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1.
Sci Rep ; 13(1): 829, 2023 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-36646782

RESUMO

High rates of new cervical cancer cases and deaths occur in low- and middle-income countries yearly, and one reason was found related to limitation of regular cervical cancer screening in local and low-resource settings. HPV has over 150 types, yet certain 14-20 high-risk and 13-14 low-risk types are common, and, thus, most conventional HPV nucleic acid assays, for examples, Cobas 4800 HPV test (Roche Diagnostics, New Jersey, USA) and REBA HPV-ID (Molecules and Diagnostics, Wonju, Republic of Korea) were developed to cover these types. We thereby utilized bioinformatics combined with recent isothermal amplification technique at 35-42 °C to firstly describe multiplex recombinase polymerase amplification assay that is specific to these common 20 high-risk and 14 low-risk types, and also L1 and E6/E7 genes that target different stages of cervical cancer development. Multiplex primer concentrations and reaction incubation conditions were optimized to allow simultaneous two gene detections at limit of detection of 1000 copies (equivalent to 2.01 fg) for L1 and 100 copies (0.0125 fg) for E6/E7, respectively. The assay was validated against urogenital and other pathogens, normal flora, and human control. In 130 real clinical sample tests, the assay demonstrated 100% specificity, 78% diagnostic accuracy, and 75% sensitivity compared with REBA HPV-ID test, and is much more rapid (15-40 min), less expensive (~ 3-4 USD/reaction) and does not require instrumentation (35-42 °C reaction condition so hand holding or tropical temperature is possible). Hence, the developed novel assay provides alternative screening tool for potential local screening. Furthermore, as this assay uses safe chemical reagents, it is safe for users.


Assuntos
Infecções por Papillomavirus , Neoplasias do Colo do Útero , Feminino , Humanos , Neoplasias do Colo do Útero/diagnóstico , Recombinases , Infecções por Papillomavirus/diagnóstico , Detecção Precoce de Câncer , Nucleotidiltransferases , Papillomaviridae/genética , Sensibilidade e Especificidade , DNA Viral/genética
2.
Sci Immunol ; 8(79): eabp9765, 2023 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-36662885

RESUMO

The mechanisms by which innate immune receptors mediate self-nonself discrimination are unclear. In this study, we found species-specific molecular determinants of self-DNA reactivity by cyclic guanosine monophosphate-adenosine monophosphate (GMP-AMP) synthase (cGAS). Human cGAS contained a catalytic domain that was intrinsically self-DNA reactive and stimulated interferon responses in diverse cell types. This reactivity was prevented by an upstream amino (N)-terminal domain. The cGAS proteins from several nonhuman primate species exhibited a similar pattern of self-DNA reactivity in cells, but chimpanzee cGAS was inactive even when its amino-terminal domain was deleted. In contrast, the N terminus of mouse cGAS promoted self-DNA reactivity. When expressed within tumors, only self-DNA-reactive cGAS proteins protected mice from tumor-induced lethality. In vitro studies of DNA- or chromatin-induced cGAS activation did not reveal species-specific activities that correlate with self-DNA reactivity observed in macrophages. Cell biological analysis revealed that self-DNA reactivity by human cGAS, but not mouse cGAS, correlated with localization to mitochondria. We found that epitope tag positions affected self-DNA reactivity in cells and that DNA present in cell lysates undermines the reliability of cGAS biochemical fractionations. These studies reveal species-specific diversity of cGAS functions, even within the primate lineage, and highlight experimental considerations for the study of this innate immune receptor.


Assuntos
DNA , Nucleotídeos Cíclicos , Animais , Camundongos , Humanos , Reprodutibilidade dos Testes , DNA/química , DNA/metabolismo , Nucleotidiltransferases/genética , Nucleotidiltransferases/química , Nucleotidiltransferases/metabolismo , Mamíferos/metabolismo
3.
Nat Struct Mol Biol ; 30(1): 72-80, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36593311

RESUMO

Cyclic GMP-AMP synthase (cGAS) is a pattern recognition receptor critical for the innate immune response to intracellular pathogens, DNA damage, tumorigenesis and senescence. Binding to double-stranded DNA (dsDNA) induces conformational changes in cGAS that activate the enzyme to produce 2'-3' cyclic GMP-AMP (cGAMP), a second messenger that initiates a potent interferon (IFN) response through its receptor, STING. Here, we combined two-state computational design with informatics-guided design to create constitutively active, dsDNA ligand-independent cGAS (CA-cGAS). We identified CA-cGAS mutants with IFN-stimulating activity approaching that of dsDNA-stimulated wild-type cGAS. DNA-independent adoption of the active conformation was directly confirmed by X-ray crystallography. In vivo expression of CA-cGAS in tumor cells resulted in STING-dependent tumor regression, demonstrating that the designed proteins have therapeutically relevant biological activity. Our work provides a general framework for stabilizing active conformations of enzymes and provides CA-cGAS variants that could be useful as genetically encoded adjuvants and tools for understanding inflammatory diseases.


Assuntos
Imunidade Inata , Nucleotidiltransferases , Nucleotidiltransferases/metabolismo , DNA/química
5.
Environ Pollut ; 319: 120988, 2023 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-36596376

RESUMO

Toxic effects of excessive manganese (Mn) from occupational or environmental exposure cause harm to human health. Excessive Mn exposure is intimately associated with neurodegeneration and cognitive dysfunction. Inflammatory responses mediated by microglia are essential contributors to the pathogenesis of Mn-induced neurotoxicity. Inhibition of microglia-mediated inflammation has been shown to alleviate Mn-induced neurotoxicity. Sesamol, derived from sesame, has neuroprotective properties in various disease models, including neurological diseases. Whether sesamol protects against Mn-induced neurological injuries has not been determined. Here, both in vivo and in vitro Mn exposure models were established to address the beneficial effects of sesamol on Mn-induced neurotoxicity. We showed that administration of sesamol mitigated learning and memory deficits of mice treated by Mn. Furthermore, sesamol reduced Mn-induced microglial activation and the expression of proinflammatory mediators (TNF-α, iNOS, and Cxcl10), while exerting a marginal effect on anti-inflammation and microglial phagocytosis. Mn exposure activated the microglial cGAS-STING pathway and sesamol inhibited this pathway by reducing the phosphorylation of STING and NF-κB, concomitantly decreasing IFN-α and IFN-ß synthesis. In summary, our novel results indicated that sesamol exerted its protective effects on Mn-induced neuroinflammation and cognitive impairment via the microglial cGAS-STING/NF-κB pathway, providing evidence that sesamol may serve as an effective therapeutic for preventing and treating Mn-induced neurotoxicity.


Assuntos
Disfunção Cognitiva , NF-kappa B , Animais , Humanos , Camundongos , Disfunção Cognitiva/induzido quimicamente , Disfunção Cognitiva/tratamento farmacológico , Manganês/toxicidade , Manganês/metabolismo , Microglia/metabolismo , Microglia/patologia , Doenças Neuroinflamatórias , NF-kappa B/metabolismo , Nucleotidiltransferases/metabolismo , Nucleotidiltransferases/farmacologia , Nucleotidiltransferases/uso terapêutico
6.
J Agric Food Chem ; 71(3): 1381-1390, 2023 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-36624936

RESUMO

High resistance to benzimidazole fungicides in Venturia carpophila is caused by the point mutation E198K of the ß-tubulin (TUB2) gene. Traditional methods for detection of fungicide resistance are time-consuming, which are routinely based on tedious operation, reliance on expensive equipment, and specially trained people. Therefore, it is important to establish efficient methods for field detection of benzimidazole resistance in V. carpophila to make suitable management strategies and ensure food safety. Based on recombinase polymerase amplification (RPA) combined with CRISPR/Cas12a, a rapid one-pot assay ORCas12a-BRVc (one-pot RPA-CRISPR/Cas12 platform) was established for the detection of benzimidazole resistance in V. carpophila. The ORCas12a-BRVc assay enabled one-pot detection by adding components at the bottom and wall of the tube separately, solving the problems of aerosol contamination and decreased sensitivity caused by competing DNA substrates between Cas12a cleavage and RPA amplification. The ORCas12a-BRVc assay could accomplish the detection with a minimum of 7.82 × 103 fg µL-1 V. carpophila genomic DNA in 45 min at 37 °C. Meanwhile, this assay showed excellent specificity due to the specific recognition ability of the Cas12a-crRNA complex. Further, we combined a method that could rapidly extract DNA from V. carpophila within 2 min with the ORCas12a-BRVc to achieve more rapid and simple detection of V. carpophila with benzimidazole resistance in fields. The ORCas12a-BRVc assay has the advantages of simplicity, rapidity, high sensitivity, high specificity, and ease of operation without the need for precision instruments and the need to isolate and culture pathogens. This assay is the first application of the one-pot platform based on the combination of RPA and CRISPR/Cas12a in fungicide resistance detection and can be used for monitoring of resistant populations in fields, providing guidance on making suitable management strategies for peach scab.


Assuntos
Fungicidas Industriais , Recombinases , Humanos , Sistemas CRISPR-Cas , Nucleotidiltransferases , Benzimidazóis/farmacologia , Técnicas de Amplificação de Ácido Nucleico
7.
Clin Sci (Lond) ; 137(2): 163-180, 2023 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-36598778

RESUMO

Cigarette smoking is a major risk factor for atherosclerosis. We previously reported that DNA damage was accumulated in atherosclerotic plaque, and was increased in human mononuclear cells by smoking. As vascular endothelial cells are known to modulate inflammation, we investigated the mechanism by which smoking activates innate immunity in endothelial cells focusing on DNA damage. Furthermore, we sought to characterize the plasma level of cell-free DNA (cfDNA), a result of mitochondrial and/or genomic DNA damage, as a biomarker for atherosclerosis. Cigarette smoke extract (CSE) increased DNA damage in the nucleus and mitochondria in human endothelial cells. Mitochondrial damage induced minority mitochondrial outer membrane permeabilization, which was insufficient for cell death but instead led to nuclear DNA damage. DNA fragments, derived from the nucleus and mitochondria, were accumulated in the cytosol, and caused a persistent increase in IL-6 mRNA expression via the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway. cfDNA, quantified with quantitative PCR in culture medium was increased by CSE. Consistent with in vitro results, plasma mitochondrial cfDNA (mt-cfDNA) and nuclear cfDNA (n-cfDNA) were increased in young healthy smokers compared with age-matched nonsmokers. Additionally, both mt-cfDNA and n-cfDNA were significantly increased in patients with atherosclerosis compared with the normal controls. Our multivariate analysis revealed that only mt-cfDNA predicted the risk of atherosclerosis. In conclusion, accumulated cytosolic DNA caused by cigarette smoke and the resultant activation of the cGAS-STING pathway may be a mechanism of atherosclerosis development. The plasma level of mt-cfDNA, possibly as a result of DNA damage, may be a useful biomarker for atherosclerosis.


Assuntos
Aterosclerose , Ácidos Nucleicos Livres , Fumar Cigarros , Humanos , DNA Mitocondrial/metabolismo , Células Endoteliais/metabolismo , Mitocôndrias/metabolismo , Nucleotidiltransferases/genética , Aterosclerose/metabolismo , Ácidos Nucleicos Livres/metabolismo
8.
Mol Cell ; 83(2): 266-280.e6, 2023 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-36638783

RESUMO

Tumor suppression by TP53 involves cell-autonomous and non-cell-autonomous mechanisms. TP53 can suppress tumor growth by modulating immune system functions; however, the mechanistic basis for this activity is not well understood. We report that p53 promotes the degradation of the DNA exonuclease TREX1, resulting in cytosolic dsDNA accumulation. We demonstrate that p53 requires the ubiquitin ligase TRIM24 to induce TREX1 degradation. The cytosolic DNA accumulation resulting from TREX1 degradation activates the cytosolic DNA-sensing cGAS/STING pathway, resulting in induction of type I interferons. TREX1 overexpression sufficed to block p53 activation of the cGAS/STING pathway. p53-mediated induction of type I interferon (IFNB1) is suppressed by cGAS/STING knockout, and p53's tumor suppressor activities are compromised by the loss of signaling through the cGAS/STING pathway. Thus, our study reveals that p53 utilizes the cGAS/STING innate immune system pathway for both cell-intrinsic and cell-extrinsic tumor suppressor activities.


Assuntos
Imunidade Inata , Interferon Tipo I , Imunidade Inata/genética , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Nucleotidiltransferases/genética , Nucleotidiltransferases/metabolismo , DNA/metabolismo , Transdução de Sinais/fisiologia , Interferon Tipo I/metabolismo
9.
Viral Immunol ; 36(1): 48-54, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36493365

RESUMO

To study the clinical significance of manganese (Mn) in the serum of children with infectious mononucleosis (IM) caused by Epstein-Barr virus (EBV) infection, we analyzed the correlation between Mn and the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway and explored the immune pathogenesis of EBV infection. Children diagnosed with IM comprised the IM group, and healthy children during the same period were selected as the normal control group. Real-time reverse transcription-polymerase chain reaction was used to detect the mRNA expression levels of cGAS, STING, Tank-binding kinase 1 (TBK1), interferon regulatory factor 3 (IRF3), and related inflammatory factors, and Mn in serum was detected by inductively coupled plasma mass spectrometry. Interferon (IFN)-α and IFN-ß expression levels in serum were detected by enzyme-linked immunosorbent assay, and the correlation between Mn levels and clinical manifestations and laboratory tests was analyzed. Mn levels and the expression levels of cGAS, STING, and related inflammatory factors were significantly higher in children with IM than in healthy children. Furthermore, Mn levels in children with IM were positively correlated with the expression levels of cGAS and related inflammatory factors. Thus, Mn, cGAS, STING, and inflammatory cytokines may be involved in the immune mechanism of IM caused by EBV infection.


Assuntos
Infecções por Vírus Epstein-Barr , Mononucleose Infecciosa , Criança , Humanos , Herpesvirus Humano 4 , Manganês , Nucleotidiltransferases/genética , Nucleotidiltransferases/metabolismo , Interferons
10.
Zool Res ; 44(1): 183-218, 2023 Jan 18.
Artigo em Inglês | MEDLINE | ID: mdl-36579404

RESUMO

The innate immune system protects the host from external pathogens and internal damage in various ways. The cGAS-STING signaling pathway, comprised of cyclic GMP-AMP synthase (cGAS), stimulator of interferon genes (STING), and downstream signaling adaptors, plays an essential role in protective immune defense against microbial DNA and internal damaged-associated DNA and is responsible for various immune-related diseases. After binding with DNA, cytosolic cGAS undergoes conformational change and DNA-linked liquid-liquid phase separation to produce 2'3'-cGAMP for the activation of endoplasmic reticulum (ER)-localized STING. However, further studies revealed that cGAS is predominantly expressed in the nucleus and strictly tethered to chromatin to prevent binding with nuclear DNA, and functions differently from cytosolic-localized cGAS. Detailed delineation of this pathway, including its structure, signaling, and regulatory mechanisms, is of great significance to fully understand the diversity of cGAS-STING activation and signaling and will be of benefit for the treatment of inflammatory diseases and cancer. Here, we review recent progress on the above-mentioned perspectives of the cGAS-STING signaling pathway and discuss new avenues for further study.


Assuntos
Imunidade Inata , Transdução de Sinais , Animais , Transdução de Sinais/fisiologia , Nucleotidiltransferases/genética , Nucleotidiltransferases/química , Nucleotidiltransferases/metabolismo , DNA
11.
Exp Neurol ; 359: 114269, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36343680

RESUMO

Stroke patients are often complicated by cerebral ischemia-reperfusion injury (CIRI) after the restoration of cerebral perfusion, and how to prevent CIRI at an early stage has received close attention. The imbalance of iron metabolism is one of the essential factors in the aggravation of CIRI, and NCOA4-mediated ferritinophagy, as a critical pathway to regulate iron metabolism, is expected to be an effective intervention target. We established a mouse model of cerebral ischemia-reperfusion (CIR) with NCOA4 silencing. We found that activation of NCOA4-mediated ferritinophagy atthe early stage of CIR mediated the onset of oxidative stress and contributed to autophagy and apoptosis, and eventually resulted in increased brain injury. This suggests that NCOA4-mediated ferritinophagy plays a vital role in early CIR and can be an effective target to prevent and treat CIRI. We next explored the upstream regulatory targets of NCOA4-mediated ferritinophagy. The previous evidence for the cGAS-STING pathway's importance during CIR and its strong relationship with autophagy attracted our attention. To investigate whether the cGAS-STING pathway regulates NCOA4-mediated ferritinophagy, we further administered a cGAS inhibitor to mice with CIR and overexpressed NCOA4. Along with the inhibition of the cGAS-STING pathway, ferritinophagy, oxidative stress, autophagy, and apoptosis were inhibited, and CIRI was ameliorated, which was attenuated by NCOA4 overexpression. In conclusion, our results suggest that activation of the cGAS-STING pathway exacerbates CIRI at the early stage of CIR, which may be achieved by mediating NCOA4-mediated ferritinophagy.


Assuntos
Isquemia Encefálica , Coativadores de Receptor Nuclear , Traumatismo por Reperfusão , Animais , Camundongos , Autofagia , Ferro/metabolismo , Coativadores de Receptor Nuclear/genética , Coativadores de Receptor Nuclear/metabolismo , Nucleotidiltransferases/metabolismo , Nucleotidiltransferases/farmacologia , Traumatismo por Reperfusão/metabolismo , Fatores de Transcrição/metabolismo , Ferritinas/metabolismo
12.
J Exp Med ; 220(3)2023 Mar 06.
Artigo em Inglês | MEDLINE | ID: mdl-36534085

RESUMO

Late cardiac toxicity is a potentially lethal complication of cancer therapy, yet the pathogenic mechanism remains largely unknown, and few treatment options exist. Here we report DNA-damaging agents such as radiation and anthracycline chemotherapies inducing delayed cardiac inflammation following therapy due to activation of cGAS- and STING-dependent type I interferon signaling. Genetic ablation of cGAS-STING signaling in mice inhibits DNA damage-induced cardiac inflammation, rescues late cardiac functional decline, and prevents death from cardiac events. Treatment with a STING antagonist suppresses cardiac interferon signaling following DNA-damaging therapies and effectively mitigates cardiac toxicity. These results identify a therapeutically targetable, pathogenic mechanism for one of the most vexing treatment-related toxicities in cancer survivors.


Assuntos
Antineoplásicos , Cardiotoxicidade , Dano ao DNA , Neoplasias , Animais , Camundongos , Imunidade Inata , Inflamação , Neoplasias/tratamento farmacológico , Nucleotidiltransferases/genética , Antineoplásicos/efeitos adversos
13.
J Agric Food Chem ; 71(1): 569-579, 2023 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-36583613

RESUMO

Di(2-ethylhexyl) phthalate (DEHP) is a highly harmful and persistent environmental pollutant. Due to its unique chemical composition, it frequently dissolves and enters the environment to endanger human and animal health. Lycopene is a natural bioactive component that can potentially reduce the risk of environmental factor-induced chronic diseases. The present study sought to explore the role and underlying mechanism of lycopene (LYC) on DEHP-induced renal inflammatory response and apoptosis. In this study, mice were orally treated with LYC (5 mg/kg BW/day) and/or DEHP (500 or 1000 mg/kg BW/day) for 28 days. Our results indicated that LYC prevented DEHP-induced histopathological alterations and ultrastructural injuries, including decreased mitochondrial membrane potential (ΔΨm), PINK1/Parkin pathway-mediated mitophagy, and mitochondrial energetic deficit. When damaged mitochondria release mitochondrial DNA (mtDNA) into cytosol, LYC can alleviate inflammation and apoptosis caused by DEHP exposure by activating the cyclic GMP-AMP synthase-stimulator of interferon gene (cGAS-STING) signal pathway. Collectively, our data demonstrate that LYC can reduce mitophagy caused by DEHP exposure by activating the PINK1/Parkin pathway and then reduce renal inflammation and apoptosis through the cGAS-STING pathway.


Assuntos
Dietilexilftalato , Animais , Camundongos , Dietilexilftalato/toxicidade , DNA Mitocondrial/metabolismo , Inflamação/genética , Interferons , Rim/metabolismo , Licopeno , Nucleotidiltransferases/metabolismo , Proteínas Quinases/metabolismo , Transdução de Sinais , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
14.
J Agric Food Chem ; 71(1): 944-951, 2023 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-36548210

RESUMO

Enterocytozoon hepatopenaei (EHP) is one of the most serious pathogens in shrimp farming. This study combines recombinase polymerase amplification (RPA) with the Argonaute from Pyrococcus furiosus (PfAgo) and establishes a sensitive and reliable method for on-site detection of EHP. With careful screening of gDNA and optimization of the reaction, the method shows a good specificity and reaches a sensitivity of single copy per reaction, which is higher than the sensitivity of the currently available molecular assays. The whole procedure can be finished within 1.5 h including the sample processing time and only requires minimum laboratory support, which is user-friendly for on-site environments. This is the first application of PfAgo for the diagnosis of infectious diseases in seafood supply chains. It provides a reliable method for on-site detection of EHP in shrimp farms and establishes a groundwork for multiplex detection of important pathogens in seafood farming using PfAgo.


Assuntos
Penaeidae , Pyrococcus furiosus , Animais , Reação em Cadeia da Polimerase/métodos , Recombinases/genética , Pyrococcus furiosus/genética , Nucleotidiltransferases
15.
Biomaterials ; 293: 121988, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36580716

RESUMO

As the clinical efficacy of immunotherapy for triple-negative breast cancer (TNBC) remains limited, exploring new immunotherapy approaches is still indispensable. Mn2+ has been proven as a cGAS-STING agonist to remarkably enhance antitumor immunity. Here, we report a combined tumor-therapeutic strategy based on Prussian blue (PB)-mediated photothermal therapy with Mn2+-augmented immunotherapy by synergistically activating the cGAS-STING pathway. Mn-enriched photonic nanomedicine (MnPB-MnOx) were constructed by integrating MnOx onto the surface of Mn-doped PB nanoparticles. All components of MnPB-MnOx are biocompatible and biodegradable, wherein sufficient Mn are endowed through rational nanostructure design, conferring easier cGAS-STING activation. Additionally, tumor hyperthermia strengthened by MnPB under near-infrared light radiation, synergistic with the generation of reactive oxygen species catalyzed by MnOx, double hits cancer cells to release abundant tumor-associated antigens for further promoting immune response stimulation. The local anti-TNBC efficacy of photothermal/immuno-therapy has been proven effective in subcutaneous 4T1-bearing mice. Especially, it has been systematically demonstrated in bilateral orthotopic 4T1-bearing mice that the as-proposed treatment could successfully activate innate and adaptive immunity, and local therapy could engender systemic responses to suppress the distant tumors. Collectively, this work represents a proof-of-concept for a non-invasive Mn-based tumor-immunotherapeutic modality, providing a paradigm for the immunotherapy of metastatic-prone tumors.


Assuntos
Hipertermia Induzida , Neoplasias , Neoplasias de Mama Triplo Negativas , Animais , Humanos , Camundongos , Catálise , Imunoterapia , Manganês , Nanomedicina , Neoplasias/terapia , Nucleotidiltransferases/metabolismo , Neoplasias de Mama Triplo Negativas/terapia
16.
Biomed Res Int ; 2022: 6189254, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36457340

RESUMO

Coronavirus illness 2019 is a significant worldwide health danger that began with severe acute respiratory syndrome coronavirus two infections. It is the largest pandemic of our lifetime to date, affecting millions of people and crippling economies globally. There is currently no viable therapy for this devastating condition. The fast spread of SARS-CoV-2 underlines the critical need for favorable treatments to prevent SARS-CoV-2 infection and dissemination. Regulating the upstream cytokine release might be a possible method for COVID-19 therapy. We propose that more consideration be paid to the dysregulated IFN-I release in COVID-19 and that cGAS and STING be considered therapeutic targets for avoiding cytokine storms and as critical components in host antiviral defense mechanisms.


Assuntos
Proteínas de Membrana , Nucleotidiltransferases , SARS-CoV-2 , Humanos , Pandemias
17.
Nat Commun ; 13(1): 7447, 2022 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-36460671

RESUMO

Control of RNA Polymerase II (pol II) elongation is a critical component of gene expression in mammalian cells. The PNUTS-PP1 complex controls elongation rates, slowing pol II after polyadenylation sites to promote termination. The Kaposi's sarcoma-associated herpesvirus (KSHV) co-opts pol II to express its genes, but little is known about its regulation of pol II elongation. We identified PNUTS as a suppressor of a KSHV reporter gene in a genome-wide CRISPR screen. PNUTS depletion enhances global KSHV gene expression and overall viral replication. Mechanistically, PNUTS requires PP1 interaction, binds viral RNAs downstream of polyadenylation sites, and restricts transcription readthrough of viral genes. Surprisingly, PNUTS also represses productive elongation at the 5´ ends of the KSHV reporter and the KSHV T1.4 RNA. From these data, we conclude that PNUTS' activity constitutes an intrinsic barrier to KSHV replication likely by suppressing pol II elongation at promoter-proximal regions.


Assuntos
Herpesviridae , Herpesvirus Humano 8 , Mieloma Múltiplo , Sarcoma de Kaposi , Animais , Herpesvirus Humano 8/genética , Nucleotidiltransferases , RNA Viral/genética , Transcrição Genética , Mamíferos
18.
Med Oncol ; 40(1): 30, 2022 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-36460853

RESUMO

Breast cancer is one of the most frequent causes of cancer related death worldwide, and despite the significant advances in therapeutic approaches, a significant proportion of patients succumb to metastasis and tumor recurrence. Breast cancer is an immunogenic cancer, and therefore, immunotherapy is considered a major therapeutic strategy. The survival rate has been increased significantly in HER2+ breast cancers after immunotherapy by monoclonal antibodies alone, or combined with chemical anti-cancer agents. Moreover, in triple negative breast cancer (TNBC), a number of novel agents called immune checkpoint inhibitors have shown optimal efficacy. The major hindrance in cancer immunotherapy is frequent development of resistance and cancer remission. cGAS-STING pathway has a key role in anti-cancer immunity as its downstream signals especially type I interferon (IFN) acts as a link between innate and adaptive immunity. Considering the roles of type I IFN in enhancing dendritic cells activity, promoting the functions of CD8+ T cells, and protecting the effector cells against apoptosis, the induction of cGAS-STING pathway demonstrated promising therapeutic effects against breast cancer, especially in triple negative breast cancers. In this review, we discuss the latest findings and the recent advances regarding the role of cGAS-STING pathway and its activation in breast cancer.


Assuntos
Linfócitos T CD8-Positivos , Neoplasias de Mama Triplo Negativas , Humanos , Recidiva Local de Neoplasia , Neoplasias de Mama Triplo Negativas/terapia , Imunoterapia , Nucleotidiltransferases
19.
PLoS One ; 17(12): e0278869, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36508428

RESUMO

Shigella spp. and enteroinvasive Escherichia coli (EIEC) are widely distributed and can cause serious food-borne diseases for humans such as dysentery. Therefore, an efficient detection platform is needed to detect Shigella and EIEC quickly and sensitively. In this study, a method called recombinase polymerase amplification combined with lateral flow dipstick (RPA-LFD) was developed for rapid detection of Shigella and EIEC. RPA primers and LFD detection probes were designed for their shared virulence gene ipaH. Primers and probes were screened, and the primer concentration, and reaction time and temperature were optimized. According to the optimization results, the RPA reaction should be performed at 39°C, and when combined with LFD, it takes less than 25 min for detection with the naked eye. The developed RPA-LFD method specifically targets gene ipaH and has no cross-reactivity with other common food-borne pathogens. In addition, the minimum detection limit of RPA-LFD is 1.29×102 copies/µL. The detection of food sample showed that the RPA-LFD method was also verified for the detection of actual samples.


Assuntos
Recombinases , Shigella , Humanos , Técnicas de Amplificação de Ácido Nucleico/métodos , Escherichia coli/genética , Sensibilidade e Especificidade , Nucleotidiltransferases , Shigella/genética
20.
Front Cell Infect Microbiol ; 12: 1019071, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36519130

RESUMO

After the outbreak of SARS-CoV-2, nucleic acid testing quickly entered people's lives. In addition to the polymerase chain reaction (PCR) which was commonly used in nucleic acid testing, isothermal amplification methods were also important nucleic acid testing methods. Among several common isothermal amplification methods like displaced amplification, rolling circle amplification, and so on, recombinase polymerase amplification (RPA) was recently paid more attention to. It had the advantages like a simple operation, fast amplification speed, and reaction at 37-42°C, et al. So it was very suitable for field detection. However, there were still some disadvantages to RPA. Herein, our review mainly summarized the principle, advantages, and disadvantages of RPA. The specific applications of RPA in bacterial detection, fungi detection, virus detection, parasite detection, drug resistance gene detection, genetically modified food detection, and SARS-CoV-2 detection were also described. It was hoped that the latest research progress on RPA could be better delivered to the readers who were interested in RPA.


Assuntos
COVID-19 , Técnicas de Amplificação de Ácido Nucleico , Humanos , COVID-19/diagnóstico , Nucleotidiltransferases/genética , Recombinases/genética , SARS-CoV-2/genética , Sensibilidade e Especificidade
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