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International Journal of Molecular... Feb 2024Dengue virus (DENV) causes dengue fever and dengue hemorrhagic fever, and DENV infection kills 20,000 people annually worldwide. Therefore, the development of anti-DENV...
Dengue virus (DENV) causes dengue fever and dengue hemorrhagic fever, and DENV infection kills 20,000 people annually worldwide. Therefore, the development of anti-DENV drugs is urgently needed. Sofosbuvir (SOF) is an effective drug for HCV-related diseases, and its triphosphorylated metabolite inhibits viral RNA synthesis by the RNA-dependent RNA polymerase (RdRp) of HCV. (2'R)-2'-Deoxy-2'-fluoro-2'-methyluridine (FMeU) is the dephosphorylated metabolite produced from SOF. The effects of SOF and FMeU on DENV1 replication were analyzed using two DENV1 replicon-based methods that we previously established. First, a replicon-harboring cell assay showed that DENV1 replicon replication in human hepatic Huh7 cells was decreased by SOF but not by FMeU. Second, a transient replicon assay showed that DENV1 replicon replication in Huh7 cells was decreased by SOF; however, in hamster kidney BHK-21 cells, it was not suppressed by SOF. Additionally, the replicon replication in Huh7 and BHK-21 cells was not affected by FMeU. Moreover, we assessed the effects of SOF on infectious DENV1 production. SOF suppressed infectious DENV1 production in Huh7 cells but not in monkey kidney Vero cells. To examine the substrate recognition of the HCV and DENV1 RdRps, the complex conformation of SOF-containing DENV1 RdRp or HCV RdRp was predicted using AlphaFold 2. These results indicate that SOF may be used as a treatment for DENV1 infection.
Topics: Animals; Cricetinae; Chlorocebus aethiops; Humans; Sofosbuvir; Antiviral Agents; Vero Cells; RNA-Dependent RNA Polymerase; Virus Replication; Hepatitis C; Hepacivirus
PubMed: 38396699
DOI: 10.3390/ijms25042022 -
Nature Communications Feb 2024Selenium is an essential multifunctional trace element in diverse organisms. The only Se-glycosyltransferase identified that catalyzes the incorporation of selenium in...
Selenium is an essential multifunctional trace element in diverse organisms. The only Se-glycosyltransferase identified that catalyzes the incorporation of selenium in selenoneine biosynthesis is SenB from Variovorax paradoxus. Although the biochemical function of SenB has been investigated, its substrate specificity, structure, and catalytic mechanism have not been elucidated. Here, we reveal that SenB exhibits sugar donor promiscuity and can utilize six UDP-sugars to generate selenosugars. We report crystal structures of SenB complexed with different UDP-sugars. The key elements N20/T23/E231 contribute to the sugar donor selectivity of SenB. A proposed catalytic mechanism is tested by structure-guided mutagenesis, revealing that SenB yields selenosugars by forming C-Se glycosidic bonds via spontaneous deprotonation and disrupting Se-P bonds by nucleophilic water attack, which is initiated by the critical residue K158. Furthermore, we functionally and structurally characterize two other Se-glycosyltransferases, CbSenB from Comamonadaceae bacterium and RsSenB from Ramlibacter sp., which also exhibit sugar donor promiscuity.
Topics: Glycosyltransferases; Selenium; Uridine Diphosphate Sugars; Carbohydrates; Sugars; Substrate Specificity; Histidine; Organoselenium Compounds
PubMed: 38395953
DOI: 10.1038/s41467-024-46065-6 -
Chemical Research in Toxicology Mar 2024The biomarker 5-chlorocytosine (5ClC) appears in the DNA of inflamed tissues. Replication of a site-specific 5ClC in a viral DNA genome results in C → T mutations,...
The biomarker 5-chlorocytosine (5ClC) appears in the DNA of inflamed tissues. Replication of a site-specific 5ClC in a viral DNA genome results in C → T mutations, which is consistent with 5ClC acting as a thymine mimic in vivo. Direct damage of nucleic acids by immune-cell-derived hypochlorous acid is one mechanism by which 5ClC could appear in the genome. A second, nonmutually exclusive mechanism involves damage of cytosine nucleosides or nucleotides in the DNA precursor pool, with subsequent utilization of the 5ClC deoxynucleotide triphosphate as a precursor for DNA synthesis. The present work characterized the mutagenic properties of 5ClC in the nucleotide pool by exposing cells to the nucleoside 5-chloro-2'-deoxycytidine (5CldC). In both and mouse embryonic fibroblasts (MEFs), 5CldC in the growth media was potently mutagenic, indicating that 5CldC enters cells and likely is erroneously incorporated into the genome from the nucleotide pool. High-resolution sequencing of DNA from MEFs derived from the Δ C57BL/6J mouse allowed qualitative and quantitative characterization of 5CldC-induced mutations; CG → TA transitions in 5'-GC(Y)-3' contexts (Y = a pyrimidine) were dominant, while TA → CG transitions appeared at a much lower frequency. The high-resolution mutational spectrum of 5CldC revealed a notable similarity to the Catalogue of Somatic Mutations in Cancer mutational signatures SBS84 and SBS42, which appear in human lymphoid tumors and in occupationally induced cholangiocarcinomas, respectively. SBS84 is associated with the expression of activation-induced cytidine deaminase (AID), a cytosine deaminase associated with inflammation, as well as immunoglobulin gene diversification during antibody maturation. The similarity between the spectra of AID activation and 5CldC could be coincidental; however, the administration of 5CldC did induce some AID expression in MEFs, which have no inherent expression of its gene. In summary, this work shows that 5CldC induces a distinct pattern of mutations in cells. Moreover, that pattern resembles human mutational signatures induced by inflammatory processes, such as those triggered in certain malignancies.
Topics: Animals; Humans; Mice; Mice, Inbred C57BL; Fibroblasts; Mutation; Deoxycytidine; Neoplasms; DNA; Mutagens; Nucleotides
PubMed: 38394377
DOI: 10.1021/acs.chemrestox.3c00358 -
Regional Anesthesia and Pain Medicine Apr 2024Postdural puncture headache has been traditionally viewed as benign, self-limited, and highly responsive to epidural blood patching (EBP) when needed. A growing body of...
BACKGROUND
Postdural puncture headache has been traditionally viewed as benign, self-limited, and highly responsive to epidural blood patching (EBP) when needed. A growing body of data from patients experiencing unintended dural puncture (UDP) in the setting of attempted labor epidural placement suggests a minority of patients will have more severe and persistent symptoms. However, the mechanisms accounting for the failure of EBP following dural puncture remain obscure. An understanding of these potential mechanisms is critical to guide management decisions in the face of severe and persistent cerebrospinal fluid (CSF) leak.
CASE PRESENTATION
We report the case of a peripartum patient who developed a severe and persistent CSF leak unresponsive to multiple EBPs following a UDP during epidural catheter placement for labor analgesia. Lumbar MRI revealed a ventral rather than dorsal epidural fluid collection suggesting that the needle had crossed the thecal sac and punctured the ventral dura, creating a puncture site not readily accessible to blood injected in the dorsal epidural space. The location of this persistent ventral dural defect was confirmed with digital subtraction myelography, permitting a transdural surgical exploration and repair of the ventral dura with resolution of the severe intracranial hypotension.
CONCLUSIONS
A ventral rather than dorsal dural puncture is one mechanism that may contribute to both severe and persistent spinal CSF leak with resulting intracranial hypotension following a UDP.
Topics: Humans; Intracranial Hypotension; Blood Patch, Epidural; Cerebrospinal Fluid Leak; Punctures; Post-Dural Puncture Headache; Iatrogenic Disease; Uridine Diphosphate
PubMed: 38388018
DOI: 10.1136/rapm-2023-105197 -
Biochemistry Mar 2024is a Gram-negative pathogenic bacterium commonly found in chickens and is the leading cause of human diarrheal disease worldwide. The various serotypes of produce...
is a Gram-negative pathogenic bacterium commonly found in chickens and is the leading cause of human diarrheal disease worldwide. The various serotypes of produce structurally distinct capsular polysaccharides (CPSs) on the exterior surfaces of the cell wall. The capsular polysaccharide from serotype HS:5 is composed of a repeating sequence of d--d--heptose and d-glucitol-6-phosphate. We previously defined the pathway for the production of d--d--heptose in . Here, we elucidate the biosynthetic pathway for the assembly of cytidine diphosphate (CDP)-6-d-glucitol by the combined action of two previously uncharacterized enzymes. The first enzyme catalyzes the formation of CDP-6-d-fructose from cytidine triphosphate (CTP) and d-fructose-6-phosphate. The second enzyme reduces CDP-6-d-fructose with NADPH to generate CDP-6-d-glucitol. Using sequence similarity network (SSN) and genome neighborhood network (GNN) analyses, we predict that these pairs of proteins are responsible for the biosynthesis of CDP-6-d-glucitol and/or CDP-d-mannitol in the lipopolysaccharides (LPSs) and capsular polysaccharides in more than 200 other organisms. In addition, high resolution X-ray structures of the second enzyme are reported, which provide novel insight into the manner in which an open-chain nucleotide-linked sugar is harbored in an active site cleft.
Topics: Animals; Humans; Campylobacter jejuni; Sorbitol; Chickens; Polysaccharides; Cytidine Diphosphate; Fructose; Polysaccharides, Bacterial
PubMed: 38386885
DOI: 10.1021/acs.biochem.3c00706 -
Biochemistry Mar 2024is a human pathogen and a leading cause of food poisoning in North America and Europe. The exterior surface of the bacterial cell wall is attached to a polymeric coat...
is a human pathogen and a leading cause of food poisoning in North America and Europe. The exterior surface of the bacterial cell wall is attached to a polymeric coat of sugar molecules known as the capsular polysaccharide (CPS) that helps protect the organism from the host immune response. The CPS is composed of a repeating sequence of common and unusual sugar residues. In the HS:11 serotype of , we identified two enzymes in the gene cluster for CPS formation that are utilized for the biosynthesis of UDP-α--acetyl-d-mannosaminuronic acid (UDP-ManNAcA). In the first step, UDP-α--acetyl-d-glucosamine (UDP-GlcNAc) is epimerized at C2 to form UDP-α--acetyl-d-mannosamine (UDP-ManNAc). This product is then oxidized by a NAD-dependent C6-dehydrogenase to form UDP-ManNAcA. In the HS:6 serotype ( strain 81116), we identified three enzymes that are required for the biosynthesis of CMP-β--acetyl-d-neuraminic acid (CMP-Neu5Ac). In the first step, UDP-GlcNAc is epimerized at C2 and subsequently hydrolyzed to form -acetyl-d-mannosamine (ManNAc) with the release of UDP. This product is then condensed with PEP by -acetyl-d-neuraminate synthase to form -acetyl-d-neuraminic acid (Neu5Ac). In the final step, CMP--acetyl-d-neuraminic acid synthase utilizes CTP to convert this product into CMP-Neu5Ac. A bioinformatic analysis of these five enzymes from serotypes HS:11 and HS:6 identified other bacterial species that can produce UDP-ManNAcA or CMP-Neu5Ac for CPS formation.
Topics: Humans; Campylobacter jejuni; Polysaccharides; Neuraminic Acids; Sugars; Uridine Diphosphate; Cytidine Monophosphate; Sialic Acids; Uronic Acids
PubMed: 38382015
DOI: 10.1021/acs.biochem.3c00664 -
Cell Reports Feb 2024Autophagy is an essential degradation and recycling process that maintains cellular homeostasis during stress or nutrient deprivation. However, certain types of tumors...
Autophagy is an essential degradation and recycling process that maintains cellular homeostasis during stress or nutrient deprivation. However, certain types of tumors such as pancreatic cancers can circumvent autophagy inhibition to sustain growth. The mechanism that autophagy-deficient pancreatic ductal adenocarcinoma (PDAC) uses to grow under nutrient deprivation is poorly understood. Our data show that nutrient deprivation in PDAC results in UDP-glucose dehydrogenase (UGDH) degradation, which is dependent on autophagic cargo receptor sequestosome 1 (p62). Moreover, we demonstrate that accumulated UGDH is indispensable for autophagy-deficient PDAC cells proliferation by promoting hyaluronic acid (HA) synthesis upon energy deprivation. Using an orthotopic mouse model of PDAC, we find that inhibition of HA synthesis by targeting UGDH in PDAC reduces tumor weight. Thus, the combined inhibition of HA and autophagy might be an attractive strategy for PDAC treatment.
Topics: Animals; Mice; Hyaluronic Acid; Pancreatic Neoplasms; Carcinoma, Pancreatic Ductal; Autophagy; Glucose Dehydrogenases; Uridine Diphosphate
PubMed: 38367236
DOI: 10.1016/j.celrep.2024.113808 -
Nature Communications Feb 2024Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of Kaposi's sarcoma (KS) and multiple types of B cell malignancies. Emerging evidence demonstrates...
Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of Kaposi's sarcoma (KS) and multiple types of B cell malignancies. Emerging evidence demonstrates that KSHV reprograms host-cell central carbon metabolic pathways, which contributes to viral persistence and tumorigenesis. However, the mechanisms underlying KSHV-mediated metabolic reprogramming remain poorly understood. Carbamoyl-phosphate synthetase 2, aspartate transcarbamoylase, and dihydroorotase (CAD) is a key enzyme of the de novo pyrimidine synthesis, and was recently identified to deamidate the NF-κB subunit RelA to promote aerobic glycolysis and cell proliferation. Here we report that KSHV infection exploits CAD for nucleotide synthesis and glycolysis. Mechanistically, KSHV vCyclin binds to and hijacks cyclin-dependent kinase CDK6 to phosphorylate Ser-1900 on CAD, thereby activating CAD-mediated pyrimidine synthesis and RelA-deamidation-mediated glycolytic reprogramming. Correspondingly, genetic depletion or pharmacological inhibition of CDK6 and CAD potently impeded KSHV lytic replication and thwarted tumorigenesis of primary effusion lymphoma (PEL) cells in vitro and in vivo. Altogether, our work defines a viral metabolic reprogramming mechanism underpinning KSHV oncogenesis, which may spur the development of new strategies to treat KSHV-associated malignancies and other diseases.
Topics: Humans; Sarcoma, Kaposi; Herpesvirus 8, Human; Glycolysis; Carcinogenesis; Pyrimidines; Nucleotides
PubMed: 38365882
DOI: 10.1038/s41467-024-45852-5 -
BMC Gastroenterology Feb 2024Sofosbuvir/Velpatasvir (Epclusa, ECS) is the first pan-genotype direct-acting antiviral agent (DAA) for hepatitis C virus (HCV) infection, and Danoprevir (DNV) is the... (Comparative Study)
Comparative Study
BACKGROUND
Sofosbuvir/Velpatasvir (Epclusa, ECS) is the first pan-genotype direct-acting antiviral agent (DAA) for hepatitis C virus (HCV) infection, and Danoprevir (DNV) is the first DAA developed by a Chinese local enterprise, which is suitable for combined use with other drugs to treat genotype 1b chronic hepatitis C. However, previous reports have never compared the real-world data of ECS and DNV.
PATIENTS AND METHODS
178 chronic hepatitis C patients were retrospectively recruited, and 94cases were accepted with Sofosbuvir/Velpatasvir ± Ribavirin (ECS group), and others (n = 84 treated with DNV combination therapy (DNV group). The HCV genotype, virological response, adverse effects and some laboratory biochemical indexes were contrasted between above two groups in the real world study.
RESULTS
DNV group had significantly lower level of alpha-fetoprotein (AFP), lower rates of decompensated cirrhosis ( P < 0.05). ECS group possessed more 6a (31.91% vs.13.10%) while DNV group was provided with more 1b (48.81% vs. 22.34%) patients. Significantly poor liver function was detected in ECS group at 4-week treatment (ALT and AST) and 12-week follow-up (AST) (all P < 0.05). The SVR12 undetectable rates of both groups were 100%, and no serious event was observed during the treatment and follow-up in both groups.
CONCLUSION
In this retrospective real-world study, the efficacy of DNV combined therapy is similar to Sofosbuvir/Velpatasvir ± Ribavirin for chronic HCV infection, and the safety is comparable. DNV based therapy is a promising regimen for chronic hepatitis C.
Topics: Humans; Antiviral Agents; Benzimidazoles; Benzopyrans; Carbamates; China; Cyclopropanes; Drug Combinations; Genotype; Hepacivirus; Hepatitis C; Hepatitis C, Chronic; Heterocyclic Compounds, 4 or More Rings; Isoindoles; Lactams, Macrocyclic; Liver Cirrhosis; Proline; Retrospective Studies; Ribavirin; Sofosbuvir; Sulfonamides; Treatment Outcome
PubMed: 38355447
DOI: 10.1186/s12876-024-03147-5 -
Plant Physiology Apr 2024Photoreactivation enzyme that repairs cyclobutane pyrimidine dimer (CPD) induced by ultraviolet-B radiation, commonly called CPD photolyase (PHR) is essential for plants...
Photoreactivation enzyme that repairs cyclobutane pyrimidine dimer (CPD) induced by ultraviolet-B radiation, commonly called CPD photolyase (PHR) is essential for plants living under sunlight. Rice (Oryza sativa) PHR (OsPHR) is a unique triple-targeting protein. The signal sequences required for its translocation to the nucleus or mitochondria are located in the C-terminal region but have yet to be identified for chloroplasts. Here, we identified sequences located in the N-terminal region, including the serine-phosphorylation site at position 7 of OsPHR, and found that OsPHR is transported/localized to chloroplasts via a vesicle transport system under the control of serine-phosphorylation. However, the sequence identified in this study is only conserved in some Poaceae species, and in many other plants, PHR is not localized to the chloroplasts. Therefore, we reasoned that Poaceae species need the ability to repair CPD in the chloroplast genome to survive under sunlight and have uniquely acquired this mechanism for PHR chloroplast translocation.
Topics: Chloroplasts; Deoxyribodipyrimidine Photo-Lyase; Ultraviolet Rays; Oryza; Plant Proteins; Pyrimidine Dimers; Poaceae; Amino Acid Sequence; Protein Transport
PubMed: 38345835
DOI: 10.1093/plphys/kiae060