmacologia , Peróxido de Hidrogênio/toxicidade , Peróxido de Hidrogênio/metabolismo , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Células Cultivadas
19.
Eur J Med Chem ; 274: 116535, 2024 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-38838546

RESUMO

Poly (ADP-ribose) polymerase (PARP) is considered an essential component in case of DNA (Deoxyribonucleic acid) damage, response by sensing DNA damage and engaging DNA repair proteins. Those proteins repair the damaged DNA via an aspect of posttranslational modification, known as poly (ADP-Ribosyl)ation (PARylation). Specifically, PARP inhibitors (PARPi) have shown better results when administered alone in a variety of cancer types with BRCA (Breast Cancer gene) mutation. The clinical therapeutic benefits of PARP inhibitors have been diminished by their cytotoxicity, progression of drug resistance, and limitation of indication, regardless of their tremendous clinical effectiveness. A growing number of PARP-1 inhibitors, particularly those associated with BRCA-1/2 mutations, have been identified as potential cancer treatments. Recently, several researchers have identified various promising scaffolds, which have resulted in the resuscitation of the faith in PARP inhibitors as cancer therapies. This review provided a comprehensive update on the anatomy and physiology of the PARP enzyme, the profile of FDA (Food and Drug Administration) and CFDA (China Food and Drug Administration)-approved drugs, and small-molecule inhibitors of PARP, including their synthetic routes, biological evaluation, selectivity, and structure-activity relationship.


Assuntos
Antineoplásicos , Inibidores de Poli(ADP-Ribose) Polimerases , Humanos , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases/química , Inibidores de Poli(ADP-Ribose) Polimerases/síntese química , Relação Estrutura-Atividade , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/síntese química , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Poli(ADP-Ribose) Polimerases/metabolismo , Estrutura Molecular , Animais
20.
Front Cell Infect Microbiol ; 14: 1414135, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38863831

RESUMO

Introduction: Acanthamoeba infection is a serious public health concern, necessitating the development of effective and safe anti-Acanthamoeba chemotherapies. Poly (ADP-ribose) polymerases (PARPs) govern a colossal amount of biological processes, such as DNA damage repair, protein degradation and apoptosis. Multiple PARP-targeted compounds have been approved for cancer treatment. However, repurposing of PARP inhibitors to treat Acanthamoeba is poorly understood. Methods: In the present study, we attempted to fill these knowledge gaps by performing anti-Acanthamoeba efficacy assays, cell biology experiments, bioinformatics, and transcriptomic analyses. Results: Using a homology model of Acanthamoeba poly (ADP-ribose) polymerases (PARPs), molecular docking of approved drugs revealed three potential inhibitory compounds: olaparib, venadaparib and AZ9482. In particular, venadaparib exhibited superior docking scores (-13.71) and favorable predicted binding free energy (-89.28 kcal/mol), followed by AZ9482, which showed a docking score of -13.20 and a binding free energy of -92.13 kcal/mol. Notably, the positively charged cyclopropylamine in venadaparib established a salt bridge (through E535) and a hydrogen bond (via N531) within the binding pocket. For comparison, AZ9482 was well stacked by the surrounding aromatic residues including H625, Y652, Y659 and Y670. In an assessment of trophozoites viability, AZ9482 exhibited a dose-and time-dependent anti-trophozoite effect by suppressing Acanthamoeba PARP activity, unlike olaparib and venadaparib. An Annexin V-fluorescein isothiocyanate/propidium iodide apoptosis assay revealed AZ9482 induced trophozoite necrotic cell death rather than apoptosis. Transcriptomics analyses conducted on Acanthamoeba trophozoites treated with AZ9482 demonstrated an atlas of differentially regulated proteins and genes, and found that AZ9482 rapidly upregulates a multitude of DNA damage repair pathways in trophozoites, and intriguingly downregulates several virulent genes. Analyzing gene expression related to DNA damage repair pathway and the rate of apurinic/apyrimidinic (AP) sites indicated DNA damage efficacy and repair modulation in Acanthamoeba trophozoites following AZ9482 treatment. Discussion: Collectively, these findings highlight AZ9482, as a structurally unique PARP inhibitor, provides a promising prototype for advancing anti-Acanthamoeba drug research.


Assuntos
Simulação de Acoplamento Molecular , Inibidores de Poli(ADP-Ribose) Polimerases , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Humanos , Piperazinas/farmacologia , Ftalazinas/farmacologia , Ftalazinas/química , Reposicionamento de Medicamentos , Poli(ADP-Ribose) Polimerases/metabolismo , Acanthamoeba/efeitos dos fármacos , Biologia Computacional , Apoptose/efeitos dos fármacos , Perfilação da Expressão Gênica , Antiprotozoários/farmacologia , Trofozoítos/efeitos dos fármacos
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