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1.
Curr Protoc ; 4(2): e984, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38327099

RESUMO

A simple, reliable, and efficient method for the gram-scale chemical synthesis of pyrimidine nucleosides functionalized with C5-carboxyl, nitrile, ester, amide, or amidine, starting from unprotected uridine and cytidine, is described. The protocol involves the synthesis of 5-trifluoromethyluridine and 5-trifluoromethylcytidine with Langlois reagent (CF3 SO2 Na) in the presence of tert-butyl hydroperoxide and subsequent transformation of the CF3 group to the C5-C 'carbon substituents' under alkaline conditions. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Synthesis and characterization of 5-trifluoromethyluridine (5-CF3 U) and 5-trifluoromethylcytidine (5-CF3 C) Basic Protocol 2: Conversion of 5-CF3 U and 5-CF3 C to several C5-substituted ribonucleosides.


Assuntos
Química Orgânica , Nucleosídeos de Pirimidina , Citidina/análogos & derivados , Nucleosídeos de Pirimidina/síntese química , Nucleosídeos de Pirimidina/química , Ribonucleosídeos/química , Uridina/análogos & derivados , Química Orgânica/métodos
2.
Int J Mol Sci ; 25(4)2024 Feb 17.
Artigo em Inglês | MEDLINE | ID: mdl-38397036

RESUMO

Nicotinamide (NA) derivatives play crucial roles in various biological processes, such as inflammation, regulation of the cell cycle, and DNA repair. Recently, we proposed that 4-pyridone-3-carboxamide-1-ß-D-ribonucleoside (4PYR), an unusual derivative of NA, could be classified as an oncometabolite in bladder, breast, and lung cancer. In this study, we investigated the relations between NA metabolism and the progression, recurrence, metastasis, and survival of patients diagnosed with different histological subtypes of renal cell carcinoma (RCC). We identified alterations in plasma NA metabolism, particularly in the clear cell RCC (ccRCC) subtype, compared to papillary RCC, chromophobe RCC, and oncocytoma. Patients with ccRCC also exhibited larger tumor sizes and elevated levels of diagnostic serum biomarkers, such as hsCRP concentration and ALP activity, which were positively correlated with the plasma 4PYR. Notably, 4PYR levels were elevated in advanced stages of ccRCC cancer and were associated with a highly aggressive phenotype of ccRCC. Additionally, elevated concentrations of 4PYR were related to a higher likelihood of mortality, recurrence, and particularly metastasis in ccRCC. These findings are consistent with other studies, suggesting that NA metabolism is accelerated in RCC, leading to abnormal concentrations of 4PYR. This supports the concept of 4PYR as an oncometabolite and a potential prognostic factor in the ccRCC subtype.


Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Piridonas , Ribonucleosídeos , Humanos , Nucleosídeos/metabolismo , Niacinamida
3.
Molecules ; 29(3)2024 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-38338431

RESUMO

In this article, we present the synthesis and the optical properties of three original molecules as potential fluorescent ribonucleoside analogues incorporating a 1,6-naphthyridin-7(6H)-one scaffold as a fluorescent nucleobase and a 1,2,3-triazole as a linkage. The nucleosides were prepared via a Cu alkyne-azide cycloaddition (CuAAC) reaction between a ribofuranosyl azide and a 4-ethynylpyridine partner. Construction of substituted 1,6-naphthyridin-7(6H)-ones was achieved through two additional steps. Optical property studies were investigated on nucleoside analogues. Powerful fluorescence properties have been evidenced with a remarkable change of emissivity depending on the polarity of the solvent, making these molecules suitable as a new class of artificial fluorescent nucleosides for investigating enzyme binding sites as well as probing nucleic acids. In addition, we are convinced that such analogues could be of great interest in the search for new antiviral or antitumoral drugs based on nucleosides.


Assuntos
Nucleosídeos , Ribonucleosídeos , Nucleosídeos/química , Azidas/química , Ribonucleosídeos/química , Corantes
4.
J Infect Dis ; 229(2): 413-421, 2024 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-37506264

RESUMO

BACKGROUND: This drug resistance analysis of a randomized trial includes 234 patients receiving maribavir and 116 receiving investigator-assigned standard therapy (IAT), where 56% and 24%, respectively, cleared cytomegalovirus DNA at week 8 (treatment responders). METHODS: Baseline and posttreatment plasma samples were tested for mutations conferring drug resistance in viral genes UL97, UL54, and UL27. RESULTS: At baseline, genotypic testing revealed resistance to ganciclovir, foscarnet, or cidofovir in 56% of patients receiving maribavir and 68% receiving IAT, including 9 newly phenotyped mutations. Among them, 63% (maribavir) and 21% (IAT) were treatment responders. Detected baseline maribavir resistance mutations were UL27 L193F (n = 1) and UL97 F342Y (n = 3). Posttreatment, emergent maribavir resistance mutations were detected in 60 (26%) of those randomized to maribavir, including 49 (48%) of 103 nonresponders and 25 (86%) of the 29 nonresponders where viral DNA initially cleared then rebounded while on maribavir. The most common maribavir resistance mutations were UL97 T409M (n = 34), H411Y (n = 26), and C480F (n = 21), first detected 26 to 130 (median 56) days after starting maribavir. CONCLUSIONS: Baseline maribavir resistance was rare. Drug resistance to standard cytomegalovirus antivirals did not preclude treatment response to maribavir. Rebound in plasma cytomegalovirus DNA while on maribavir strongly suggests emerging drug resistance. CLINICAL TRIALS REGISTRATION: NCT02931539.


Assuntos
Infecções por Citomegalovirus , Diclororribofuranosilbenzimidazol , Ribonucleosídeos , Humanos , Antivirais/uso terapêutico , Antivirais/farmacologia , Benzimidazóis/uso terapêutico , Citomegalovirus/genética , Infecções por Citomegalovirus/tratamento farmacológico , Diclororribofuranosilbenzimidazol/análogos & derivados , DNA , Farmacorresistência Viral/genética , Ganciclovir/uso terapêutico , Mutação , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Ribonucleosídeos/uso terapêutico , Transplantados
5.
Nucleic Acids Res ; 52(3): 1207-1225, 2024 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-38117983

RESUMO

Abundant ribonucleoside-triphosphate (rNTP) incorporation into DNA by DNA polymerases in the form of ribonucleoside monophosphates (rNMPs) is a widespread phenomenon in nature, resulting in DNA-structural change and genome instability. The rNMP distribution, characteristics, hotspots and association with DNA metabolic processes in human mitochondrial DNA (hmtDNA) remain mostly unknown. Here, we utilize the ribose-seq technique to capture embedded rNMPs in hmtDNA of six different cell types. In most cell types, the rNMPs are preferentially embedded on the light strand of hmtDNA with a strong bias towards rCMPs; while in the liver-tissue cells, the rNMPs are predominately found on the heavy strand. We uncover common rNMP hotspots and conserved rNMP-enriched zones across the entire hmtDNA, including in the control region, which links the rNMP presence to the frequent hmtDNA replication-failure events. We show a strong correlation between coding-sequence size and rNMP-embedment frequency per nucleotide on the non-template, light strand in all cell types, supporting the presence of transient RNA-DNA hybrids preceding light-strand replication. Moreover, we detect rNMP-embedment patterns that are only partly conserved across the different cell types and are distinct from those found in yeast mtDNA. The study opens new research directions to understand the biology of hmtDNA and genomic rNMPs.


Assuntos
Replicação do DNA , Genoma Mitocondrial , Ribonucleosídeos , Humanos , DNA Mitocondrial/genética , DNA Mitocondrial/metabolismo , Ribonucleosídeos/metabolismo , Ribonucleotídeos/genética , Ribonucleotídeos/metabolismo
6.
J Chromatogr A ; 1715: 464561, 2024 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-38154259

RESUMO

Ribonucleoside hydrolase C (RihC, EC 3.2.2.1-3.2.2.3, 3.2.2.7, 3.2.2.8) belongs to the family of ribonucleoside hydrolases that catalyze the cleavage of both purine and pyrimidine ribonucleosides to nitrogenous bases and ribose. Its most efficient reaction is the cleavage of uridine with the highest reaction rate. The reaction cannot be detected by a simple spectrophotometric method because of the same absorption maximum for the substrate and reaction product or requires time- and labor-consuming sample preparation for ribose. Reversed-phase HPLC is currently used to register enzymatic activity, where the time of one chromatographic run takes about 10 min. Since a large number of analyses is required to measure the kinetics of an enzymatic reaction, the total time is significant. In this work, we obtained new recombinant RihC from Limosilactobacillus reuteri by gene cloning and expression in E.coli cells. We proposed a new approach for determining the enzymatic activity of the new RihC using hydrophilic interaction liquid chromatography (HILIC). The novel column was developed for this procedure providing the determination of uracil and uridine with high efficiency and retention times of 0.9 and 1.7 min, respectively. Kinetic parameters for RihC uridine cleavage were determined. The proposed approach provided significant rapidity for measurement of the enzyme kinetics being 5 times faster as compared to reversed-phase HPLC.


Assuntos
Hidrolases de Éster Carboxílico , Ribonucleosídeos , Ribose , Ribonucleosídeos/análise , Cromatografia Líquida , Uridina , Escherichia coli/genética , Escherichia coli/metabolismo , Interações Hidrofóbicas e Hidrofílicas
7.
Org Lett ; 25(50): 9002-9007, 2023 12 22.
Artigo em Inglês | MEDLINE | ID: mdl-38051027

RESUMO

Nucleoside analogues are effective antiviral agents, and the continuous emergence of pathogenic viruses demands the development of novel and structurally diverse analogues. Here, we present the design and synthesis of novel nucleoside analogues with a carbobicyclic core, which mimics the conformation of natural ribonucleosides. Employing a divergent synthetic route featuring an intermolecular Diels-Alder reaction, we successfully synthesized carbobicyclic nucleoside analogues with high antiviral efficacy against respiratory syncytial virus.


Assuntos
Nucleosídeos , Ribonucleosídeos , Antivirais/farmacologia , Conformação Molecular
8.
Carbohydr Res ; 534: 108981, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37992558

RESUMO

An easy and efficient large-scale synthesis of 1, 2,-di-O-acetyl-5-O-benzoyl-3-O-methyl-d-ribofuranose (8) was accomplished from commercial 1,2:5,6-di-O-isopropylidene-α-d-allofuranose in 7-steps and 30 % overall yield. The utility of protected 8 was demonstrated via synthesis of 9-(3'-O-methyl-ß-d-ribofuranosyl)-6-chloropurine (21) and six other nucleoside analogues in good yields. A library of five novel base modified nucleosides were generated starting from purine nucleoside 21 via functional group manipulations. The 3'-O-modified nucleosides are known to act as chain terminator exerting antiviral activity. The synthesis strategy described herein offers direct access to 3'-O-alkylated nucleosides with wide range of applications, including cap analogues for mRNA vaccine production. This protocol provides a route to exclusive synthesis of 3'-O-alkylated nucleosides, devoid of isomeric 2'-O-alkylated products essential for both therapeutic and biological research.


Assuntos
Ribonucleosídeos , Nucleosídeos
9.
J Am Chem Soc ; 145(43): 23781-23793, 2023 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-37856825

RESUMO

Among the many prebiotic phosphorylation chemistries investigated, diamidophosphate (DAP) has shown promising potential for nucleoside phosphorylation. Herein, we show that DAP's phosphorylation capability is enhanced significantly (up to 90%) in wet-dry cycles by a range of prebiotically plausible pHs (6-10) and temperatures (up to 80 °C) in the presence of additives such as formamide, cyanamide, urea, guanidine, 2-aminoimidazole, and hydantoin. For ribonucleosides, the main products are the 2',3'-cyclic phosphates along with the corresponding 2'- and 3'-phosphates, while deoxyribonucleosides form 5'- and 3'-phosphates, the ratios of which are affected by cycles and the presence and nature of the additives. A simple change of temperature to 80 °C with additives leads to higher conversion yields (≈80-90%) with an increased level of 5'-phosphorylation (≈40-49%). This demonstration of enhancing and controlling the regioselectivity of DAP-mediated phosphorylation by a range of additives and conditions potentiates transitioning to the search for more efficient catalysts, enabling regiospecific phosphorylations and oligonucleotide formation in the same milieu and setting.


Assuntos
Nucleosídeos , Ribonucleosídeos , Fosforilação , Fosfatos
10.
Biomolecules ; 13(9)2023 09 12.
Artigo em Inglês | MEDLINE | ID: mdl-37759775

RESUMO

Ribonucleoside hydrolases are enzymes that catalyze the cleavage of ribonucleosides to nitrogenous bases and ribose. These enzymes are found in many organisms: bacteria, archaea, protozoa, metazoans, yeasts, fungi and plants. Despite the simple reaction catalyzed by these enzymes, their physiological role in most organisms remains unclear. In this review, we compare the structure, kinetic parameters, physiological role, and potential applications of different types of ribonucleoside hydrolases discovered and isolated from different organisms.


Assuntos
Hidrolases , Ribonucleosídeos , Fungos , Leveduras
11.
Chembiochem ; 24(22): e202300544, 2023 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-37666794

RESUMO

Given the importance of mRNA with 5'-cap, easy access to RNA substrates with different 7m G caps, of high quality and in large quantities is essential to elucidate the roles of RNA and the regulation of underlying processes. In addition to existing synthetic routes to 5'-cap RNA based on enzymatic, chemical or chemo-enzymatic methods, we present here an all-chemical method for synthetic RNA capping. The novelty of this study lies in the fact that the capping reaction is performed on solid-support after automated RNA assembly using commercial 2'-O-propionyloxymethyl ribonucleoside phosphoramidites, which enable final RNA deprotection under mild conditions while preserving both 7m G-cap and RNA integrity. The capping reaction is efficiently carried out between a 5'-phosphoroimidazolide RNA anchored on the support and 7m GDP in DMF in the presence of zinc chloride. Substantial amounts of 7m G-cap RNA (from 1 to 28 nucleotides in length and of any sequence with or without internal methylations) containing various cap structures (7m GpppA, 7m GpppAm , 7m Gpppm6 A, 7m Gpppm6 Am , 7m GpppG, 7m GpppGm ) were obtained with high purity after IEX-HPLC purification. This capping method using solid-phase chemistry is convenient to perform and provides access to valuable RNA substrates as useful research tools to unravel specific issues regarding cap-related processes.


Assuntos
Metiltransferases , Ribonucleosídeos , Metiltransferases/metabolismo , Capuzes de RNA , Metilação , RNA Mensageiro
12.
RNA ; 29(11): 1818-1836, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37582618

RESUMO

The conserved family of RNA-binding proteins (RBPs), IGF2BPs, plays an essential role in posttranscriptional regulation controlling mRNA stability, localization, and translation. Mammalian cells express three isoforms of IGF2BPs: IGF2BP1-3. IGF2BP3 is highly overexpressed in cancer cells, and its expression correlates with a poor prognosis in various tumors. Therefore, revealing its target RNAs with high specificity in healthy tissues and in cancer cells is of crucial importance. Photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) identifies the binding sites of RBPs on their target RNAs at nucleotide resolution in a transcriptome-wide manner. Here, we optimized the PAR-CLIP protocol to study RNA targets of endogenous IGF2BP3 in a human colorectal carcinoma cell line. To this end, we first established an immunoprecipitation protocol to obtain highly pure endogenous IGF2BP3-RNA complexes. Second, we modified the protocol to use highly sensitive infrared (IR) fluorescent dyes instead of radioactive probes to visualize IGF2BP3-crosslinked RNAs. We named the modified method "IR-PAR-CLIP." Third, we compared RNase cleavage conditions and found that sequence preferences of the RNases impact the number of the identified IGF2BP3 targets and introduce a systematic bias in the identified RNA motifs. Fourth, we adapted the single adapter circular ligation approach to increase the efficiency in library preparation. The optimized IR-PAR-CLIP protocol revealed novel RNA targets of IGF2BP3 in a human colorectal carcinoma cell line. We anticipate that our IR-PAR-CLIP approach provides a framework for studies of other RBPs.


Assuntos
Neoplasias do Colo , Neoplasias Colorretais , Ribonucleosídeos , Animais , Humanos , RNA/genética , Imunoprecipitação , Proteínas de Ligação a RNA/metabolismo , Sítios de Ligação , Ribonucleases/metabolismo , Ribonucleosídeos/química , Mamíferos/genética
13.
Methods Mol Biol ; 2712: 29-43, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37578694

RESUMO

Ferroptosis is a regulatory cell death process that is accompanied by large amounts of iron ion accumulation and lipid peroxidation. Photoactivated ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) is a method used to identify the binding sites of RNA-binding proteins (RBPs) on target RNAs with high resolution at the nucleotide level. By inserting photosensitive ribonucleoside analogs into new RNA transcripts of living cells, characteristic mutations can be generated during reverse transcription and be used to accurately locate the crosslinking position of RNAs and RBPs. The use of PAR-CLIP to detect interactions and determine precise crosslinking sites between RNAs and RBPs, or to search for RNAs upstream or downstream of ferroptosis pathways genes through known proteins, can help to clarify and verify the occurrence and regulation mechanisms of the various signaling pathways of ferroptosis. Furthermore, it may reveal new targets for ferroptosis detection and improve the treatment efficiency of ferroptosis-related diseases such as cancer and neurodegenerative diseases. Here, we introduce a specific PAR-CLIP protocol for monitoring the ferroptosis process.


Assuntos
Ferroptose , Ribonucleosídeos , RNA/genética , Imunoprecipitação , Proteínas de Ligação a RNA/metabolismo , Sítios de Ligação , Ribonucleosídeos/química
14.
EMBO J ; 42(18): e114990, 2023 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-37548337

RESUMO

The building blocks for RNA and DNA are made in the cytosol, meaning mitochondria depend on the import and salvage of ribonucleoside triphosphates (rNTPs) and deoxyribonucleoside triphosphates (dNTPs) for the synthesis of their own genetic material. While extensive research has focused on mitochondrial dNTP homeostasis due to its defects being associated with various mitochondrial DNA (mtDNA) depletion and deletion syndromes, the investigation of mitochondrial rNTP homeostasis has received relatively little attention. In this issue of the EMBO Journal, Grotehans et al provide compelling evidence of a major role for NME6, a mitochondrial nucleoside diphosphate kinase, in the conversion of pyrimidine ribonucleoside diphosphates into the corresponding triphosphates. These data also suggest a significant physiological role for NME6, as its absence results in the depletion of mitochondrial transcripts and destabilization of the electron transport chain (Grotehans et al, 2023).


Assuntos
Ribonucleosídeos , Ribonucleotídeos , Ribonucleotídeos/genética , Mitocôndrias/genética , DNA Mitocondrial/genética , Nucleotídeos
15.
Talanta ; 263: 124697, 2023 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-37262985

RESUMO

Although next-generation sequencing technology has been used to delineate RNA modifications in recent years, the paucity of appropriate converting reactions or specific antibodies impedes the accurate characterization and quantification of numerous RNA modifications, especially when these modifications demonstrate wide variations across developmental stages and cell types. In this study, we developed a high-throughput analytical platform coupling ultra-performance liquid chromatograph (UPLC) with complementary mass spectrometry (MS) to identify and quantify RNA modifications in both synthetic and biological samples. Sixty-four types of RNA modifications, including positional isomers and hypermodified ribonucleosides, were successfully monitored within a 16-min single run of UPLC-MS. Two independent methods to cross-validate the purity of RNA extracted from Caenorhabditis elegans (C. elegans) were developed using the coexisting C. elegans and Escherichia coli (E. coli) as a surveillance system. To test the validity of the method, we investigated the RNA modification landscape of three model organisms, C. elegans, E. coli, and Arabidopsis thaliana (A. thaliana). Both the identity and molarity of modified ribonucleosides markedly varied among the species. Moreover, our platform is not only useful for exploring the dynamics of RNA modifications in response to environmental cues (e.g., cold shock) but can also help with the identification of RNA-modifying enzymes in genetic studies. Cumulatively, our method presents a novel platform for the comprehensive analysis of RNA modifications, which will be of benefit to both analytical chemists involved in biomarker discovery and biologists conducting functional studies of RNA modifications.


Assuntos
Arabidopsis , Ribonucleosídeos , Animais , Cromatografia Líquida/métodos , Caenorhabditis elegans/metabolismo , Escherichia coli/metabolismo , Espectrometria de Massas em Tandem/métodos , Cromatografia Líquida de Alta Pressão/métodos , RNA/genética , RNA/química , Ribonucleosídeos/química , Arabidopsis/genética , Controle de Qualidade
16.
J Am Chem Soc ; 145(21): 11611-11621, 2023 05 31.
Artigo em Inglês | MEDLINE | ID: mdl-37192367

RESUMO

Nucleosides are essential cornerstones of life, and nucleoside derivatives and synthetic analogues have important biomedical applications. Correspondingly, production of non-canonical nucleoside derivatives in animal model systems is of particular interest. Here, we report the discovery of diverse glucose-based nucleosides in Caenorhabditis elegans and related nematodes. Using a mass spectrometric screen based on all-ion fragmentation in combination with total synthesis, we show that C. elegans selectively glucosylates a series of modified purines but not the canonical purine and pyrimidine bases. Analogous to ribonucleosides, the resulting gluconucleosides exist as phosphorylated and non-phosphorylated forms. The phosphorylated gluconucleosides can be additionally decorated with diverse acyl moieties from amino acid catabolism. Syntheses of representative variants, facilitated by a novel 2'-O- to 3'-O-dibenzyl phosphoryl transesterification reaction, demonstrated selective incorporation of different nucleobases and acyl moieties. Using stable-isotope labeling, we further show that gluconucleosides incorporate modified nucleobases derived from RNA and possibly DNA breakdown, revealing extensive recycling of oligonucleotide catabolites. Gluconucleosides are conserved in other nematodes, and biosynthesis of specific subsets is increased in germline mutants and during aging. Bioassays indicate that gluconucleosides may function in stress response pathways.


Assuntos
Nucleosídeos , Ribonucleosídeos , Animais , Caenorhabditis elegans , Oligonucleotídeos
17.
J Nutr ; 153(5): 1636-1645, 2023 05.
Artigo em Inglês | MEDLINE | ID: mdl-36907444

RESUMO

BACKGROUND: Ribonucleosides and RNA are an underappreciated nutrient group essential during Drosophila larval development and growth. Detection of these nutrients requires at least one of the 6 closely related taste receptors encoded by the Gr28 genes, one of the most conserved insect taste receptor subfamilies. OBJECTIVES: We investigated whether blow fly larvae and mosquito larvae, which shared the last ancestor with Drosophila about 65 and 260 million years ago, respectively, can taste RNA and ribose. We also tested whether the Gr28 homologous genes of the mosquitoes Aedes aegypti and Anopheles gambiae can sense these nutrients when expressed in transgenic Drosophila larvae. METHODS: Taste preference in blow flies was examined by adapting a 2-choice preference assay that has been well-established for Drosophila larvae. For the mosquito Aedes aegypti, we developed a new 2-choice preference assay that accommodates the aquatic environment of these insect larvae. Finally, we identified Gr28 homologs in these species and expressed them in Drosophila melanogaster to determine their potential function as RNA receptors. RESULTS: Larvae of the blow fly Cochliomyia macellaria and Lucilia cuprina are strongly attracted to RNA (0.5 mg/mL) in the 2-choice feeding assays (P < 0.05). Similarly, the mosquito Aedes aegypti larvae showed a strong preference for RNA (2.5 mg/mL) in an aquatic 2-choice feeding assay. Moreover, when Gr28 homologs of Aedes or Anopheles mosquitoes are expressed in appetitive taste neurons of Drosophila melanogaster larvae lacking their Gr28 genes, preference for RNA (0.5 mg/mL) and ribose (0.1 M) is rescued (P < 0.05). CONCLUSIONS: The appetitive taste for RNA and ribonucleosides in insects emerged about 260 million years ago, the time mosquitoes and fruit flies diverged from their last common ancestor. Like sugar receptors, receptors for RNA have been highly conserved during insect evolution, suggesting that RNA is a critical nutrient for fast-growing insect larvae.


Assuntos
Aedes , Ribonucleosídeos , Animais , RNA/genética , Drosophila melanogaster/genética , Paladar/fisiologia , Ribose , Drosophila/genética , Larva/genética , Aedes/genética
18.
Biochemistry ; 62(8): 1376-1387, 2023 04 18.
Artigo em Inglês | MEDLINE | ID: mdl-36972568

RESUMO

Adenosine deaminases acting on RNA (ADARs) are RNA editing enzymes that catalyze the hydrolytic deamination of adenosine (A) to inosine (I) in dsRNA. In humans, two catalytically active ADARs, ADAR1 and ADAR2, perform this A-to-I editing event. The growing field of nucleotide base editing has highlighted ADARs as promising therapeutic agents while multiple studies have also identified ADAR1's role in cancer progression. However, the potential for site-directed RNA editing as well as the rational design of inhibitors is being hindered by the lack of detailed molecular understanding of RNA recognition by ADAR1. Here, we designed short RNA duplexes containing the nucleoside analog, 8-azanebularine (8-azaN), to gain insight into molecular recognition by the human ADAR1 catalytic domain. From gel shift and in vitro deamination experiments, we validate ADAR1 catalytic domain's duplex secondary structure requirement and present a minimum duplex length for binding (14 bp, with 5 bp 5' and 8 bp 3' to editing site). These findings concur with predicted RNA-binding contacts from a previous structural model of the ADAR1 catalytic domain. Finally, we establish that neither 8-azaN as a free nucleoside nor a ssRNA bearing 8-azaN inhibits ADAR1 and demonstrate that the 8-azaN-modified RNA duplexes selectively inhibit ADAR1 and not the closely related ADAR2 enzyme.


Assuntos
Ribonucleosídeos , Humanos , Nucleosídeos de Purina , RNA de Cadeia Dupla , Adenosina , Adenosina Desaminase/metabolismo
19.
Astrobiology ; 23(5): 605-615, 2023 05.
Artigo em Inglês | MEDLINE | ID: mdl-36862128

RESUMO

Boron, as borate (or boric acid), is known as a mediator of the synthesis of ribose, ribonucleosides, and ribonucleotides (precursors of RNA) under plausible prebiotic conditions. With regard to these phenomena, the potential participation of this chemical element (as a constituent of minerals or hydrogels) for the emergence of prebiological homochirality is considered. This hypothesis is based on characteristics of crystalline surfaces as well as solubility of some minerals of boron in water or specific features of hydrogels with ester bonds from reaction of ribonucleosides and borate.


Assuntos
Boro , Ribonucleosídeos , Humanos , Boro/química , Boratos/química , Minerais/química , Ribonucleosídeos/química , Hidrogéis
20.
Life Sci ; 318: 121462, 2023 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-36736767

RESUMO

AIMS: Cardiomyopathy is a diabetic comorbidity with few molecular targets. To address this, we evaluated transfer RNA (tRNA) modifications in the diabetic heart because tRNA modifications have been implicated in diabetic etiologies. MAIN METHODS: tRNA was isolated from aorta, apex, and atrial tissue of healthy and diabetic murine hearts and related hyperglycemic cell models. tRNA modifications and canonical ribonucleosides were quantified by liquid-chromatography tandem mass spectrometry (LC-MS/MS) using stable isotope dilution. Correlations between ribonucleosides and diabetic comorbidity pathology were assessed using statistical analyses. KEY FINDINGS: Total tRNA ribonucleoside levels were analyzed from cell types and healthy and diabetic murine heart tissue. Each heart structure had characteristic ribonucleoside profiles and quantities. Several ribonucleosides were observed as significantly different in hyperglycemic cells and diabetic tissues. In hyperglycemic models, ribonucleosides N4-acetylcytidine (ac4C), 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U), 5-methylcytidine (m5C), and N1-methylguanosine (m1G) were anomalous. Specific tRNA modifications known to be on murine tRNAIni(CAU) were higher in diabetic heart tissue which suggests that tRNA modifications could be regulating translation in diabetes. SIGNIFICANCE: We identified tRNA ribonucleosides and tRNA species associated with hyperglycemia and diabetic etiology.


Assuntos
Diabetes Mellitus , Ribonucleosídeos , Animais , Camundongos , Ribonucleosídeos/análise , Ribonucleosídeos/química , Ribonucleosídeos/metabolismo , Cromatografia Líquida/métodos , Espectrometria de Massas em Tandem , RNA de Transferência/genética , Mamíferos/metabolismo
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