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Archives of Microbiology Jun 2024Vibrio parahaemolyticus possesses two distinct type VI secretion systems (T6SS), namely T6SS1 and T6SS2. T6SS1 is predominantly responsible for adhesion to Caco-2 and...
Vibrio parahaemolyticus possesses two distinct type VI secretion systems (T6SS), namely T6SS1 and T6SS2. T6SS1 is predominantly responsible for adhesion to Caco-2 and HeLa cells and for the antibacterial activity of V. parahaemolyticus, while T6SS2 mainly contributes to HeLa cell adhesion. However, it remains unclear whether the T6SS systems have other physiological roles in V. parahaemolyticus. In this study, we demonstrated that the deletion of icmF2, a structural gene of T6SS2, reduced the biofilm formation capacity of V. parahaemolyticus under low salt conditions, which was also influenced by the incubation time. Nonetheless, the deletion of icmF2 did not affect the biofilm formation capacity in marine-like growth conditions, nor did it impact the flagella-driven swimming and swarming motility of V. parahaemolyticus. IcmF2 was found to promote the production of the main components of the biofilm matrix, including extracellular DNA (eDNA) and extracellular proteins, and cyclic di-GMP (c-di-GMP) in V. parahaemolyticus. Additionally, IcmF2 positively influenced the transcription of cpsA, mfpA, and several genes involved in c-di-GMP metabolism, including scrJ, scrL, vopY, tpdA, gefA, and scrG. Conversely, the transcription of scrA was negatively impacted by IcmF2. Therefore, IcmF2-dependent biofilm formation was mediated through its effects on the production of eDNA, extracellular proteins, and c-di-GMP, as well as its impact on the transcription of cpsA, mfpA, and genes associated with c-di-GMP metabolism. This study confirmed new physiological roles for IcmF2 in promoting biofilm formation and c-di-GMP production in V. parahaemolyticus.
Topics: Vibrio parahaemolyticus; Biofilms; Type VI Secretion Systems; Bacterial Proteins; Cyclic GMP; Humans; Gene Expression Regulation, Bacterial; HeLa Cells
PubMed: 38907796
DOI: 10.1007/s00203-024-04060-x -
Nature Communications Jun 2024Adenosine-5'-triphosphate (ATP), the primary energy currency in cellular processes, drives metabolic activities and biosynthesis. Despite its importance, understanding...
Adenosine-5'-triphosphate (ATP), the primary energy currency in cellular processes, drives metabolic activities and biosynthesis. Despite its importance, understanding intracellular ATP dynamics' impact on bioproduction and exploiting it for enhanced bioproduction remains largely unexplored. Here, we harness an ATP biosensor to dissect ATP dynamics across different growth phases and carbon sources in multiple microbial strains. We find transient ATP accumulations during the transition from exponential to stationary growth phases in various conditions, coinciding with fatty acid (FA) and polyhydroxyalkanoate (PHA) production in Escherichia coli and Pseudomonas putida, respectively. We identify carbon sources (acetate for E. coli, oleate for P. putida) that elevate steady-state ATP levels and boost FA and PHA production. Moreover, we employ ATP dynamics as a diagnostic tool to assess metabolic burden, revealing bottlenecks that limit limonene bioproduction. Our results not only elucidate the relationship between ATP dynamics and bioproduction but also showcase its value in enhancing bioproduction in various microbial species.
Topics: Adenosine Triphosphate; Biosensing Techniques; Escherichia coli; Pseudomonas putida; Fatty Acids; Polyhydroxyalkanoates; Energy Metabolism; Carbon; Oleic Acid
PubMed: 38906854
DOI: 10.1038/s41467-024-49579-1 -
Bioorganic & Medicinal Chemistry Jul 2024N-(Benzothiazole-2-yl)pyrrolamide DNA gyrase inhibitors with benzyl or phenethyl substituents attached to position 3 of the benzothiazole ring or to the carboxamide...
Exploring the interaction of N-(benzothiazol-2-yl)pyrrolamide DNA gyrase inhibitors with the GyrB ATP-binding site lipophilic floor: A medicinal chemistry and QTAIM study.
N-(Benzothiazole-2-yl)pyrrolamide DNA gyrase inhibitors with benzyl or phenethyl substituents attached to position 3 of the benzothiazole ring or to the carboxamide nitrogen atom were prepared and studied for their inhibition of Escherichia coli DNA gyrase by supercoiling assay. Compared to inhibitors bearing the substituents at position 4 of the benzothiazole ring, the inhibition was attenuated by moving the substituent to position 3 and further to the carboxamide nitrogen atom. A co-crystal structure of (Z)-3-benzyl-2-((4,5-dibromo-1H-pyrrole-2-carbonyl)imino)-2,3-dihydrobenzo[d]-thiazole-6-carboxylic acid (I) in complex with E. coli GyrB24 (ATPase subdomain) was solved, revealing the binding mode of this type of inhibitor to the ATP-binding pocket of the E. coli GyrB subunit. The key binding interactions were identified and their contribution to binding was rationalised by quantum theory of atoms in molecules (QTAIM) analysis. Our study shows that the benzyl or phenethyl substituents bound to the benzothiazole core interact with the lipophilic floor of the active site, which consists mainly of residues Gly101, Gly102, Lys103 and Ser108. Compounds with substituents at position 3 of the benzothiazole core were up to two orders of magnitude more effective than compounds with substituents at the carboxamide nitrogen. In addition, the 6-oxalylamino compounds were more potent inhibitors of E. coli DNA gyrase than the corresponding 6-acetamido analogues.
Topics: Topoisomerase II Inhibitors; DNA Gyrase; Binding Sites; Escherichia coli; Structure-Activity Relationship; Benzothiazoles; Adenosine Triphosphate; Molecular Structure; Quantum Theory; Anti-Bacterial Agents; Models, Molecular
PubMed: 38906068
DOI: 10.1016/j.bmc.2024.117798 -
Plant Signaling & Behavior Dec 2024Extracellular ATP (eATP) orchestrates vital processes in plants, akin to its role in animals. P2K1 is a crucial receptor mediating eATP effects. Immunoprecipitation...
Extracellular ATP (eATP) orchestrates vital processes in plants, akin to its role in animals. P2K1 is a crucial receptor mediating eATP effects. Immunoprecipitation tandem mass spectrometry data highlighted FERONIA's significant interaction with P2K1, driving us to explore its role in eATP signaling. Here, we investigated putative P2K1-interactor, FERONIA, which is a versatile receptor kinase pivotal in growth and stress responses. We employed a FERONIA loss-of-function mutant, , to dissect its effects on eATP signaling. Interestingly, showed distinct calcium responses compared to wild type, while eATP-responsive genes were constitutively upregulated in . Additionally, displayed insensitivity to eATP-regulated root growth and reduced cell wall accumulation. Together, these results uncover a role for FERONIA in regulating eATP signaling. Overall, our study deepens our understanding of eATP signaling, revealing the intricate interplay between P2K1 and FERONIA impacting the interface between growth and defense.
Topics: Plant Roots; Signal Transduction; Arabidopsis Proteins; Arabidopsis; Adenosine Triphosphate; Gene Expression Regulation, Plant; Phosphotransferases; Protein Serine-Threonine Kinases
PubMed: 38905329
DOI: 10.1080/15592324.2024.2370706 -
Proceedings of the National Academy of... Jun 2024Hydrogen isotope ratios (δH) represent an important natural tracer of metabolic processes, but quantitative models of processes controlling H-fractionation in aquatic...
Hydrogen isotope ratios (δH) represent an important natural tracer of metabolic processes, but quantitative models of processes controlling H-fractionation in aquatic photosynthetic organisms are lacking. Here, we elucidate the underlying physiological controls of H/H fractionation in algal lipids by systematically manipulating temperature, light, and CO(aq) in continuous cultures of the haptophyte . We analyze the hydrogen isotope fractionation in alkenones (α), a class of acyl lipids specific to this species and other haptophyte algae. We find a strong decrease in the α with increasing CO(aq) and confirm α correlates with temperature and light. Based on the known biosynthesis pathways, we develop a cellular model of the δH of algal acyl lipids to evaluate processes contributing to these controls on fractionation. Simulations show that longer residence times of NADPH in the chloroplast favor a greater exchange of NADPH with H-richer intracellular water, increasing α. Higher chloroplast CO(aq) and temperature shorten NADPH residence time by enhancing the carbon fixation and lipid synthesis rates. The inverse correlation of α to CO(aq) in our cultures suggests that carbon concentrating mechanisms (CCM) do not achieve a constant saturation of CO at the Rubisco site, but rather that chloroplast CO varies with external CO(aq). The pervasive inverse correlation of α with CO(aq) in the modern and preindustrial ocean also suggests that natural populations may not attain a constant saturation of Rubisco with the CCM. Rather than reconstructing growth water, α may be a powerful tool to elucidate the carbon limitation of photosynthesis.
Topics: Carbon Dioxide; Haptophyta; Lipids; Photosynthesis; Hydrogen; Chloroplasts; Deuterium; NADP; Temperature; Chemical Fractionation; Lipid Metabolism
PubMed: 38905238
DOI: 10.1073/pnas.2318570121 -
Nucleosides, Nucleotides & Nucleic Acids Jun 2024The chemical synthesis of guanosine nucleosides generates both the and regioisomers, which require careful separation to obtain the desired isomer. To preferentially...
The chemical synthesis of guanosine nucleosides generates both the and regioisomers, which require careful separation to obtain the desired isomer. To preferentially obtain the isomer, a bulky diphenylcarbamoyl (DPC) group can be installed at the position of guanine. However, installation of the DPC group presents a challenging task due to low solubility of the -acetyl protected guanine. Here we report the usage of commercially available 2-amino-6-chloro purine as a new strategy that offers a more efficient route to the synthesis of the guanine phosphoramidite of threose nucleic acid (TNA).
PubMed: 38904107
DOI: 10.1080/15257770.2024.2369688 -
NPJ Biofilms and Microbiomes Jun 2024Bacteria induced metamorphosis observed in nearly all marine invertebrates. However, the mechanism of bacteria regulating the larvae-juvenile metamorphosis remains...
Bacteria induced metamorphosis observed in nearly all marine invertebrates. However, the mechanism of bacteria regulating the larvae-juvenile metamorphosis remains unknown. Here, we test the hypothesis that c-di-GMP, a ubiquitous bacterial second-messenger molecule, directly triggers the mollusc Mytilus coruscus larval metamorphosis via the stimulator of interferon genes (STING) receptor. We determined that the deletion of c-di-GMP synthesis genes resulted in reduced c-di-GMP levels and biofilm-inducing activity on larval metamorphosis, accompanied by alterations in extracellular polymeric substances. Additionally, c-di-GMP extracted from tested varying marine bacteria all exhibited inducing activity on larval metamorphosis. Simultaneously, through pharmacological and molecular experiments, we demonstrated that M. coruscus STING (McSTING) participates in larval metamorphosis by binding with c-di-GMP. Our findings reveal that new role of bacterial c-di-GMP that triggers mussel larval metamorphosis transition, and extend knowledge in the interaction of bacteria and host development in marine ecosystems.
Topics: Animals; Metamorphosis, Biological; Larva; Cyclic GMP; Biofilms; Mytilus; Bacteria; Membrane Proteins
PubMed: 38902226
DOI: 10.1038/s41522-024-00523-7 -
Nature Communications Jun 2024C2'-halogenation has been recognized as an essential modification to enhance the drug-like properties of nucleotide analogs. The direct C2'-halogenation of the...
C2'-halogenation has been recognized as an essential modification to enhance the drug-like properties of nucleotide analogs. The direct C2'-halogenation of the nucleotide 2'-deoxyadenosine-5'-monophosphate (dAMP) has recently been achieved using the Fe(II)/α-ketoglutarate-dependent nucleotide halogenase AdaV. However, the limited substrate scope of this enzyme hampers its broader applications. In this study, we report two halogenases capable of halogenating 2'-deoxyguanosine monophosphate (dGMP), thereby expanding the family of nucleotide halogenases. Computational studies reveal that nucleotide specificity is regulated by the binding pose of the phosphate group. Based on these findings, we successfully engineered the substrate specificity of these halogenases by mutating second-sphere residues. This work expands the toolbox of nucleotide halogenases and provides insights into the regulation mechanism of nucleotide specificity.
Topics: Substrate Specificity; Protein Engineering; Halogenation; Nucleotides; Deoxyguanine Nucleotides; Escherichia coli
PubMed: 38898020
DOI: 10.1038/s41467-024-49147-7 -
Experimental Cell Research Jul 2024Glaucoma is characterized by pathological elevation of intraocular pressure (IOP) due to dysfunctional trabecular meshwork (TM), which is the primary cause of...
Glaucoma is characterized by pathological elevation of intraocular pressure (IOP) due to dysfunctional trabecular meshwork (TM), which is the primary cause of irreversible vision loss. There are currently no effective treatment strategies for glaucoma. Mitochondrial function plays a crucial role in regulating IOP within the TM. In this study, primary TM cells treated with dexamethasone were used to simulate glaucomatous changes, showing abnormal cellular cytoskeleton, increased expression of extracellular matrix, and disrupted mitochondrial fusion and fission dynamics. Furthermore, glaucomatous TM cell line GTM3 exhibited impaired mitochondrial membrane potential and phagocytic function, accompanied by decreased oxidative respiratory levels as compared to normal TM cells iHTM. Mechanistically, lower NAD + levels in GTM3, possibly associated with increased expression of key enzymes CD38 and PARP1 related to NAD + consumption, were observed. Supplementation of NAD + restored mitochondrial function and cellular viability in GTM3 cells. Therefore, we propose that the aberrant mitochondrial function in glaucomatous TM cells may be attributed to increased NAD + consumption dependent on CD38 and PARP1, and NAD + supplementation could effectively ameliorate mitochondrial function and improve TM function, providing a novel alternative approach for glaucoma treatment.
Topics: Trabecular Meshwork; Mitochondria; Glaucoma; NAD; Humans; Membrane Potential, Mitochondrial; Intraocular Pressure; Cell Survival; ADP-ribosyl Cyclase 1; Cell Line; Poly (ADP-Ribose) Polymerase-1; Dexamethasone; Cells, Cultured
PubMed: 38897410
DOI: 10.1016/j.yexcr.2024.114137 -
Science Advances Jun 2024Chloroplasts are the powerhouse of the plant cell, and their activity must be matched to plant growth to avoid photooxidative damage. We have identified a...
Chloroplasts are the powerhouse of the plant cell, and their activity must be matched to plant growth to avoid photooxidative damage. We have identified a posttranslational mechanism linking the eukaryotic target of rapamycin (TOR) kinase that promotes growth and the guanosine tetraphosphate (ppGpp) signaling pathway of prokaryotic origins that regulates chloroplast activity and photosynthesis in particular. We find that RelA SpoT homolog 3 (RSH3), a nuclear-encoded enzyme responsible for ppGpp biosynthesis, interacts directly with the TOR complex via a plant-specific amino-terminal region which is phosphorylated in a TOR-dependent manner. Down-regulating TOR activity causes a rapid increase in ppGpp synthesis in RSH3 overexpressors and reduces photosynthetic capacity in an RSH-dependent manner in wild-type plants. The TOR-RSH3 signaling axis therefore regulates the equilibrium between chloroplast activity and plant growth, setting a precedent for the regulation of organellar function by TOR.
Topics: Photosynthesis; Chloroplasts; Arabidopsis Proteins; Signal Transduction; Arabidopsis; Phosphorylation; Protein Processing, Post-Translational; Gene Expression Regulation, Plant; Guanosine Tetraphosphate; TOR Serine-Threonine Kinases; Phosphatidylinositol 3-Kinases
PubMed: 38896607
DOI: 10.1126/sciadv.adj3268