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1.
BMC Genomics ; 23(1): 33, 2022 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-34996349

RESUMO

BACKGROUND: Long non-coding RNAs (lncRNAs) play a critical role in the pathogenesis of hypoxic pulmonary hypertension (HPH). The role of N7-methylguanosine (m7G) modification in lncRNAs has received increased attentions in recent years. However, the m7G-methylation of lncRNA in HPH has yet to be determined. We have therefore performed a transcriptome-wide analysis of m7G lncRNAs in HPH. RESULTS: Differentially-expressed m7Gs were detected in HPH, and m7G lncRNAs were significantly upregulated compared with non-m7G lncRNAs in HPH. Importantly, this was the first time that the upregulated m7G lncXR_591973 and m7G lncXR_592398 were identified in HPH. CONCLUSION: This study provides the first m7G transcriptome-wide analysis of HPH. Importantly, two HPH-associated m7G lncRNAs were identified, although their clinical significance requires further validation.


Assuntos
Hipertensão Pulmonar , RNA Longo não Codificante , Animais , Guanosina/análogos & derivados , Hipertensão Pulmonar/genética , Hipóxia/genética , RNA Longo não Codificante/genética , Ratos
2.
J Hazard Mater ; 421: 126801, 2022 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-34388916

RESUMO

A novel chemosensor is developed for the sensitive and facile detection of trace strontium ions (Sr2+) based on the ion-imprinted hydrogels. With Sr2+ as the templates, the ion-imprinted hydrogels are synthesized by copolymerizing the ion-responsive units 5'-O-acryloyl-2',3'-O-isopropylidene guanosine (APG) and the thermo-responsive units N-isopropylacrylamide (NIPAM). In the presence of Sr2+, APG units can self-assemble to form planar G-quartets via the complexation with Sr2+, which are introduced into the gel network during polymerization. Then Sr2+ templates can be removed by multiple repeated washing. When re-exposed to Sr2+, the relaxed G-quartets can recognize Sr2+, leading to the weakening of electrostatic repulsion between the four oxygen atoms in the G-quartets and inducing the shrinkage of the hydrogels. In this work, the Sr2+-imprinted chemosensors are designed as the grating systems for detecting trace Sr2+. Based on the array of hydrogel strings synthesized on a nano-scale, the smart grating systems thus constructed can convert and amplify the Sr2+ concentration signals to the easily-measurable optical signals. With the Sr2+-imprinted hydrogel gratings, trace Sr2+ (10-11 M) in an aqueous solution can be detected sensitively. Moreover, the proposed Sr2+-imprinted chemosensors can be integrated with other smart systems for developing various detectors with high performance.


Assuntos
Hidrogéis , Estrôncio , Guanosina , Íons , Polimerização
3.
J Phys Chem A ; 126(1): 68-79, 2022 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-34941276

RESUMO

8-Bromoguanosine is generated in vivo as a biomarker for early inflammation. Its formation and secondary reactions lead to a variety of biological sequelae at inflammation sites, most of which are mutagenic and linked to cancer. Herein, we report the formation of radical cations of 8-bromoguanine (8BrG•+) and 8-bromoguanosine (8BrGuo•+) and their reactions toward the lowest excited singlet molecular oxygen (1O2)─a common reactive oxygen species generated in biological systems. This work aims to investigate synergistic, oxidatively generated damage of 8-brominated guanine and guanosine that may occur upon ionizing radiation, one-electron oxidation, and 1O2 oxidation. Capitalizing on measurements of reaction product ions and cross sections of 8BrG•+ and 8BrGuo•+ with 1O2 using guided-ion beam tandem mass spectrometry and augmented by computational modeling of the prototype reaction system, 8BrG•+ + 1O2, using the approximately spin-projected ωB97XD/6-31+G(d,p) density functional theory, the coupled cluster DLPNO-CCSD(T)/aug-cc-pVTZ and the multireference CASPT2(21,15)/6-31G**, probable reaction products, and potential energy surfaces (PESs) were mapped out. 8BrG•+ and 8BrGuo•+ present similar exothermic oxidation products, and their reaction efficiencies with 1O2 increase with decreasing collision energy. Both single- and multireference theories predicted that the two most energetically favorable reaction pathways correspond to 1O2-addition to the C8 and C5-positions of 8BrG•+, respectively. The CASPT2-calculated PES represents the best quantitative agreement with the experimental benchmark, in that the oxidation exothermicity is close to the water hydration energy of product ions and, thus, is able to eliminate a water ligand in the product ions.


Assuntos
Guanina , Guanosina , Cátions , Guanina/análogos & derivados , Guanosina/análogos & derivados , Cinética
4.
Curr Protoc ; 1(11): e297, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34837670

RESUMO

A convenient synthetic method for preparing 3-deazapurine nucleosides (3-deazainosine, 3-deazaadenosine, and 3-deazaguanosine) from inosine via a 5-ethynyl-1-ß-D-ribofuranosylimidazole-4-carboxamide (EICAR) derivative, which is a key intermediate, is described. A large-scale synthesis of an EICAR derivative starting from inosine was achieved in six steps via dinitrophenylation at the N1 position followed by ring opening, iodination of the resulting 5-amino group, and a palladium-catalyzed cross-coupling reaction. The resulting EICAR derivative was then converted into 3-deazainosine, 3-deazaadenosine, and 3-deazaguanosine. This route enabled us to synthesize 3-deazapurine nucleosides conveniently in good yields. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Preparation of 5-ethynyl-1-ß-D-ribofuranosylimidazole-4-carboxamide (EICAR) derivative 6 Basic Protocol 2: Preparation of 3-deazapurine nucleosides 8, 11, and 14.


Assuntos
Inosina , Nucleosídeos , Guanosina/análogos & derivados , Tubercidina
5.
Analyst ; 146(19): 5866-5872, 2021 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-34570847

RESUMO

DNA-tuned dye assemblies have received considerable attention toward developing various devices. Owing to easy conformation implementation, G-quadruplexes (G4s) have been extensively used as initiators to grow dye assemblies with controllable chiralities. However, programmed chirality regulation of dye assemblies for a given G4 sequence has not been realized in a straightforward manner. In this work, we replaced a middle guanine in the G-tracts of a human telomeric G4 with an apurinic site (AP site) to meet the programmed dye assemblies. Although all of the AP site replacements altered the G4 conformation from the hybrid to the antiparallel folding, the handedness of pinacyanol (PIN) assemblies grown on the AP site-containing G4 was programmably regulated. The G4 with the AP site at the 5'-most G-tract grew right-handed assemblies, while that with the AP site at the 3'-most G-tract grew left-handed assemblies. The handedness of assemblies almost totally mirrored each other within 450-700 nm. Interestingly, we found that the AP site provided a specific binding site for guanosine and guanine, and this binding event sensitively broke the chiral assemblies. Thus, dye assembly-based sensors can be easily established based on the chiral responses with a high selectivity and sensitivity. Our work first demonstrates the AP site programmed chirality regulation of G4-grown dye assemblies and will find wide application in chiral devices.


Assuntos
Quadruplex G , DNA , Guanina , Guanosina , Humanos , Telômero
6.
Methods Enzymol ; 658: 25-47, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34517949

RESUMO

Precise and reliable mapping of modified nucleotides in RNA is a challenging task in epitranscriptomics analysis. Only deep sequencing-based methods are able to provide both, a single-nucleotide resolution and sufficient selectivity and sensitivity. A number of protocols employing specific chemical reagents to distinguish modified RNA nucleotides from canonical parental residues have already proven their performance. We developed a deep-sequencing analytical pipeline for simultaneous detection of several modified nucleotides of different nature (methylation, hydroxylation, reduction) in RNA. The AlkAniline-Seq protocol uses intrinsic fragility of the N-glycosidic bond present in certain modified residues (7-methylguanosine (m7G), 3-methylcytidine (m3C), dihydrouridine (D) and 5-hydroxycytidine (ho5C)) to induce cleavage under heat combined with alkaline conditions. The resulting RNA abasic site is decomposed by aniline-driven ß-elimination and creates a 5'-phosphate (5'-P) at the adjacent N+1 residue. This 5'-P is the crucial entry point for a highly selective ligation of sequencing adapters during the subsequent Illumina library preparation protocol. AlkAniline-Seq protocol has a very low background, and is both highly sensitive and specific. Applications of AlkAniline-Seq include mapping of m7G, m3C, D, and ho5C in variety of cellular RNAs, including in particular rRNAs and tRNAs.


Assuntos
Citidina , Guanosina , Citidina/análogos & derivados , Guanosina/análogos & derivados , Sequenciamento de Nucleotídeos em Larga Escala , RNA/genética , RNA de Transferência/genética , Análise de Sequência de RNA
7.
Zhongguo Zhong Yao Za Zhi ; 46(12): 2912-2922, 2021 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-34467681

RESUMO

The dried fruit body of Phylloporia ribis(Hymenochaetaceae), which prefers to live on the stumps of Lonicera japonica(Caprifoliaceae), has a variety of activities, whereas its pharmacodynamic material basis is not completely clear and there are few reports on its quality control and evaluation. In this study, an UPLC-Q-TOF-MS method was used to analyze the nucleosides and nucleobases in P. ribis and a HPLC method was established for simultaneous determination of 10 nucleosides and nucleobases. MS and MS/MS data were acquired in positive ion mode. Based on the data comparison of the sample and the reference substance, the literature data and the compound databases of ChemSpider and PubChem, 18 nucleosides and nucleobases were identified qualitatively from the water extract of P. ribis for the first time. After optimization, the HPLC was performed using a Welch Ultimate AQ C_(18) column(4.6 mm×250 mm, 5 µm) by gradient elution with acetonitrile and water as mobile phase, the flow rate of 1.0 mL·min~(-1), the detection wavelength of 260 nm, and the column temperature of 30 ℃. Through the investigation of the extraction method, solvent and time, it was determined that the test solution should be obtained by cold water extraction for 18 h. At the present HPLC conditions, 10 components of uracil, cytidine, hypoxanthine, uridine, thymine, inosine, guanosine, 2'-deoxyinosine, 2'-deoxyguanosine and thymidine could be well separated(R > 1.5) and showed good linearity(r > 0.999 9) in the concentration ranges of 0.247-24.7, 0.283-28.3, 0.273-27.3, 0.256-25.6, 0.257-25.7, 0.318-31.8, 0.245-24.5, 0.267-26.7, 0.250-25.0 and 0.267-26.7 mg·L~(-1), respectively. The average reco-veries of 10 components were 95.78%-104.5%, and the RSDs were 2.2%-5.2%(n=6). The contents of 10 nucleosides and nucleobases in different samples of P. ribis varied greatly, which were 0.021-0.122, 0.004-0.029, 0.014-0.226, 0.009-0.442, 0.003-0.014, 0.002-0.146, 0.007-0.098, 0-0.054, 0.005-0.069, 0.004-0.081 and 0.072-1.28 mg·g~(-1) for uracil, cytidine, hypoxanthine, uridine, thymine, inosine, guanosine, 2'-deoxyinosine, 2'-deoxyguanosine, thymidine and total 10 components, respectively. These results demonstrated that the components had significant differences in the internal quality, and good quality control was needed to ensure the medical efficacy. This study provides a scientific basis for the discovery of pharmacodynamic ingredients, quality control and evaluation of P. ribis.


Assuntos
Nucleosídeos , Espectrometria de Massas em Tandem , Basidiomycota , Cromatografia Líquida de Alta Pressão , Guanosina
8.
J Chem Phys ; 155(9): 094305, 2021 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-34496579

RESUMO

DNA strands are polymeric ligands that both protect and tune molecular-sized silver cluster chromophores. We studied single-stranded DNA C4AC4TC3XT4 with X = guanosine and inosine that form a green fluorescent Ag10 6+ cluster, but these two hosts are distinguished by their binding sites and the brightness of their Ag10 6+ adducts. The nucleobase subunits in these oligomers collectively coordinate this cluster, and fs time-resolved infrared spectra previously identified one point of contact between the C2-NH2 of the X = guanosine, an interaction that is precluded for inosine. Furthermore, this single nucleobase controls the cluster fluorescence as the X = guanosine complex is ∼2.5× dimmer. We discuss the electronic relaxation in these two complexes using transient absorption spectroscopy in the time window 200 fs-400 µs. Three prominent features emerged: a ground state bleach, an excited state absorption, and a stimulated emission. Stimulated emission at the earliest delay time (200 fs) suggests that the emissive state is populated promptly following photoexcitation. Concurrently, the excited state decays and the ground state recovers, and these changes are ∼2× faster for the X = guanosine compared to the X = inosine cluster, paralleling their brightness difference. In contrast to similar radiative decay rates, the nonradiative decay rate is 7× higher with the X = guanosine vs inosine strand. A minor decay channel via a dark state is discussed. The possible correlation between the nonradiative decay and selective coordination with the X = guanosine/inosine suggests that specific nucleobase subunits within a DNA strand can modulate cluster-ligand interactions and, in turn, cluster brightness.


Assuntos
DNA de Cadeia Simples/química , Guanosina/química , Inosina/química , Prata/química , Sítios de Ligação , Fluorescência
9.
Mol Cell ; 81(16): 3339-3355.e8, 2021 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-34352206

RESUMO

Cancer cells selectively promote translation of specific oncogenic transcripts to facilitate cancer survival and progression, but the underlying mechanisms are poorly understood. Here, we find that N7-methylguanosine (m7G) tRNA modification and its methyltransferase complex components, METTL1 and WDR4, are significantly upregulated in intrahepatic cholangiocarcinoma (ICC) and associated with poor prognosis. We further reveal the critical role of METTL1/WDR4 in promoting ICC cell survival and progression using loss- and gain-of-function assays in vitro and in vivo. Mechanistically, m7G tRNA modification selectively regulates the translation of oncogenic transcripts, including cell-cycle and epidermal growth factor receptor (EGFR) pathway genes, in m7G-tRNA-decoded codon-frequency-dependent mechanisms. Moreover, using overexpression and knockout mouse models, we demonstrate the crucial oncogenic function of Mettl1-mediated m7G tRNA modification in promoting ICC tumorigenesis and progression in vivo. Our study uncovers the important physiological function and mechanism of METTL1-mediated m7G tRNA modification in the regulation of oncogenic mRNA translation and cancer progression.


Assuntos
Colangiocarcinoma/genética , Proteínas de Ligação ao GTP/genética , Metiltransferases/genética , Biossíntese de Proteínas , Animais , Carcinogênese/genética , Colangiocarcinoma/patologia , Progressão da Doença , Receptores ErbB/genética , Guanosina/análogos & derivados , Guanosina/genética , Humanos , Camundongos , Processamento Pós-Transcricional do RNA/genética , RNA Mensageiro/genética , RNA de Transferência/genética
10.
Mol Cell ; 81(16): 3323-3338.e14, 2021 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-34352207

RESUMO

The emerging "epitranscriptomics" field is providing insights into the biological and pathological roles of different RNA modifications. The RNA methyltransferase METTL1 catalyzes N7-methylguanosine (m7G) modification of tRNAs. Here we find METTL1 is frequently amplified and overexpressed in cancers and is associated with poor patient survival. METTL1 depletion causes decreased abundance of m7G-modified tRNAs and altered cell cycle and inhibits oncogenicity. Conversely, METTL1 overexpression induces oncogenic cell transformation and cancer. Mechanistically, we find increased abundance of m7G-modified tRNAs, in particular Arg-TCT-4-1, and increased translation of mRNAs, including cell cycle regulators that are enriched in the corresponding AGA codon. Accordingly, Arg-TCT expression is elevated in many tumor types and is associated with patient survival, and strikingly, overexpression of this individual tRNA induces oncogenic transformation. Thus, METTL1-mediated tRNA modification drives oncogenic transformation through a remodeling of the mRNA "translatome" to increase expression of growth-promoting proteins and represents a promising anti-cancer target.


Assuntos
Carcinogênese/genética , Metiltransferases/genética , Neoplasias/genética , tRNA Metiltransferases/genética , Guanosina/análogos & derivados , Guanosina/genética , Humanos , Metilação , Neoplasias/patologia , Oncogenes/genética , Processamento Pós-Transcricional do RNA/genética , RNA Mensageiro/genética , RNA de Transferência/genética
11.
Psychopharmacology (Berl) ; 238(9): 2555-2568, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34342672

RESUMO

RATIONALE: Guanosine has been shown to potentiate ketamine's antidepressant-like actions, although its ability to augment the anxiolytic effect of ketamine remains to be determined. OBJECTIVE: This study investigated the anxiolytic-like effects of a single administration with low doses of ketamine and/or guanosine in mice subjected to chronic administration of corticosterone and the role of NLRP3-driven signaling. METHODS: Corticosterone (20 mg/kg, p.o.) was administered for 21 days, followed by a single administration of ketamine (0.1 mg/kg, i.p.), guanosine (0.01 mg/kg, p.o.), or ketamine (0.1 mg/kg, i.p.) plus guanosine (0.01 mg/kg, p.o.). Anxiety-like behavior and NLRP3-related targets were analyzed 24 h following treatments. RESULTS: Corticosterone reduced the time spent in the open arms and the central zone in the elevated plus-maze test and open-field test, respectively. Corticosterone raised the number of unsupported rearings and the number and time of grooming, and decreased the latency to start grooming in the open-field test. Disturbances in regional distribution (increased rostral grooming) and grooming transitions (increased aborted and total incorrect transitions) were detected in corticosterone-treated mice. These behavioral alterations were accompanied by increased immunocontent of Iba-1, ASC, NLRP3, caspase-1, TXNIP, and IL-1ß in the hippocampus, but not in the prefrontal cortex. The treatments with ketamine, guanosine, and ketamine plus guanosine were effective to counteract corticosterone-induced anxiety-like phenotype, but not disturbances in the hippocampal NLRP3 pathway. CONCLUSIONS: Our study provides novel evidence that low doses of ketamine and/or guanosine reverse corticosterone-induced anxiety-like behavior and shows that the NLRP3 inflammasome pathway is likely unrelated to this response.


Assuntos
Ketamina , Animais , Ansiedade/induzido quimicamente , Ansiedade/tratamento farmacológico , Comportamento Animal , Corticosterona , Depressão , Guanosina , Hipocampo , Inflamassomos , Ketamina/farmacologia , Camundongos , Proteína 3 que Contém Domínio de Pirina da Família NLR
12.
Antimicrob Agents Chemother ; 65(11): e0098821, 2021 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-34424050

RESUMO

Every year, millions of people worldwide are infected with dengue virus (DENV), with a significant number developing severe life-threatening disease. There are currently no broadly indicated vaccines or therapeutics available for treatment of DENV infection. Here, we show that AT-281, the free base of AT-752, an orally available double prodrug of a guanosine nucleotide analog, was a potent inhibitor of DENV serotypes 2 and 3 in vitro, requiring concentrations of 0.48 and 0.77 µM, respectively, to inhibit viral replication by 50% (EC50) in Huh-7 cells. AT-281 was also a potent inhibitor of all other flaviviruses tested, with EC50 values ranging from 0.19 to 1.41 µM. Little to no cytotoxicity was observed for AT-281 at concentrations up to 170 µM. After oral administration of AT-752, substantial levels of the active triphosphate metabolite AT-9010 were formed in vivo in peripheral blood mononuclear cells of mice, rats, and monkeys. Furthermore, AT-9010 competed with GTP in RNA template-primer elongation assays with DENV2 RNA polymerase, which is essential for viral replication, with incorporation of AT-9010 resulting in termination of RNA synthesis. In AG129 mice infected with DENV D2Y98P, treatment with AT-752 significantly reduced viremia and morbidity and increased survival. The demonstrated in vitro and in vivo activity of AT-752 suggests that it is a promising compound for the treatment of dengue virus infection and is currently under evaluation in clinical studies.


Assuntos
Vírus da Dengue , Dengue , Flavivirus , Pró-Fármacos , Animais , Antivirais/farmacologia , Antivirais/uso terapêutico , Dengue/tratamento farmacológico , Guanosina/farmacologia , Guanosina/uso terapêutico , Leucócitos Mononucleares , Camundongos , Nucleotídeos/uso terapêutico , Pró-Fármacos/farmacologia , Pró-Fármacos/uso terapêutico , Ratos , Replicação Viral
13.
Nucleic Acids Res ; 49(14): 8247-8260, 2021 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-34244755

RESUMO

Transfer RNAs (tRNAs) are key players in protein synthesis. To be fully active, tRNAs undergo extensive post-transcriptional modifications, including queuosine (Q), a hypermodified 7-deaza-guanosine present in the anticodon of several tRNAs in bacteria and eukarya. Here, molecular and biochemical approaches revealed that in the protozoan parasite Trypanosoma brucei, Q-containing tRNAs have a preference for the U-ending codons for asparagine, aspartate, tyrosine and histidine, analogous to what has been described in other systems. However, since a lack of tRNA genes in T. brucei mitochondria makes it essential to import a complete set from the cytoplasm, we surprisingly found that Q-modified tRNAs are preferentially imported over their unmodified counterparts. In turn, their absence from mitochondria has a pronounced effect on organellar translation and affects function. Although Q modification in T. brucei is globally important for codon selection, it is more so for mitochondrial protein synthesis. These results provide a unique example of the combined regulatory effect of codon usage and wobble modifications on protein synthesis; all driven by tRNA intracellular transport dynamics.


Assuntos
Mitocôndrias/genética , Conformação de Ácido Nucleico , Nucleosídeo Q/genética , RNA de Transferência/genética , Anticódon/genética , Núcleo Celular/genética , Núcleo Celular/ultraestrutura , Códon/genética , Citoplasma/genética , Citoplasma/ultraestrutura , Guanosina/genética , Biossíntese de Proteínas/genética , Processamento Pós-Transcricional do RNA/genética , RNA de Transferência/ultraestrutura , Trypanosoma brucei brucei/genética
14.
Int J Mol Sci ; 22(14)2021 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-34298953

RESUMO

A novel siphovirus, vB_PagS_MED16 (MED16) was isolated in Lithuania using Pantoea agglomerans strain BSL for the phage propagation. The double-stranded DNA genome of MED16 (46,103 bp) contains 73 predicted open reading frames (ORFs) encoding proteins, but no tRNA. Our comparative sequence analysis revealed that 26 of these ORFs code for unique proteins that have no reliable identity when compared to database entries. Based on phylogenetic analysis, MED16 represents a new genus with siphovirus morphology. In total, 35 MED16 ORFs were given a putative functional annotation, including those coding for the proteins responsible for virion morphogenesis, phage-host interactions, and DNA metabolism. In addition, a gene encoding a preQ0 DNA deoxyribosyltransferase (DpdA) is present in the genome of MED16 and the LC-MS/MS analysis indicates 2'-deoxy-7-amido-7-deazaguanosine (dADG)-modified phage DNA, which, to our knowledge, has never been experimentally validated in genomes of Pantoea phages. Thus, the data presented in this study provide new information on Pantoea-infecting viruses and offer novel insights into the diversity of DNA modifications in bacteriophages.


Assuntos
DNA Viral , Genoma Viral , Guanosina , Fases de Leitura Aberta , Pantoea/virologia , Siphoviridae , Proteínas Virais , DNA Viral/genética , DNA Viral/metabolismo , Guanosina/análogos & derivados , Guanosina/química , Guanosina/metabolismo , Siphoviridae/genética , Siphoviridae/metabolismo , Proteínas Virais/genética , Proteínas Virais/metabolismo
15.
Anal Chem ; 93(31): 10825-10833, 2021 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-34324303

RESUMO

DNA/RNA synthesis precursors are especially vulnerable to damage induced by reactive oxygen species occurring following oxidative stress. Guanosine triphosphates are the prevalent oxidized nucleotides, which can be misincorporated during replication, leading to mutations and cell death. Here, we present a novel method based on micro-Raman spectroscopy, combined with ab initio calculations, for the identification, detection, and quantification of oxidized nucleotides at low concentration. We also show that the Raman signature in the terahertz spectral range (<100 cm-1) contains information on the intermolecular assembly of guanine in tetrads, which allows us to further boost the oxidative damage detection limit. Eventually, we provide evidence that similar analyses can be carried out on samples in very small volumes at very low concentrations by exploiting the high sensitivity of surface-enhanced Raman scattering combined with properly designed superhydrophobic substrates. These results pave the way for employing such advanced spectroscopic methods for quantitatively sensing the oxidative damage of nucleotides in the cell.


Assuntos
Ácidos Nucleicos , Análise Espectral Raman , Guanosina , Nucleotídeos , Estresse Oxidativo
16.
Int J Mol Sci ; 22(13)2021 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-34199004

RESUMO

Guanosine (Guo) is a nucleotide metabolite that acts as a potent neuromodulator with neurotrophic and regenerative properties in neurological disorders. Under brain ischemia or trauma, Guo is released to the extracellular milieu and its concentration substantially raises. In vitro studies on brain tissue slices or cell lines subjected to ischemic conditions demonstrated that Guo counteracts destructive events that occur during ischemic conditions, e.g., glutaminergic excitotoxicity, reactive oxygen and nitrogen species production. Moreover, Guo mitigates neuroinflammation and regulates post-translational processing. Guo asserts its neuroprotective effects via interplay with adenosine receptors, potassium channels, and excitatory amino acid transporters. Subsequently, guanosine activates several prosurvival molecular pathways including PI3K/Akt (PI3K) and MEK/ERK. Due to systemic degradation, the half-life of exogenous Guo is relatively low, thus creating difficulty regarding adequate exogenous Guo distribution. Nevertheless, in vivo studies performed on ischemic stroke rodent models provide promising results presenting a sustained decrease in infarct volume, improved neurological outcome, decrease in proinflammatory events, and stimulation of neuroregeneration through the release of neurotrophic factors. In this comprehensive review, we discuss molecular signaling related to Guo protection against brain ischemia. We present recent advances, limitations, and prospects in exogenous guanosine therapy in the context of ischemic stroke.


Assuntos
Guanosina/farmacologia , AVC Isquêmico/metabolismo , Fármacos Neuroprotetores/farmacologia , Animais , Biomarcadores , Sistema Nervoso Central/efeitos dos fármacos , Sistema Nervoso Central/metabolismo , Gerenciamento Clínico , Suscetibilidade a Doenças , Humanos , AVC Isquêmico/tratamento farmacológico , AVC Isquêmico/etiologia , AVC Isquêmico/patologia , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteína Quinase C/metabolismo , Transdução de Sinais
17.
Nat Cell Biol ; 23(7): 684-691, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34253897

RESUMO

Members of the mammalian AlkB family are known to mediate nucleic acid demethylation1,2. ALKBH7, a mammalian AlkB homologue, localizes in mitochondria and affects metabolism3, but its function and mechanism of action are unknown. Here we report an approach to site-specifically detect N1-methyladenosine (m1A), N3-methylcytidine (m3C), N1-methylguanosine (m1G) and N2,N2-dimethylguanosine (m22G) modifications simultaneously within all cellular RNAs, and discovered that human ALKBH7 demethylates m22G and m1A within mitochondrial Ile and Leu1 pre-tRNA regions, respectively, in nascent polycistronic mitochondrial RNA4-6. We further show that ALKBH7 regulates the processing and structural dynamics of polycistronic mitochondrial RNAs. Depletion of ALKBH7 leads to increased polycistronic mitochondrial RNA processing, reduced steady-state mitochondria-encoded tRNA levels and protein translation, and notably decreased mitochondrial activity. Thus, we identify ALKBH7 as an RNA demethylase that controls nascent mitochondrial RNA processing and mitochondrial activity.


Assuntos
Enzimas AlkB/metabolismo , Mitocôndrias/enzimologia , Proteínas Mitocondriais/metabolismo , Processamento Pós-Transcricional do RNA , RNA Mitocondrial/metabolismo , Adenosina/análogos & derivados , Adenosina/metabolismo , Enzimas AlkB/genética , Citidina/análogos & derivados , Citidina/metabolismo , Guanosina/análogos & derivados , Guanosina/metabolismo , Células HEK293 , Células HeLa , Células Hep G2 , Humanos , Mitocôndrias/genética , Proteínas Mitocondriais/genética , Biossíntese de Proteínas , RNA Mitocondrial/genética , RNA de Transferência/genética , RNA de Transferência/metabolismo
18.
J Org Chem ; 86(15): 9970-9978, 2021 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-34279932

RESUMO

The hierarchical self-assembly of various lipophilic guanosines exposing either a phenyl or a ferrocenyl group in the C(8) position was investigated. In a solution, all the derivatives were found to self-assemble primarily into isolated guanine (G)-quartets. In spite of the apparent similar bulkiness of the two substituents, most of the derivatives form disordered structures in the solid state, whereas a specific 8-phenyl derivative self-assembles into an unprecedented, cation-free stacked G-quartet architecture.


Assuntos
Quadruplex G , Cátions , Guanina , Guanosina
19.
Molecules ; 26(12)2021 Jun 09.
Artigo em Inglês | MEDLINE | ID: mdl-34207872

RESUMO

Five new compounds including three pairs of enantiomeric xanthine analogues, parvaxanthines D-F (1-3), two new guanosine derivatives, asponguanosines C and D (6 and 7), along with two known adenine derivatives were isolated from the insect Cyclopelta parva. Racemic 1-3 were further separated by chiral HPLC. Their absolute configurations were assigned by spectroscopic and computational methods. It is interesting that all of these isolates are natural product hybrids. Antiviral, immunosuppressive, antitumor and anti-inflammatory properties of all the isolates were evaluated.


Assuntos
Anti-Inflamatórios/farmacologia , Antineoplásicos/farmacologia , Antivirais/farmacologia , Produtos Biológicos/farmacologia , Guanosina/química , Insetos/química , Xantinas/química , Animais , Produtos Biológicos/química , Células Cultivadas , Chlorocebus aethiops , Cromatografia Líquida de Alta Pressão/métodos , Camundongos , Camundongos Endogâmicos C57BL , Estrutura Molecular , Estereoisomerismo
20.
Mol Cell Biol ; 41(9): e0030321, 2021 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-34228493

RESUMO

Germline mutations in the mismatch repair (MMR) genes MSH2, MSH6, MLH1, and PMS2 are linked to cancer of the colon and other organs, characterized by microsatellite instability and a large increase in mutation frequency. Unexpectedly, mutations in EXO1, encoding the only exonuclease genetically implicated in MMR, are not linked to familial cancer and cause a substantially weaker mutator phenotype. This difference could be explained if eukaryotic cells possessed additional exonucleases redundant with EXO1. Analysis of the MLH1 interactome identified FANCD2-associated nuclease 1 (FAN1), a novel enzyme with biochemical properties resembling EXO1. We now show that FAN1 efficiently substitutes for EXO1 in MMR assays and that this functional complementation is modulated by its interaction with MLH1. FAN1 also contributes to MMR in vivo; cells lacking both EXO1 and FAN1 have an MMR defect and display resistance to N-methyl-N-nitrosourea (MNU) and 6-thioguanine (TG). Moreover, FAN1 loss amplifies the mutational profile of EXO1-deficient cells, suggesting that the two nucleases act redundantly in the same antimutagenic pathway. However, the increased drug resistance and mutator phenotype of FAN1/EXO1-deficient cells are less prominent than those seen in cells lacking MSH6 or MLH1. Eukaryotic cells thus apparently possess additional mechanisms that compensate for the loss of EXO1.


Assuntos
Proteínas Aviárias/metabolismo , Reparo de Erro de Pareamento de DNA , Endodesoxirribonucleases/metabolismo , Exodesoxirribonucleases/metabolismo , Enzimas Multifuncionais/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Linhagem Celular , Galinhas , Endodesoxirribonucleases/química , Exodesoxirribonucleases/química , Exodesoxirribonucleases/deficiência , Exodesoxirribonucleases/genética , Guanosina/análogos & derivados , Células HEK293 , Humanos , Metilnitronitrosoguanidina , Enzimas Multifuncionais/química , Mutação/genética , Tionucleosídeos
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