bluuebunny/crossref_metadata_2025_split huggingface dataset:
Functional loss ofrffGandrfbB,encoding dTDP-glucose 4,6-dehydratase, changes colony morphology, cell shape, motility and virulence inSalmonellaTyphimurium
Authors: Chakravorty Subhashish , Banerjee Pip , Joseph Joel P. , Pathak Sanmoy , Verma Taru , Das Mrinmoy , Nandi Dipankar 
Published: June 2023

AbstractLipopolysaccharide (LPS) O-antigen and enterobacterial common antigen (ECA) play crucial roles in maintaining the structural integrity of the outer membrane in Gram-negative bacteria. Previous studies conducted with either LPS or ECA mutants have highlighted the importance of these cell surface polysaccharides in the physiology ofSalmonella entericaserovar Typhimurium (S. Typhimurium). However, the functional consequences resulting from the abrogation of both O-antigen and ECA synthesis inS. Typhimurium are not well studied. In the present study, we generated single and double gene-deleted mutants ofrffGandrfbB, which are paralogs, encoding dTDP-glucose 4,6-dehydratase that catalyze steps in the synthesis of both O-antigen and ECA. The functional loss of bothrffGandrfbB(ΔrffGΔrfbB), but not in single gene-deleted strains, results in a round cell morphology, smaller colony formation and altered LPS profile. In addition, the ΔrffGΔrfbBstrain displays defects in outer membrane permeability, causing hypersensitivity to bile and cell wall targeting antibiotics, e.g., meropenem and polymyxin B. Transcriptomic analysis identified flagellar and SPI-1 pathway to be highly down-regulated in the ΔrffGΔrfbBstrain which leads to impaired swimming and swarming motility and lower adhesion and invasion of HeLa cells. Importantly, the ΔrffGΔrfbBstrain is less proficient in colonizing Peyer’s patches, spleen and liver, is unable to induce pro-inflammatory cytokines and is attenuated in both the oral and intra-peritoneal models ofS. Typhimurium infection in mice. Overall, this study highlights the importance ofrffGandrfbBin maintaining cell wall integrity, colony and cellular morphology, motility and virulence inS. Typhimurium.
A novel role for the peptidyl-prolylcis-transisomerase Cyclophilin A in DNA-repair following replication fork stalling via the MRE11-RAD50-NBS1 complex
Authors: Bedir Marisa , Outwin Emily , Colnaghi Rita , Bassett Lydia , Abramowicz Iga , O’Driscoll Mark 
Published: June 2023

AbstractWe previously reported that non-homologous end-joining (NHEJ)-defective humanLIG4-/-pre-B lymphocytes were unexpectedly sensitive to killing by the cyclic peptide Cyclosporin A (CsA), a common component of bone marrow transplantation conditioning and maintenance regimes. We also found that CsA induced DNA double strand breaks (DSBs) in LIG4 syndrome patient fibroblasts, specifically upon transit through S-phase. The molecular basis underlying these CsA impacts has not been described hitherto. We postulated that CsA-induced genomic instability may reflect a direct role of Cyclophilin A (CYPA) in DNA repair, as CYPA is the primary physiological target interactor of CsA.CYPA is the founding member of the Cyclophilin family of peptidyl-prolylcis-transisomerases (PPIs). CsA inhibits the PPI activity of CYPA through occupation of the latter’s enzymatic active site. Using an integrated approach involving CRISPR/Cas9-engineering, siRNA, BioID, co-immunoprecipitation, pathway-specific DNA repair investigations as well as protein expression-interaction analysis, we describe novel impacts of CYPA loss and inhibition of its PPI activity on DNA repair. Prompted by findings from our CYPA-BioID proximity interactome, we validate CYPA interactions with different components of the DNA end resection machinery. Moreover, we characterise a novel and direct CYPA interaction with the NBS1 component of the MRE11-RAD50-NBS1 (MRN) complex, providing evidence that the PPI function of CYPA actively influences DNA repair via direct protein-protein interaction at the level of DNA end resection. Consequently, we demonstrate that CYPA loss or inhibition impairs Homologous Recombination Repair (HRR) following DNA replication fork stalling.Additionally, we define a set of genetic vulnerabilities associated with CYPA loss and inhibition, identifying DNA replication fork protection as an important determinant of viability herein. Leveraging the novel insights into CYPA biology we have uncovered; we explore examples of how CYPA PPI inhibition may be exploited to selectively kill cells from a variety of different cancers with a shared characteristic genomic instability profile. These findings propose a potential new disease application or repurposing strategy for the non-immunosuppressive CsA analogue class of Cyclophilin inhibitors.
Multiple pathways for glucose phosphate transport and utilization support growth ofCryptosporidium parvum
Authors: Xu Rui , Beatty Wandy L. , Greigert Valentin , Witola William H. , Sibley L. David 
Published: June 2023

AbstractCryptosporidium parvumis an obligate intracellular parasite with a highly reduced mitochondrion that lacks the TCA cycle and the ability to generate ATP, making the parasite reliant on glycolysis. Genetic ablation experiments demonstrated that neither of the two putative glucose transporters CpGT1 and CpGT2 were essential for growth. Surprisingly, hexokinase was also dispensable for parasite growth while the downstream enzyme aldolase was required, suggesting the parasite has an alternative way of obtaining phosphorylated hexose. Complementation studies inE. colisupport a role for direct transport of glucose-6-phosphate from the host cell by the parasite transporters CpGT1 and CpGT2, thus bypassing a requirement for hexokinase. Additionally, the parasite obtains phosphorylated glucose from amylopectin stores that are released by the action of the essential enzyme glycogen phosphorylase. Collectively, these findings reveal thatC. parvumrelies on multiple pathways to obtain phosphorylated glucose both for glycolysis and to restore carbohydrate reserves.
Glycoprofiling of proteins as prostate cancer biomarkers: a multinational population study
Authors: Pinkeova Andrea , Tomikova Adela , Bertokova Aniko , Fabinyova Eva , Bartova Radka , Jane Eduard , Hroncekova Stefania , Sievert Karl-Dietrich , Sokol Roman , Jirasko Michal , Kucera Radek , Eder Iris E. , Horninger Wolfgang , Klocker Helmut , Ďubjaková Petra , Fillo Juraj , Bertok Tomas , Tkac Jan 
Published: June 2023

AbstractThe glycoprofiling of two proteins, the free form of the prostate-specific antigen (fPSA) and zinc-α-2-glycoprotein (ZA2G), was assessed to determine their suitability as prostate cancer (PCa) biomarkers. The glycoprofiling of proteins was performed by analysing changes in the glycan composition on fPSA and ZA2G using lectins (proteins recognising glycans,i.e. complex carbohydrates). The specific glycoprofiling of the proteins was performed using magnetic beads (MBs) modified with horseradish peroxidase (HRP) and antibodies that selectively enriched fPSA or ZA2G from human serum samples. Subsequently, the antibody-captured glycoproteins were incubated on lectin-coated ELISA plates. In addition, a novel glycoprotein standard (GPS) was used to calibrate the assay. The glycoprofiling of fPSA and ZA2G was performed in human serum samples obtained from men undergoing prostate biopsy after an elevated serum PSA, and prostate cancer patients with or without prior therapy. The results are presented in the form of a ROC (Receiver Operating Curve). A DCA (Decision Curve Analysis) to evaluate the clinical performance and net benefit of fPSA glycan-based biomarkers was also performed. While the glycoprofiling of ZA2G showed little promise as a potential PCa biomarker, the glycoprofiling of fPSA would appear to have significant clinical potential. Hence, the GIA (Glycobiopsy ImmunoAssay) test integrates the glycoprofiling of fPSA (i.e. two glycan forms of fPSA). The GIA test could be used for early diagnoses of PCa (AUC=0.84; n=501 samples) with a potential for use in therapy-monitoring (AUC=0.85; n=168 samples). Moreover, the analysis of a subset of serum samples (n=215) revealed that the GIA test (AUC=0.81) outperformed the PHI (Prostate Health Index) test (AUC=0.69) in discriminating between men with prostate cancer and those with benign serum PSA elevation.
The Sde phosphoribosyl-linked ubiquitin transferases exploit reticulons to protect the integrity of theLegionella-containing vacuole
Authors: Zhang Mengyun , Kim Seongok , Isberg Ralph R. 
Published: June 2023

AbstractLegionella pneumophilagrows within host cells by forming a specialized membrane-bound compartment via the Icm/Dot type IV secretion system (T4SS). T4SS translocated Sde proteins promote phosphoribosyl-linked ubiquitination (pR-Ub) of several mammalian cell proteins, including Rtn4. In response to modification, Rtn4 forms tubular ER aggregates around theLegionella-containing vacuole (LCV). The loss ofsdetogether withsdhAresults in severe vacuole disruption at early infection timepoints. We tested if Rtn4 modification allowed it to serve as a physical barrier to protect its degradation from host-derived disruptive compartments. The challenge of thertn4-/-macrophages with ΔsdhApartially phenocopied the challenge ofrtn4+/+with ΔsdeΔsdhA, indicating that Rtn4 plays a role in protecting LCV integrity. Depletion ofrtn1inrtn4-/-macrophages potentiated vacuole permeability when compared tortn4-/-macrophages, consistent with Sde targeting multiple reticulon family members to support vacuole integrity. These results indicate thatL. pneumophilaexploits Rtn4 in cooperation with Rtn1 to establish a replication niche by promoting Sde-mediated tubular ER aggregates, arguing that these proteins function as a physical barrier during early steps of LCV biogenesis.
Prevalence of pathogenic bacteria detected by qPCR from cultured Nile tilapia (Oreochromis niloticusLinnaeus, 1758) in southwest Mexico
Authors: Soto-Rodriguez Sonia A. , Lopez Francis I. Marrujo , Aguilar-Rendon Karla G. 
Published: June 2023

ABSTRACTNile tilapia (Oreochromis niloticus) is one of the most important aquaculture species in the world. When bacteria are present in cultured tilapia but do not cause a declared disease, it makes them asymptomatic carrier organisms. Once environmental or nutritional conditions change, an outbreak may occur. This is why it is so important to detect pathogens before outbreaks occur. This is the first study that use molecular techniques based on PCR to estimate prevalence of fish pathogens in southwest Mexico. During 2018, 2019, 2020 and 2022 samples of internal organs and lesions of Nile tilapia were taken and analyzed for detection of the main bacterial tilapia pathogens using one-step PCR or qPCR. A total of 2396 samples from the internal organs of Nile tilapia pond and cage cultured come from the Mexican Pacific southwest states of Guerrero, Oaxaca and Chiapas were analyzed. Most of the sampled tilapias were apparently healthy and had no relation between the clinical signs and the pathogens detection was found. NoFrancisellasp. was detected in any sample,Staphylococcussp. was the most prevalent bacterial genus from the three states over time (from 0 to 64 %). Prevalence ofAeromonassp. was from 0 to 4.3 %, although the fish pathogenA. dhakensiswas not detected. Meanwhile,S. iniaewas only detected in Chiapas in 2019 at low prevalence (1.4 %) andS. agalactiaewas detected in the three sites at high prevalence (from 0 to 59 %). BothStreptococcuscan cause streptococcosis, the most dangerous re-emergent disease to cultured tilapia, which means a great risk for tilapia farming in Mexico.
Synaptotagmin-9 In Mouse Retina
Authors: Mesnard Chris S. , Hays Cassandra L. , Townsend Lou E. , Barta Cody L. , Gurumurthy Channabasavaiah B. , Thoreson Wallace B. 
Published: June 2023

AbstractSynaptotagmin-9 (Syt9) is a Ca2+sensor mediating fast synaptic release expressed in various parts of the brain. The presence and role of Syt9 in retina is unknown. We found evidence for Syt9 expression throughout the retina and created mice to conditionally eliminate Syt9 in a cre-dependent manner. We crossed Syt9fl/flmice with Rho-iCre, HRGP-Cre, and CMV-cre mice to generate mice in which Syt9 was eliminated from rods (rodSyt9CKO), cones (coneSyt9CKO), or whole animals (CMVSyt9). CMVSyt9mice showed an increase in scotopic electroretinogram (ERG) b-waves evoked by bright flashes with no change in a-waves. Cone-driven photopic ERG b-waves were not significantly different in CMVSyt9knockout mice and selective elimination of Syt9 from cones had no effect on ERGs. However, selective elimination from rods decreased scotopic and photopic b-waves as well as oscillatory potentials. These changes occurred only with bright flashes where cone responses contribute. Synaptic release was measured in individual rods by recording anion currents activated by glutamate binding to presynaptic glutamate transporters. Loss of Syt9 from rods had no effect on spontaneous or depolarization-evoked release. Our data show that Syt9 is acts at multiple sites in the retina and suggest that it may play a role in regulating transmission of cone signals by rods.
Homocysteine Metabolites Impair the PHF8/H4K20me1/mTOR/Autophagy Pathway by Upregulating the Expression of Histone Demethylase PHF8-targeting microRNAs in Human Vascular Endothelial Cells and Mice
Authors: Witucki Łukasz , Jakubowski Hieronim 
Published: June 2023

AbstractThe inability to efficiently metabolize homocysteine (Hcy) due to nutritional and genetic deficiencies, leads to hyperhomocysteinemia (HHcy) and endothelial dysfunction, a hallmark of atherosclerosis which underpins cardiovascular disease (CVD). PHF8 is a histone demethylase that demethylates H4K20me1, which affects the mammalian target of rapamycin (mTOR) signaling and autophagy, processes that play important roles in CVD. PHF8 is regulated by microRNA (miR) such as miR-22-3p and miR-1229-3p. Biochemically, HHcy is characterized by elevated levels of Hcy, Hcy-thiolactone andN-Hcy-protein. Here, we examined effects of these metabolites on miR-22-3p, miR-1229-3p, and their target PHF8, as well as on the downstream consequences of these effects on H4K20me1, mTOR-, and autophagy-related proteins and mRNAs expression in human umbilical vein endothelial cells (HUVEC). We found that treatments withN-Hcy-protein, Hcy-thiolactone, or Hcy upregulated miR-22-3p and miR-1229-3p, attenuated PHF8 expression, upregulated H4K20me1, mTOR, and phospho-mTOR. Autophagy-related proteins (BECN1, ATG5, ATG7, lipidated LC3-II, and LC3-II/LC3-I ratio) were significantly downregulated by at least one of these metabolites. We also found similar changes in the expression of miR-22-3p, Phf8, mTOR- and autophagy-related proteins/mRNAs in vivo in hearts ofCbs-/-mice, which show severe HHcy and endothelial dysfunction. Treatments with inhibitors of miR-22-3p or miR-1229-3p abrogated the effects of Hcy-thiolactone,N-Hcy-protein, and Hcy on miR expression and on PHF8, H4K20me1, mTOR-, and autophagy-related proteins/mRNAs in HUVEC. Taken together, these findings show that Hcy metabolites upregulate miR-22-3p and miR-1229-3p expression, which then dysregulate the PHF8/H4K20me1/mTOR/autophagy pathway, important for vascular homeostasis.
Bacterial SEAL domains undergo autoproteolysis and function in regulated intramembrane proteolysis
Authors: Brogan Anna P. , Habib Cameron , Hobbs Samuel J. , Kranzusch Philip J. , Rudner David Z. 
Published: June 2023

ABSTRACTGram-positive bacteria use SigI/RsgI-family sigma factor/anti-sigma factor pairs to sense and respond to cell wall defects and plant polysaccharides. InBacillus subtilisthis signal transduction pathway involves regulated intramembrane proteolysis (RIP) of the membrane-anchored anti-sigma factor RsgI. However, unlike most RIP signaling pathways, site-1 cleavage of RsgI on the extracytoplasmic side of the membrane is constitutive and the cleavage products remain stably associated, preventing intramembrane proteolysis. The regulated step in this pathway is their dissociation, which is hypothesized to involve mechanical force. Release of the ectodomain enables intramembrane cleavage by the RasP site-2 protease and activation of SigI. The constitutive site-1 protease has not been identified for any RsgI homolog. Here, we report that RsgI’s extracytoplasmic domain has structural and functional similarities to eukaryotic SEA domains that undergo autoproteolysis and have been implicated in mechanotransduction. We show that site-1 proteolysis inB. subtilisand Clostridial RsgI family members is mediated by enzyme-independent autoproteolysis of these SEA-like (SEAL) domains. Importantly, the site of proteolysis enables retention of the ectodomain through an undisrupted ß-sheet that spans the two cleavage products. Autoproteolysis can be abrogated by relief of conformational strain in the scissile loop, in a mechanism analogous to eukaryotic SEA domains. Collectively, our data support the model that RsgI-SigI signaling is mediated by mechanotransduction in a manner that has striking parallels with eukaryotic mechanotransducive signaling pathways.SIGNIFICANCESEA domains are broadly conserved among eukaryotes but absent in bacteria. They are present on diverse membrane-anchored proteins some of which have been implicated in mechanotransducive signaling pathways. Many of these domains have been found to undergo autoproteolysis and remain noncovalently associated following cleavage. Their dissociation requires mechanical force. Here, we identify a family of bacterial SEA-like (SEAL) domains that arose independently from their eukaryotic counterparts but have structural and functional similarities. We show these SEAL domains autocleave and the cleavage products remain stably associated. Importantly, these domains are present on membrane-anchored anti-sigma factors that have been implicated in mechanotransduction pathways analogous to those in eukaryotes. Our findings suggest that bacterial and eukaryotic signaling systems have evolved a similar mechanism to transduce mechanical stimuli across the lipid bilayer.
Global diversity and inferred ecophysiology of microorganisms with the potential for dissimilatory sulfate/sulfite reduction
Authors: Diao Muhe , Dyksma Stefan , Koeksoy Elif , Ngugi David Kamanda , Anantharaman Karthik , Loy Alexander , Pester Michael 
Published: June 2023

AbstractSulfate/sulfite-reducing microorganisms (SRM) are ubiquitous in nature, driving the global sulfur cycle. A hallmark of SRM is the dissimilatory sulfite reductase encoded by the paralogous genesdsrAB. Based on analysis of 950 mainly metagenome-deriveddsrAB-encoding genomes, we redefine the global diversity of microorganisms with the potential for dissimilatory sulfate/sulfite reduction and uncover genetic repertoires that challenge earlier generalizations regarding their mode of energy metabolism. We show: (i) 19 out of 23 bacterial and 2 out of 4 archaeal phyla harbor uncharacterized SRM, (ii) four phyla including theDesulfobacterotaharbor microorganisms with the genetic potential to switch between sulfate/sulfite reduction and sulfur oxidation, and (iii) the combination as well as presence/absence of differentdsrAB-types, dsrL-types anddsrDprovides guidance on the inferred direction of dissimilatory sulfur metabolism. We further provide an updateddsrABdatabase including >60% taxonomically resolved, uncultured family-level lineages and recommendations on existingdsrABprimers for environmental surveys. Our work summarizes insights into the inferred ecophysiology of newly discovered SRM, puts SRM diversity into context of the major recent changes in bacterial and archaeal taxonomy, and provides an up-to-date framework to study SRM in a global context.One sentence summarySulfate/sulfite reducing microorganisms are shaping Earth’s interconnected sulfur and carbon cycles since the Archaean: this legacy unfolds in 27 archaeal and bacterial phyla encountered in diverse marine, terrestrial, and deep-subsurface environments.
The use of plant extracts and bacteriophages as an alternative therapy approach in combating bacterial infections: the study of lytic phages andStevia rebaudiana
Authors: Stachurska Xymena , Mizielińska Małgorzata , Ordon Magdalena , Nawrotek Paweł 
Published: June 2023

AbstractIntroductionIn the light of the problem of antibiotic-resistance, the use of alternative combined therapies in combating bacterial-related issues has gained popularity. Therefore, using up-to-date laboratory techniques, possible interactions of lytic bacteriophages (MS2, T4 and Phi6), acetone and methanolS. rebaudianaextracts (SRa and SRm) in the bacterial environment have been examined.Material and MethodsUsing a microdilution method, phages-extracts coincubation assay, static interactions synographies and dynamic growth profile experiments in a bioreactor, it was found that the interactions in a static environment differ from interactions in a dynamic environment.ResultsDynamic conditions alter the influence of extracts in a concentration-dependent manner. The effects of the SRa and SRm extracts on bacterial growth in a dynamic environment depend on the species of the phage and bacterial host. The greatest differences were observed forE. colistrains and their phages, whereasP. syringaeand Phi6 phage reacted very similar to both extracts. The differences also emerged for single extracts withinE. colistrains and their phages.ConclusionsEvery extract type should be tested on a case-by-case basis and experiments outcomes should not be generalized before gathering data. Moreover, many varied experiments should be performed, especially when examining such multifactorial mixtures. Tested mixtures could be potentially used in multi-drug resistant bacterial infections treatments.
An engineered biosensor enables dynamic aspartate measurements in living cells
Authors: Davidsen Kristian , Marvin Jonathan S , Aggarwal Abhi , Brown Timothy A , Sullivan Lucas B 
Published: June 2023

AbstractIntracellular levels of the amino acid aspartate are responsive to changes in metabolism in mammalian cells and can correspondingly alter cell function, highlighting the need for robust tools to measure aspartate abundance. However, comprehensive understanding of aspartate metabolism has been limited by the throughput, cost, and static nature of the mass spectrometry based measurements that are typically employed to measure aspartate levels. To address these issues, we have developed a GFP-based sensor of aspartate (jAspSnFR3), where the fluorescence intensity corresponds to aspartate concentration. As a purified protein, the sensor has a 20-fold increase in fluorescence upon aspartate saturation, with dose dependent fluorescence changes covering a physiologically relevant aspartate concentration range and no significant off target binding. Expressed in mammalian cell lines, sensor intensity correlated with aspartate levels measured by mass spectrometry and could resolve temporal changes in intracellular aspartate from genetic, pharmacological, and nutritional manipulations. These data demonstrate the utility of jAspSnFR3 and highlight the opportunities it provides for temporally resolved and high throughput applications of variables that affect aspartate levels.