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A theoretical thermodynamic study was conducted to investigate the antioxidant activity and mechanism of 1,3,4-oxadiazol-2-ylthieno[2,3-d]pyrimidin-4-amine derivatives (OTP) using a Density Functional Theory (DFT) approach. The study assessed how solvent environments influence the antioxidant properties of these derivatives. With the increasing prevalence of diseases linked to oxidative stress, such as cancer and cardiovascular diseases, antioxidants are crucial in mitigating the damage caused by free radicals. Previous research has demonstrated the remarkable scavenging abilities of 1,3,4-oxadiazole derivatives, prompting this investigation into their potential using computational methods. DFT calculations were employed to analyze key parameters, including bond dissociation enthalpy (BDE), ionization potential (IP), proton dissociation enthalpy (PDE), and electron transfer enthalpy (ETE), to delineate the antioxidant mechanisms of these compounds. Our findings indicate that specific electron-donating groups such as amine on the phenyl rings significantly enhance the antioxidant activities of these derivatives. The study also integrates global and local reactivity descriptors, such as Fukui functions and HOMO-LUMO energies, to predict the stability and reactivity of these molecules, providing insights into their potential as effective synthetic antioxidants in pharmaceutical applications.
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The P-glycoprotein (ABCB1)-mediated multidrug resistance (MDR) has emerged as a significant impediment to the efficacy of cancer chemotherapy in clinical therapy, which could promote the development of effective agents for MDR reversal. In this work, we reported the exploration of novel pyrazolo [1,5-a]pyrimidine derivatives as potent reversal agents capable of enhancing the sensitivity of ABCB1-mediated MDR MCF-7/ADR cells to paclitaxel (PTX). Among them, compound 16q remarkably increased the sensitivity of MCF-7/ADR cells to PTX at 5 µM (IC50 = 27.00 nM, RF = 247.40) and 10 µM (IC50 = 10.07 nM, RF = 663.44). Compound 16q could effectively bind and stabilize ABCB1, and does not affect the expression and subcellular localization of ABCB1 in MCF-7/ADR cells. Compound 16q inhibited the function of ABCB1, thereby increasing PTX accumulation, and interrupting the accumulation and efflux of the ABCB1-mediated Rh123, thus resulting in exhibiting good reversal effects. In addition, due to the potent reversal effects of compound 16q, the abilities of PTX to inhibit tubulin depolymerization, and induce cell cycle arrest and apoptosis in MCF-7/ADR cells under low-dose conditions were restored. These results indicate that compound 16q might be a promising potent reversal agent capable of revising ABCB1-mediated MDR, and pyrazolo [1,5-a]pyrimidine might represent a novel scaffold for the discovery of new ABCB1-mediated MDR reversal agents.
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Glioblastoma (GBM) is a brain tumor characterized by its aggressive and invasive properties. It is found that STAT3 is abnormally activated in GBM, and inhibiting STAT3 signaling can effectively suppress tumor progression. In this study, novel pyrimidine compounds, BY4003 and BY4008, were synthesized to target the JAK3/STAT3 signaling pathway, and their therapeutic efficacy and mechanisms of action were evaluated and compared with Tofacitinib in U251, A172, LN428 and patient-derived glioblastoma cells. The ADP-Glo™ kinase assay was utilized to assessed the inhibitory effects of BY4003 and BY4008 on JAK3, a crucial member of the JAK family. The results showed that both compounds significantly inhibited JAK3 enzyme activity, with IC50 values in the nanomolar range. The antiproliferative effects of BY4003, BY4008, and Tofacitinib on GBM and patient-derived glioblastoma cells were evaluated by MTT and H&E assays. The impact of BY4003 and BY4008 on GBM cell migration and apoptosis induction was assessed through wound healing, transwell, and TUNEL assays. STAT3-regulated protein expression and relative mRNA levels were analyzed by western blotting, immunocytochemistry, immunofluorescence, and qRT-PCR. It was found that BY4003, BY4008 and Tofacitinib could inhibit U251, A172, LN428 and patient-derived glioblastoma cells growth and proliferation. Results showed decreased expression of STAT3-associated proteins, including p-STAT3, CyclinD1, and Bcl-2, and increased expression of Bax, a pro-apoptotic protein, as well as significant down-regulation of STAT3 and STAT3-related genes. These findings suggested that BY4003 and BY4008 could inhibit GBM growth by suppressing the JAK3/STAT3 signaling pathway, providing valuable insights into the therapeutic development of GBM.
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The rapid emergence of drug resistance severely reduces the clinical response of human immunodeficiency virus-1 (HIV-1) to non-nucleoside reverse transcriptase inhibitors (NNRTIs). Herein, a series of 2,4,6-trisubstituted pyrimidine derivatives was designed and synthesized, with the aim to identify novel anti-HIV-1 agents with improved drug resistance profiles. The antiviral activity results demonstrated that all compounds showed excellent potency to wild-type (WT) HIV-1 strain (EC50 = 3.61-15.5 nM). Moreover, 13c was proved to be the most potent inhibitor against the whole tested viral panel, with EC50 ranging from 4.68 to 229 nM. In addition, 13c yielded moderate HIV-1 RT inhibition with IC50 value of 0.231 µM, which demonstrated it was a classical NNRTI. Molecular docking was further conducted to illustrate its binding mode with HIV-1 RT. These encouraging results indicated that 13c can be used as a lead compound for further study.
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In this study 5-((2-((3-methoxy benzylidene)-amino)-phenyl)-diazenyl)-4,6-diphenyl pyrimidine-2(5H)-thione was synthesized. The pharmacological applications of pyrimidine analogs are restricted due to their poor pharmacokinetic properties. As a solution, a microbial exopolysaccharide (curdlan gum) was used to synthesize folic acid-conjugated pyrimidine-2(5H)-thione-encapsulated curdlan gum-PEGamine nanoparticles (FA-Py-CG-PEGamine NPs). The results of physicochemical properties revealed that the fabricated FA-Py-CG-PEGamine NPs were between 100 and 400 nm in size with a majorly spherical shaped, crystalline nature, and the encapsulation efficiency and loading capacity were 79.04 ± 0.79 %, and 8.12 ± 0.39 % respectively. The drug release rate was significantly higher at pH 5.4 (80.14 ± 0.79 %) compared to pH 7.2. The cytotoxic potential of FA-Py-CG-PEGamine NPs against MCF-7 cells potentially reduced the number of cells after 24 h with 42.27 µg × mL-1 as IC50 value. The higher intracellular accumulation of pyrimidine-2(5H)-thione in MCF-7 cells leads to apoptosis, observed by AO/EBr staining and flow cytometry analysis. The highest pyrimidine-2(5H)-thione internalization in MCF-7 cells may be due to folate conjugated on the surface of curdlan gum nanoparticles. Further, internalized pyrimidine-2(5H)-thione increases the intracellular ROS level, leading to apoptosis and inducing the decalin in mitochondrial membrane potential. These outcomes demonstrated that the FA-Py-CG-PEGamine NPs were specificity-targeting folate receptors on the plasma membranes of MCF-7 Cells.
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Eukaryotic NMEs/NDP kinases are a family of 10 multifunctional proteins that occur in different cellular compartments and interact with various cellular components (proteins, membranes, and DNA). In contrast to the well-studied Group I NMEs (NME1-4), little is known about the more divergent Group II NMEs (NME5-9). Three recent publications now shed new light on NME6. First, NME6 is a third mitochondrial NME, largely localized in the matrix space, associated with the mitochondrial inner membrane. Second, while its monomeric form is inactive, NME6 gains NDP kinase activity through interaction with mitochondrial RCC1L. This challenges the current notion that mammalian NMEs require the formation of hexamers to become active. The formation of complexes between NME6 and RCC1L, likely heterodimers, seemingly obviates the necessity for hexamer formation, stabilizing a NDP kinase-competent conformation. Third, NME6 is involved in mitochondrial gene maintenance and expression by providing (d)NTPs for replication and transcription (in particular the pyrimidine nucleotides) and by a less characterized mechanism that supports mitoribosome function. This review offers an overview of NME evolution and structure and highlights the new insight into NME6. The new findings position NME6 as the most comprehensively studied protein in NME Group II and may even suggest it as a new paradigm for related family members.
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Mitocôndrias , Humanos , Animais , Mitocôndrias/metabolismo , Proteínas Mitocondriais/metabolismo , Proteínas Mitocondriais/genética , Nucleosídeo NM23 Difosfato Quinases/metabolismo , Nucleosídeo NM23 Difosfato Quinases/genética , Nucleosídeo Difosfato Quinase D/metabolismo , Nucleosídeo Difosfato Quinase D/genéticaRESUMO
Rho-associated coiled-coil kinase (ROCK) is involved in multiple cellular activities regulating the actin cytoskeleton, such as cell morphology, adhesion, and migration. The inhibition of ROCK is a feasible strategy to suppress breast cancer metastasis. Herein, based on Belumosudil, a series of pyrazolo[1,5-a]pyrimidine derivatives as selective ROCK2 inhibitors were designed and synthesized. Through systematic investigation of SARs, the piperazine analog 7u was identified with optimum ROCK2 inhibitory activity (IC50 = 36.8 nM) and excellent selectivity over the isoform protein ROCK1 (>250-fold). Intriguingly, upon treatment with 7u, the arrangement of the MDA-MB-231 cytoskeleton was affected accompanied by the alteration of morphology. Furthermore, cell scratch and transwell assays indicated that 7u inhibited MDA-MB-231 cell migration and invasion in a dose-dependent manner. Ultimately, the binding model of 7u with ROCK2 well accounted for the superior activities of 7u as a promising ROCK2 inhibitor with the potential application in breast cancer metastasis treatment.
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A series of pyrimidine-2,4-diamine analogues were designed and synthesized. Their anticancer activity and the underlying mechanism against colorectal cancer (CRC) HCT116 cells and non-small cell lung cancer (NSCLC) A549 cells were investigated. The results demonstrated that the active compound Y18 significantly inhibited cancer cell proliferation by inducing robust cell cycle arrest and cell senescence through the persistence of DNA damage. Additionally, Y18 exhibited significant inhibitory effects on the adhesion, migration and invasion of cancer cells in vitro. Mechanistically, Y18 achieved these anticancer activities by suppressing GTSE1 transcription and expression. Y18 also effectively inhibited tumor growth in vivo with minimal side effects. Furthermore, Y18 exhibited a suitable half-life and oral bioavailability (16.27%), with limited inhibitory activity on CYP isoforms. Taken together, these results suggested that Y18 could be a potential chemotherapeutic drug for cancer treatment, particularly in cases of GTSE1 overexpressed cancers.
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Molnupiravir (MO) is a pyrimidine nucleoside anti-SARS-CoV-2 drug. MO treatment could cause mild liver injury. However, the underlying mechanism of MO-induced liver injury and the metabolic pathway of MO in vivo are unclear. In this study, metabolomics analysis and molecular biology methods were used to explore these issues. Through metabolomics analysis, it was found that the homeostasis of pyrimidine, purine, lysophosphatidylcholine (LPC), and amino acids in mice was destroyed after MO treatment. A total of 80 changed metabolites were detected. Among these changed metabolites, 4-ethylphenyl sulfate, dihydrouracil, and LPC 20:0 was related to the elevation of alkaline phosphatase (ALP), interleukin-6 (IL6), and nuclear factor kappa-B (NF-κB). The levels of 4-ethylphenyl sulfate, dihydrouracil, and LPC 20:0 in plasma were positively correlated with their levels in the liver, suggesting that these metabolites were associated with MO-induced liver injury. MO treatment could increase NHC and cytidine levels, activate cytidine deaminase (CDA), and increase LPC levels. CDA and LPC could increase the mRNA expression level of toll-like receptor (TLR). The current study indicated that the elevation of hepatic TLR may be an important reason for MO leading to the liver injury.
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In this work, a series of new diarylpyrimidine derivatives as microtubule destabilizers were designed, synthesized, and evaluated for anticancer activities. Based on restriction configuration strategy, we introduced the pyrimidine moiety containing the hydrogen-bond acceptors as cis-olefin bond of CA-4 analogs to improve structural stability. Compounds 11a-t exerted antiproliferative activities against three human cancer cell lines (SGC-7901, HeLa, and MCF-7), due to tubulin polymerization inhibition, showing high selectivity toward cancer cells in comparison with non-tumoral HSF cells, as evidenced by MTT assays. In mechanistic investigations, compound 11s remarkably inhibited tubulin polymerization and disorganized microtubule in SGC-7901 cells by binding to tubulin. Moreover, 11s caused G2/M phase cell cycle arrest in SGC-7901 cells in a concentration-dependent manner. Furthermore, molecular modeling analysis revealed that 11s interacts with tubulin through binding to the colchicine site. In addition, the prediction of physicochemical properties disclosed that 11s conformed well to the Lipinski's rule of five. This work offered a fresh viewpoint for the discovery of new tubulin-targeting anticancer drugs.
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OBJECTIVES: Previous studies elucidated that capecitabine (CAP) works as an anti-tumor agent with putative immunosuppressive effects. However, the intricate mechanisms underpinning these effects remain to be elucidated. In this study, we aimed to unravel the molecular pathways by which CAP exerts its immunosuppressive effects to reduce allograft rejection. METHODS: Hearts were transplanted from male BALB/c donors to male C57BL/6 recipients and treated with CAP for seven days. The rejection of these heart transplants was assessed using a range of techniques, including H&E staining, immunohistochemistry, RNA sequencing, LS-MS/MS, and flow cytometry. In vitro, naïve CD4+ T cells were isolated and cultured under Th1 condition medium with varying treatments, flow cytometry, LS-MS/MS were employed to delineate the role of thymidine synthase (TYMS) during Th1 differentiation. RESULTS: CAP treatment significantly mitigated acute allograft rejection and enhanced graft survival by reducing graft damage, T cell infiltration, and levels of circulating pro-inflammatory cytokines. Additionally, it curtailed CD4+ T cell proliferation and the presence of Th1 cells in the spleen. RNA-seq showed that TYMS, the target of CAP, was robustly increased post-transplantation in splenocytes. In vitro, TYMS and its metabolic product dTMP were differentially expressed in Th0 and Th1, and were required after activation of CD4+ T cell and Th1 differentiation. TYMS-specific inhibitor, raltitrexed, and the metabolite of capecitabine, 5-fluorouracil, could inhibit the proliferation and differentiation of Th1. Finally, the combined use of CAP and the commonly used immunosuppressant rapamycin can induce long-term survival of allograft. CONCLUSION: CAP undergoes metabolism conversion to interfere pyrimidine metabolism, which targets TYMS-mediated differentiation of Th1, thereby playing a significant role in mitigating acute cardiac allograft rejection in murine models.
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In the upcoming decades, the incidence and mortality rates of cancer are expected to rise globally, with colorectal and prostate cancers among the most prevalent types. Despite advancements in molecular targeted therapy, platinum-based chemotherapies remain the cornerstone of treatment, especially for colorectal and prostate cancer, with oxaliplatin and cisplatin being extremely effective due to their DNA-targeting capabilities. In our pursuit of new platinum-based chemotherapeutics that are potentially less toxic and more effective, we have explored the combination of the Pt-binding groups of the diaminocyclohexane ring used in oxaliplatin, with the stable amino-pyrimidine hemicurcumin moiety. This new derivative exhibit improved stability in physiological conditions and increased solubility in aqueous media, demonstrating promising effects on cell proliferation of both colorectal and prostate cells. We report herein the complete synthesis and chemical characterization in solution of the new derivative [(1R,2R)-N1-(3-(4-((E)-2-(2-Amino-6-methylpyrimidin-4-yl)vinyl)-2-methoxyphenoxy) propyl) cyclohexane-1,2-diamine] (MPYD). Our analysis includes an examination of its acid-base equilibria, speciation and stability in physiological conditions. The synthesis and in situ formation of Pt(II) complexes were investigated by nuclear magnetic resonance spectroscopy, while density functional theory calculations were employed to elucidate the chemical structure in solution. Results on the biological activity were obtained through cell viability assays on different colorectal and prostate cell lines (HCT116, HT29, PC3 and LNCaP).
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Orotic acid is widely used in healthcare and cosmetic industries. However, orotic acid-producing microorganisms are auxotrophic, which results in inefficient microbial production. Herein, a plasmid-free, uninduced, non-auxotrophic orotic acid hyperproducer was constructed from Escherichia coli W3110. Initially, the orotic acid degradation pathway was blocked and the carbamoyl phosphate supply was enriched. Subsequently, pyr operon from Bacillus subtilis F126 was heterologously expressed and precursors' supply was optimized. Thereafter, pyrE was dynamically regulated to reconstruct the non-auxotrophic pathway. Employing fed-batch cultivation, orotic acid titer, yield, and productivity of strain Ora21 reached 182.5â¯g/L, 0.58â¯g/g, and 3.80â¯g/L/h, respectively, the highest levels reported so far. Finally, a novel "Chaos to Order Cycles (COC)" fermentation was developed, which effectively increased the yield to 0.63â¯g/g. This research is a remarkable achievement in orotic acid production by microbial fermentation and has vast potential for industrial applications.
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Introduction: Nuclear factor kappa (NF-κB) plays a key role in cancer cell proliferation; thus, small molecule inhibitors of NF-κB activity can effectively inhibit breast cancer (BC) progression. We have previously reported oxazine and piperazine-linked pyrimidines as novel anti-cancer agents that can suppress NF-κB activation in BC cells. Moreover, the TRX-01 compound, an oxazine-linked pyrimidine, inhibited MCF-7 cells at a concentration of 9.17 µM in the Alamar Blue assay. Methods: This work involved the analysis of frontier molecular orbitals, HOMO-LUMO interactions, and molecular electrostatic potential for the TRX-01 structure. Additionally, the TRX-01 compound was studied for cytotoxicity, and migration as well as invasion assays were performed on BC cells. Results: Finally, TRX-01 blocked the translocation of NF-κB from the cytoplasm to the nucleus in MCF-7 cells and reduced NF-κB and IκBα levels in a dose-dependent manner. It also suppressed migratory and invasive properties of BC cells. Conclusion: Overall, the data indicates that TRX-01 can function as a novel blocker of BC growth and metastasis by targeting NF-κB activation.
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A series of 4H-Chromene Based Indole - Pyrimidine Hybrids synthesized using simple and efficient multicomponent reaction. The title molecules were evaluated for their invitro antioxidant and antimicrobial activities. Compounds 8g containing bromo substituted naphthalene displayed potent antioxidant activity with IC50 value of 1.09±0.34 µM and 1.10±0.36 µM. Compound 10a, a 4-methylphenyl derivative presented potent activity with antioxidant activity with IC50 value of 1.29±0.35 µM and 1.43±0.38 µM. Subsequently, compounds 8a, 8b, 8d and 10g had shown prominent percentage of inhibition and derived effective IC50 values in comparison to reference drug Ascorbic Acid. The invitro antimicrobial activity carried out against two gram positive and two gram-negative bacteria, and two fungal strains using Ampicillin and Itraconazole as refence drugs. Compound 10f exhibited exceptional efficacy against all types of bacterial and fungal strains compared to Ampicillin and Itraconazole, compounds 8e and 8g showed activity against bacterial strains whereas compound 10g exhibited the most effective zone of inhibition against fungal strains. The molecular docking study against crystal structure of NADPH oxidase obtained supporting docking scores and showed notable binding interactions such as H-bond and hydrophobic.
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Fluorescent drugs and pyrimidine-indole scaffolds have been shown to have advantages in cancer treatment. Fluorescent antitumor drugs BF3-o, m, p-phenylenediamine pyrimidine-indole derivatives (PYB1, PYB2, and PYB3) were synthesized by linking pyrimidine and indole groups with aniline through a simple step and introducing BF3. The drugs exhibit promising antitumor activity and their fluorescent properties make them useful for imaging purposes. The optical properties of the three compounds have been investigated. All of them have fluorescent properties and compound PYB2 has good fluorescent properties. Additionally, the in vitro cytotoxicity of these compounds was evaluated against the human cancer cell line HeLa and the human normal cell line L02. The inhibition rates of HeLa cells treated with PYB1, PYB2, and PYB3 at a concentration of 19.2 µg/mL were 80.91%, 77.72%, and 65.94%, respectively. These results indicate a strong inhibitory effect on cancer cells. Further through the cell imaging experiment, we can see that PYB2 can enter the cell through the cell membrane through the fluorescence scattering diagram, which has good biocompatibility. In addition, the possible interactions between the compounds and Ras protein active sites were analyzed by molecular docking. The three compounds can bind well to Ras protein through hydrogen bonding. This study provides a basis for the development and modification of pyrimidine-indole fluorescent anticancer drugs. Compound PYB2 shows potential as a fluorescent anticancer drug.
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INTRODUCTION: Cyclin-dependent protein kinase 4/6 (CDK4/6) is a class of serine/threonine protein kinases that plays a key role in the regulation of the cell cycle. CDK4/6 is highly expressed in cancers such as breast cancer, melanoma, and non-small cell lung cancer (NSCLC). Currently, a variety of CDK4/6 inhibitors have been developed, aiming to develop effective inhibitors to solve CDK4/6 resistance and toxicity. AREAS COVERED: This article searches patents through Espacenet and reviews the development of widely studied CDK inhibitors and FDA-approved CDK4/6 inhibitors, as well as the latest progress of patented inhibitors with good inhibitory activity against CDK4/6 from 2020 to now. EXPERT OPINION: CDK4/6 is highly expressed in many tumors and has become an important anti-tumor target. Among the patents from 2020 to the present, many inhibitors have good kinase inhibitory effects on CDK4/6 and also show great development potential in anti-tumor. However, there is still an urgent need to develop novel CDK4/6 inhibitors that address challenges such as drug resistance, toxicity, and selectivity.
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Antineoplásicos , Quinase 4 Dependente de Ciclina , Quinase 6 Dependente de Ciclina , Desenvolvimento de Medicamentos , Resistencia a Medicamentos Antineoplásicos , Neoplasias , Patentes como Assunto , Inibidores de Proteínas Quinases , Humanos , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Antineoplásicos/farmacologia , AnimaisRESUMO
Small molecules that exhibit broad-spectrum enteroviral inhibitory activity by targeting viral replication proteins are highly desired in antiviral drug discovery studies. To discover new human rhinovirus (hRV) inhibitors, we performed a high-throughput screening of 100,000 compounds from the Korea Chemical Bank library. This search led to identification of two phosphatidylinositol-4-kinase IIIß (PI4KIIIß) inhibitors having the pyrazolo-pyrimidine core structure, which display moderate anti-rhinoviral activity along with mild cytotoxicity. The results of a study aimed at optimizing the activity of the hit compounds showed that the pyrazolo-pyrimidine derivative 6f exhibits the highest activity (EC50 = 0.044, 0.066, and 0.083 µM for hRV-B14, hRV-A16, and hRV-A21, respectively) and moderate toxicity (CC50 = 31.38 µM). Furthermore, 6f has broad-spectrum activities against various hRVs, coxsackieviruses and other enteroviruses, such as EV-A71, EV-D68. An assessment of kinase inhibition potencies demonstrated that 6f possesses a high and selective kinase inhibition activity against PI4KIIIß (IC50 value of 0.057 µM) and not against PI4KIIIα (>10 µM). Moreover, 6f exhibits modest hepatic stability (46.9 and 55.3 % remaining after 30 min in mouse and human liver microsomes, respectively). Finally, an in vivo study demonstrated that 6f possesses a desirable pharmacokinetic profile reflected in low systemic clearance (0.48 Lâh-1 kg-1) and modest oral bioavailability (52.4 %). Hence, 6f (KR-26549) appears to be an ideal lead for the development of new antiviral drugs.
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Antivirais , Pirimidinas , Rhinovirus , Replicação Viral , Humanos , Rhinovirus/efeitos dos fármacos , Antivirais/farmacologia , Antivirais/química , Antivirais/síntese química , Replicação Viral/efeitos dos fármacos , Pirimidinas/farmacologia , Pirimidinas/química , Pirimidinas/síntese química , Animais , Relação Estrutura-Atividade , Estrutura Molecular , Pirazóis/farmacologia , Pirazóis/química , Pirazóis/síntese química , Camundongos , Relação Dose-Resposta a Droga , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/síntese química , Testes de Sensibilidade Microbiana , Fosfotransferases (Aceptor do Grupo Álcool)RESUMO
Dihydroorotase (DHOase) is the third enzyme in the six enzymatic reaction steps of the endogenous pyrimidine nucleotide de novo biosynthesis pathway, which is a metabolic pathway conserved in both bacteria and eukaryotes. However, research on the biological function of DHOase in plant pathogenic fungi is very limited. In this study, we identified and named MoPyr4, a homologous protein of Saccharomyces cerevisiae DHOase Ura4, in the rice blast fungus Magnaporthe oryzae and investigated its ability to regulate fungal growth, pathogenicity, and autophagy. Deletion of MoPYR4 led to defects in growth, conidiation, appressorium formation, the transfer and degradation of glycogen and lipid droplets, appressorium turgor accumulation, and invasive hypha expansion in M. oryzae, which eventually resulted in weakened fungal pathogenicity. Long-term replenishment of exogenous uridine-5'-phosphate (UMP) can effectively restore the phenotype and virulence of the ΔMopyr4 mutant. Further study revealed that MoPyr4 also participated in the regulation of the Pmk1-MAPK signaling pathway, co-localized with peroxisomes for the oxidative stress response, and was involved in the regulation of the Osm1-MAPK signaling pathway in response to hyperosmotic stress. In addition, MoPyr4 interacted with MoAtg5, the core protein involved in autophagy, and positively regulated autophagic degradation. Taken together, our results suggested that MoPyr4 for UMP biosynthesis was crucial for the development and pathogenicity of M. oryzae. We also revealed that MoPyr4 played an essential role in the external stress response and pathogenic mechanism through participation in the Pmk1-MAPK signaling pathway, peroxisome-related oxidative stress response mechanism, the Osm1-MAPK signaling pathway and the autophagy pathway.
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Autofagia , Proteínas Fúngicas , Oryza , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/genética , Oryza/microbiologia , Virulência/genética , Peroxissomos/metabolismo , Doenças das Plantas/microbiologia , Ascomicetos/patogenicidade , Ascomicetos/genética , Ascomicetos/enzimologia , Sistema de Sinalização das MAP Quinases , Estresse OxidativoRESUMO
Irradiation of the major conformation of duplex DNA found in cells (B form) produces cyclobutane pyrimidine dimers (CPDs) from adjacent pyrimidines in a head-to-head orientation (syn) with the C5 substituents in a cis stereochemistry. These CPDs have crucial implications in skin cancer. Irradiation of G-quadruplexes and other non-B DNA conformations in vitro produces, however, CPDs between nonadjacent pyrimidines in nearby loops with syn and head-to-tail orientations (anti) with both cis and trans stereochemistry to yield a mixture of six possible isomers of the T=T dimer. This outcome is further complicated by formation of mixtures of nonadjacent CPDs of C=T, T=C, and C=C, and successful analysis depends on development of specific and sensitive methods. Toward meeting this need, we investigated whether ion mobility mass spectrometry (IMMS) and MS/MS can distinguish the cis,syn and trans,anti T=T CPDs. Ion mobility can afford baseline separation and give relative mobilities that are in accord with predicted cross sections. Complementing this ability to distinguish isomers is MS/MS collisional activation where fragmentation also distinguishes the two isomers and confirms conclusions drawn from ion mobility analysis. The observations offer early support that ion mobility and MS/MS can enable the distinction of DNA photoproduct isomers.