ARTICLE | doi:10.20944/preprints202010.0080.v1
Subject: Medicine & Pharmacology, Allergology Keywords: bioflavonoids; superoxide generation; oxidative phosphorylation; translocation
Online: 5 October 2020 (12:13:08 CEST)
In present work, the effects of bioflavonoids (ginkgetin and sciadopitysin) on stimulus-induced superoxide generation, tyrosyl and serine/threonine phosphorylation of proteins in human neutrophils, and the translocation of cytosolic compounds (p47phox, p67phox and Rac) to cell membrane were studied, which were isolated from the needles of Taxus media var. Hicksii. Meanwhile, three normal flavonoids (apigenin, quercetin and isoquercetin) were involved as contrasts. The results indicated that ginkgetin and sciadopitysin were capable of concentration-dependently inhibitory effects on the superoxide generation induced by N-formyl-methionyl-leucyl-phenylalanine (fMLP), arachidonic acid (AA) and phorbol-12-myristate 13-acetate (PMA). And they also suppressed fMLP- and AA- induced tyrosyl or PMA-induced serine/threonine phosphorylation and the translocation of cytosolic compounds (p47phox, p67phox and Rac) to cell membrane, which were in parallel with the suppression of the stimulus-induced superoxide generation. The effect of these compounds on the radical-scavenging was also investigated. Ginkgetin and sciadopitysin did not show remarkable effect on DPPH radical-scavenging activity, and they didn’t display the radical-scavenging activity on superoxide anion generated by phenagine methoxysulfate (PMS)-NADH system. Apparently, ginkgetin and sciadopitysin had great performance in pharmacological value and they are worthy of in-depth study.
Subject: Life Sciences, Biochemistry Keywords: nanopore; translocation slowdown; mobility; polymer sequencing; bandwidth; diffusion
Online: 14 January 2021 (09:53:00 CET)
A major obstacle faced by nanopore-based polymer sequencing and analysis is the high speed of translocation of an analyte (nucleotide, DNA, amino acid (AA), peptide) through the pore; the rate currently exceeds available detector bandwidth. Except for one method that uses an enzyme ratchet to sequence DNA, attempts to resolve the problem satisfactorily have been largely unsuccessful. Here a counterintuitive method based on reversing the pore voltage, and, with some analytes, increasing their mobility, is described. A simplified Fokker-Planck model shows a significant increase in translocation times for single nucleotides and AAs (up from ~10 ns to ~1 ms). More realistic simulations show that with a bi-level positive-negative pore voltage profile all four nucleotides in DNA (dAMP, dTMP, dCMP, and dGMP) and the 20 proteinogenic amino acids can be trapped inside the pore long enough for detection with bandwidths of ~1-10 Khz.
ARTICLE | doi:10.20944/preprints202107.0596.v1
Subject: Medicine & Pharmacology, Allergology Keywords: schizophrenia; neuro-immune; inflammation; physiological stress; bacterial translocation; psychiatry; LPS
Online: 27 July 2021 (09:16:54 CEST)
There is evidence that schizophrenia is characterized by activation of the immune-inflammatory response (IRS) and compensatory immune-regulatory (CIRS) systems and lowered neuroprotection. Studies performed on antipsychotic-naïve first episode psychosis (AF-FEP) and schizophrenia (FES) patients are important as they may disclose the pathogenesis of the disease. However, the interactome of FEP/FES is not well delineated. The aim of the current study was to delineate the characteristics of the protein-protein interaction (PPI) network of AN-FEP and its transition to FES and the biological functions, pathways, and molecular patterns, which are over-represented in FEP/FES. PPI network analysis shows that FEP and FEP/FES are strongly associated with a response to a bacterium, TNF, NFκB, RELA, SP1, JAK-STAT, death receptor and TLR4 signaling, and tyrosine phosphorylation of STAT proteins. Specific molecular complexes of the peripheral immune response are associated with microglial activation, neuroinflammation and gliogenesis. FEP/FES is accompanied by lowered protection against inflammation in part attributable to dysfunctional miRNA maturation, deficits in neurotrophin/Trk, RTK and Wnt/catenin signaling and adherens junction organization. Lowered neuroprotection due to reduced neurotrophin/Trk and Wnt/catenin signaling, and DISC1 expression and multiple interactions between lowered BDNF, CDH1, CTNNB, and DISC1 expression, increase the vulnerability to the neurotoxic effects of immune products including cytokines and complement factors. All pathways or molecular patterns enriched in the interactome of FEP/FES are directly or indirectly affected by LPS. In summary: FEP appears to be triggered by a biotic stimulus (e.g. Gram-negative bacteria) which may induce neuro-immune toxicity cascades especially when anti-inflammatory and neurotrophic protections are deficient.
ARTICLE | doi:10.20944/preprints202101.0623.v1
Subject: Medicine & Pharmacology, Allergology Keywords: depression; neuroimmune; inflammation; oxidative and nitrosative stress; autoimmune; bacterial translocation
Online: 29 January 2021 (13:17:48 CET)
The approach towards myalgic encephalomyelitis / chronic fatigue syndrome (ME/CFS) remains in a permanent state of crisis with fierce competition between the psychosocial school, which attributes ME/CFS to the perception of effort, and the medical approach (Maes and Twisk, BMC Med, 2010,8,35). The aim of this paper is to review how to construct a nomothetic model of ME/CFS using Partial Least Squares (PLS) path analysis and ensembling causome (bacterial translocation as assessed with IgM/IgA responses to LPS), protectome (lowered coenzyme Q10), adverse outcome pathways (AOP) including increased lysozyme, CD38+ T cell activation, cell-mediated immune activation (CMI), and IgM responses to oxidative specific epitopes and NO-adducts (IgM OSENO). Using PLS, we trained, tested and validated this knowledge- and data-driven causal ME/CFS model, which showed adequate convergence, construct and replicability validity. This bottom-up explicit data model of ME/CFS objectivates the descriptive narratives of the ME/CFS phenome, using causome-protectome-AOP data, whereby the abstract concept ME/CFS is translated into pathways, thereby securing the reification of the ME/CFS phenome. We found that 31.6% of the variance in the physiosomatic symptom dimension of ME/CFS was explained by the cumulative effects of CMI and CD38+ activation, IgM OSENO, IgA LPS, lysozyme (all positive) and coenzyme Q10 (inversely). Cluster analysis performed on the PLS-generated latent vector scores of all feature sets exposed three distinct immune groups of ME/CFS, namely one with increased lysozyme, one with increased CMI + CD38 activation + depressive symptoms, and another with increased bacterial translocation + autoimmune responses to OSENO.
REVIEW | doi:10.20944/preprints202004.0465.v1
Subject: Biology, Plant Sciences Keywords: terrestrial orchids; threatened; species; conservation; propagation; translocation; pollination; adaptive management
Online: 26 April 2020 (02:17:01 CEST)
This paper presents a comprehensive and adaptive framework for orchid conservation programs illustrated with data from published and unpublished case studies. There is a specific focus on West Australian terrestrial orchids, but many of the approaches have universal relevance. Aspects of the framework include (1) setting appropriate objectives, (2) establishing effective collaborations between scientists, volunteers and regulators to fill knowledge and funding gaps, (3) use of survey and demographics data to determine extinction risks and management requirements for species, (4) effective habitat management to overcome threats such as grazing, (5) finding potential new habitats by modelling climate and site data, (6) investigating the effectiveness of pollinators and (7) using seed baiting to detect mycorrhizal fungi. The relative cost and effectiveness of different methods used to propagate orchids for translocation are compared. Methods known to be successful, in order of complexity, include placement of seed in situ, vegetative propagation, symbiotic germination in non-sterile organic matter, symbiotic germination in sterile culture, asymbiotic sterile germination and clonal division in tissue culture. These form a continuum of complexity, cost, time required, faculties needed, as well as the capacity to maintain genetic diversity and produce seedlings preadapted to survive in situ. They all start with seed collection and lead to seed storage, living collections used as tuber banks and seed orchards, as well as translocation for conservation. They could also lead to commercial availability and sustainable ecotourism, both of which are needed to reduce pressure on wild plants. Overall, there has been a strong preference to use relatively complex, expensive and time-consuming methods for orchid conservation, despite evidence that simpler approaches have also been successful. These simpler methods, which include in situ seed placement and non-sterile germination on inorganic substrates, should be trialled in combination with more complex orchid propagation methods as part of an adaptive management framework. It is essential that orchid conservation projects harness the unique biological features of orchids, such as abundant seed production and mycorrhizal fungi which are far more widespread than their hosts. This is necessary to increase the efficiency and coverage of recovery actions for the largest and most threatened plant family.
ARTICLE | doi:10.20944/preprints201811.0513.v1
Subject: Biology, Agricultural Sciences & Agronomy Keywords: dryland wheat; subsoiling; sowing date; nitrogen accumulation; nitrogen translocation; yield
Online: 21 November 2018 (04:51:17 CET)
Dryland winter wheat in Loess Plateau is facing yield reduction due to shortage of soil moisture and delayed sowing time. Field experiment was conducted at Loess Plateau in Shanxi Province, China from 2012 to 2014, to study the effect of subsoiling and conventional tillage and different sowing dates on the soil water storage and contribution of N accumulation and remobilization to yield of winter wheat. The results showed that subsoiling significantly improved the soil water storage at 0-300 cm depth, improved the number of tillers and pre-anthesis N translocation in various organs of wheat and post-anthesis N accumulation, eventually increased the yield up to 17-36%. Delaying sowing time had reduced the soil water storage at sowing and winter accumulated temperature by about 180˚C. The contribution of N translocation to grain yield was maximum in glume+spike followed by in leaves and minimum by stem+sheath. In addition a close relationship was found between the N accumulation and translocation and the soil moisture in the 20-300 cm. Subsoiling during the fallow period and the medium sowing date was beneficial for improving the soil water storage and increased the N translocation to grain, thereby increasing the yield of wheat, especially in dry year.
ARTICLE | doi:10.20944/preprints201811.0227.v1
Subject: Chemistry, Other Keywords: coal fly ash; leachates; chemical species; pot culture experiments; translocation; bioconcentration
Online: 9 November 2018 (03:12:39 CET)
This study evaluated the physicochemical, mineralogical properties, mobile chemical species’ bioavailability and translocation in Brassica juncea and Spinacea oleracea L plants of a South African coal fired power utility. Coal fly ash (CFA) disposal is associated with various environmental and health risks including air, soil, surface and ground water pollution due to the leaching of toxic chemical species; these ends up in food webs affecting human health, while repeated inhalation causes bronchitis, silicosis, hair loss and lung cancer. The morphology, chemical, and mineralogical composition of CFA were determined using Scanning Electron Microscopy (SEM), X-ray fluorescence (XRF) and X-ray Diffraction, respectively. In pot culture experiments, S. oleracea L and B. juncea plants were grown in three sets of pots containing CFA (set 1), soil (set 2) and a mixture of CFA plus soil at ratio 1:1 (50% CFA: 50% soil) (set 3), while no plants were grown in set 4 as a control for the leachate samples. SEM showed that surface morphology of CFA has a lower degree of sphericity with irregular agglomerations of many particles. The XRF results revealed that CFA contains 43.65%, 22.68% and 10.89% of SiO2, Al2O3 and Fe2O3 respectively which indicate that the CFA is an alumino-silicate material. While XRD showed that the coal CFA contains mullite as a major phase followed by quartz mineral phases. Chemical species such as Fe, Mn, B, Ba and Zn were accumulated highly in most parts of the plant species. However, B. juncea showed higher potential to accumulate chemical species as compared to S. oleracea L. The bioconcentration and translocation factors (BF and TF) showed that B. juncea was the most effective in terms of bioconcentration and translocation of most of the chemical species. This indicates that B. juncea has potential in application for phytoremediation of CFA dumps and could contribute to remediation of CFA dumps and reduction of potential health and environmental impacts associated with CFA.
ARTICLE | doi:10.20944/preprints202105.0182.v1
Subject: Medicine & Pharmacology, Allergology Keywords: chronic fatigue syndrome; Myalgic encephalomyelitis; oxidative stress; neuro-immune; inflammation; bacterial translocation
Online: 10 May 2021 (12:27:45 CEST)
Background: A meaningful part of schizophrenia patients suffer from physiosomatic symptoms (formerly named psychosomatic) which are reminiscent of chronic fatigue syndrome and fibromyalgia (FF) and are associated with signs of immune activation and increased levels of tryptophan catabolites (TRYCATs). Aims: To examine whether FF symptoms in schizophrenia are associated with breakdown of the paracellular pathway, zonulin, lowered natural IgM responses to oxidative specific epitopes (OSEs); and whether FF symptoms belong to the behavioral-cognitive-physical-psychosocial-(BCPS)-worsening index consisting of indices of a general cognitive decline (G-CoDe), symptomatome of schizophrenia, and quality of life (QoL)-phenomenome. Methods: FF symptoms were assessed using the Fibromyalgia and Chronic Fatigue Rating scale in 80 schizophrenia patients and 40 healthy controls and serum cytokines/chemokines, IgA levels to TRYCATs, IgM to OSEs, zonulin and transcellular/paracellular (TRANS/PARA) molecules were assayed using ELISA methods. Results: A large part (42.3%) of the variance in the total FF score was explained by the regression on the PARA/TRANS ratio, pro-inflammatory cytokines, IgM to zonulin, IgA to TRYCATs (all positively) and IgM to OSEs (inversely). There were highly significant correlations between the total FF score and G-CoDe, symtopmatome, QoL phenomenome and BCPS-worsening score. FF symptoms belong to a common core shared by G-CoDe, symtopmatome, and QoL phenomenome. Discussion: The physio-somatic symptoms of schizophrenia are driven by various pathways including increased zonulin, breakdown of the paracellular tight-junctions pathway, immune activation with induction of the TRYCAT pathway, and consequent neurotoxicity. It is concluded that FF symptoms are part of the phenome of schizophrenia and BCPS-worsening as well.
ARTICLE | doi:10.20944/preprints202001.0349.v1
Subject: Life Sciences, Biochemistry Keywords: MAP kinase; ERK5; Bmk1; SUMO; nuclear translocation; transcription; cell proliferation; cancer; Hsp90; Cdc37
Online: 29 January 2020 (05:00:39 CET)
The MAP kinase ERK5 contains an N-terminal kinase domain and a unique C-terminal tail including a nuclear localization signal and a transcriptional activation domain. ERK5 is activated in response to growth factors and stresses, and regulates transcription at the nucleus by either phosphorylation or interaction with transcription factors. MEK5-ERK5 pathway plays an important role regulating cancer cell proliferation and survival. Therefore, it is important to define the precise molecular mechanisms implicated in ERK5 nucleo-cytoplasmic shuttling. We previously described that the molecular chaperone Hsp90 stabilizes and anchors ERK5 at the cytosol, and that ERK5 nuclear shuttling requires Hsp90 dissociation. Here, we show that MEK5 or Cdc37 overexpression -mechanisms that induce nuclear ERK5- induced ERK5 SUMO-2 modification at residues Lys6/Lys22 in cancer cells. We also show that overexpression of the SUMO protease SENP2 completely abolished endogenous ERK5 nuclear localization in response to EGF stimulation. Furthermore, mutation of these SUMO sites abolished the ability of ERK5 to translocate to the nucleus and to promote prostatic cancer PC-3 cell proliferation. These results allow us to propose a more precise mechanism: in response to MEK5 activation, ERK5 SUMOylation favors the dissociation of Hsp90 from the complex, allowing ERK5 nuclear shuttling and activation of transcription.
ARTICLE | doi:10.20944/preprints201907.0293.v1
Subject: Medicine & Pharmacology, Psychiatry & Mental Health Studies Keywords: depression; bipolar disorder; gut; bacterial translocation; LPS; oxidative stress; neuro-immune; immunology; psychiatry
Online: 26 July 2019 (00:38:09 CEST)
Major depression (MDD) is accompanied by higher serum IgM/IgA responses to LPS of Gram-negative bacteria, suggesting increased bacterial translocation and gut dysbiosis. Gut dysbiosis may occur in bipolar disorder (BD) and there are differences between MDD and BD type 1 (BP1) and -2 (BP2) in nitro-oxidative stress biomarkers associated with leaky gut. This study examines serum IgM/IgA responses directed to LPS of 6 Gram-negative bacteria in 29 BP1, 37 BP2, 44 MDD and 30 healthy individuals. MDD plus BD was best discriminated from controls by increased IgM/IgA responses to Pseudomonas aeruginosa. BP1 patients showed higher IgM responses to Morganella morganii as compared with MDD and BP2 patients. Patients with melancholia showed higher IgA responses to Citrobacter koseri as compared to controls and non-melancholic depression. The total score on the Hamilton Depression Rating Scale was significantly associated with IgA responses, especially C. koseri. IgG responses to oxidized low-density lipoprotein were significantly associated with signs of increased bacterial translocation. In conclusion, not only MDD but also BP1 and BP2 are accompanied by an immune response due to the increased load of plasma LPS of gut commensal bacteria while these aberrations in the gut-brain axis are most pronounced in BP1 and patients with melancholic features. Activated oxidative stress pathways and autoimmune responses to oxidative specific epitopes in mood disorders may be driven by a breakdown in gut paracellular, transcellular and/or vascular pathways. If replicated, drugs that protect the integrity of the gut barrier may offer novel therapeutic opportunities for BP1 and MDD.
REVIEW | doi:10.20944/preprints201904.0155.v1
Subject: Biology, Physiology Keywords: Rho GTPases; metabolism; glucose homeostasis; GLUT4 translocation; skeletal muscle; pancreas; insulin; diabetes; ageing
Online: 13 April 2019 (05:20:35 CEST)
Rho guanosine triphosphatases (GTPases) are key regulators in a number of cellular functions, including actin cytoskeleton remodeling and vesicle traffic. Traditionally, Rho GTPases are studied because of their function in cell migration and cancer, while their roles in metabolism are less documented. However, emerging evidence implicates Rho GTPases as regulators of processes of crucial importance for maintaining metabolic homeostasis. Thus, the time is now ripe for reviewing Rho GTPases in the context of metabolic health. Rho GTPase-mediated key processes include the release of insulin from pancreatic β-cells, glucose uptake into skeletal muscle and adipose tissue, and muscle mass regulation. Through the current review, we cast light on the important role of Rho GTPases in skeletal muscle, adipose tissue, and the pancreas and mechanisms by which Rho GTPases act to regulate glucose metabolism in health and disease. We also describe challenges and goals for future research.
ARTICLE | doi:10.20944/preprints202109.0395.v1
Subject: Biology, Plant Sciences Keywords: 2,4-D; upland cotton; chromosome substitution lines; herbicide tolerance; 2,4-D absorption and translocation
Online: 23 September 2021 (08:10:50 CEST)
Upland cotton is sensitive to 2,4-dichlorophenoxyacetic acid (2,4-D), and the identification of potentially 2,4-D tolerant cotton chromosome substitution (CS) lines and understanding tolerance mechanisms provide a significant step into the development and genetic improvement of upland cotton to reduce yield loss caused by 2,4-D herbicide effects including the drifts. Experiments were conducted to understand the possible mechanism of herbicide tolerance in CS-T04-15, CS-T07, and CS-B15sh, 2,4-D herbicide-tolerant cotton CS lines compared with TM-1, the 2,4-D herbicide susceptible recurrent parent of the CS line as control, using [14C]2,4-D. Percent absorption rate and translocation patterns of the 14C-labeled herbicide application at 5.17 kBq at 6 to 48 hours after treatment (HAT) were determined. The tolerant cotton CS lines showed 15-19% [14C]2,4-D uptake while TM-1 exhibited a reduced uptake of only 1.4% [14C]2,4-D at 24 HAT. Distribution of the absorbed [14C]2,4-D showed that 2-5% was translocated outside the treated leaf. In TM-1, 77% of the herbicide was translocated above and below the treated leaf, contrasting with the reduced translocation of 14C-labeled herbicide observed in the tolerant CS lines. Interestingly, CS-T04-15 showed a restricted movement of 14C below the treated leaf at 6 to 48 HAT, suggesting a novel mechanism of herbicide tolerance. This finding is the first report on upland cotton demonstrating a complex differential uptake and translocation associated with herbicide tolerance for [14C]2,4-D in cotton CS lines.
ARTICLE | doi:10.20944/preprints201901.0139.v1
Subject: Life Sciences, Microbiology Keywords: E. coli O157:H7; non-intact beef; mechanical tenderization; blade tenderization; antimicrobial interventions; translocation
Online: 14 January 2019 (12:15:28 CET)
The USDA-FSIS considers mechanically-tenderized beef as ‘non-intact’ and a food safety concern because of the potential for translocation of surface E. coli O157:H7 into the interior of the meat that may be cooked ‘rare or medium-rare’ and consumed. We evaluated 14 potential spray interventions on E. coli O157:H7-inoculated lean beef wafers (~106 CFU/cm2 , n=80) passing through a spray system (18 sec dwell time, ~40 PSI) integrated into the front end of a Ross TC-700MC tenderizer. Inoculated and processed beef wafers were stomached with D/E neutralizing broth and plated immediately, or were held in refrigerated storage for 1-, 7-, or 14 days prior to microbial plating. Seven antimicrobials that showed better performance in preliminary screening on beef wafers were selected for further testing on beef subprimals in conjunction with blade tenderization. Boneless top sirloin beef subprimals were inoculated at ~2 x 104 CFU/cm2 with a four-strain cocktail of Escherichia coli O157:H7 and passed once, lean side up, through an integrated spray system and blade tenderizer. Core samples obtained from each subprimal were examined for the presence/absence of E. coli O157:H7. Absence of E coli O157:H7 translocated into core samples correlated with the ability of the antimicrobials to reduce bacterial levels on the surface of beef prior to blade tenderization.
REVIEW | doi:10.20944/preprints202108.0411.v1
Subject: Life Sciences, Biotechnology Keywords: abiotic stresses; gene-expression; genomics; ion homeostasis; plant growth and development; plasma membrane; sugar translocation
Online: 20 August 2021 (11:43:31 CEST)
Membrane transporters (MTs) are mainly localized at the plasma membrane (PM), tonoplast and vacuolar membrane (VM) of cells in all plant organs. Their work is to maintain the cellular homeostasis by controlling ionic movements across PM channels from roots to upper plant parts, xylem loading and remobilization of sugar molecules from photosynthesis tissues in the leaf (source) to roots, stem and seeds (sink) via phloem loading. The plant’s whole source-to-sink relationship is regulated by multiple transporting proteins in a highly sophisticated manner and driven based on different stages of plant growth and development (PG&D), and environmental changes. The MTs play a pivotal role in PG&D in terms of increased plant height, branches/tiller numbers, enhanced numbers, length and filled panicles per plant, seed yield and grain quality. Dynamic climatic changes disturbed the ionic balance (salt, drought and heavy metals) and sugar supply (cold and heat stress). Due to poor selectivity, some of the MTs also uptake toxic elements in the roots that negatively impact on PG&D, later on also exported to upper parts and then deteriorate the grain quality. As an adaptive strategy, in response to salt and HMs plants activated PM and VM localized MTs that export toxic elements into vacuole, and also translocate in the root’s tips and shoot. However, in case of drought, cold and heat stresses, MTs increased the water and sugar supply to all organs. In this review, we mainly reviewed recent literature from Arabidopsis, halophytes, and major field crops such as rice, wheat, maize and oilseed rape to argue on the global role of MTs in PG&D and abiotic stress tolerance. We also discussed the gene expression level changes and genomic variations within a species as well as within a family in response to developmental and environmental cues.
REVIEW | doi:10.20944/preprints201909.0144.v1
Subject: Life Sciences, Immunology Keywords: immunoglobulin; IVIG; LcrV; PcrV; translocation; type III secretory toxin; type III secretion system; V-antigen
Online: 14 September 2019 (19:18:28 CEST)
The mechanisms underlying the effects of γ-globulin therapy for bacterial infections are thought to involve bacterial cell lysis via complement activation, phagocytosis via bacterial opsonization, toxin neutralization, and antibody-dependent cell-mediated cytotoxicity. Nevertheless, recent advances in the study of pathogenicity in gram-negative bacteria have raised the possibility of an association between γ-globulin and bacterial toxin secretion. Over time, new toxin secretion systems like the type III secretion system have been discovered in many pathogenic gram-negative bacteria. With this system, the bacterial toxins are directly injected into the cytoplasm of the target cell through a special secretory apparatus without any exposure to the extracellular environment and, therefore, with no opportunity for antibodies to neutralize the toxin. However, because antibodies against the V-antigen, which is located on the needle-shaped tip of the bacterial secretion apparatus, can inhibit toxin translocation, this raises the hope that the toxin might be susceptible to antibody targeting. Because multi-drug resistant bacteria are now prevalent, inhibiting this secretion mechanism is attractive as an alternative or adjunctive therapy against lethal bacterial infections. Thus, it would not be unreasonable to define the blocking effect of anti-V-antigen antibodies as the fifth mechanism for immunoglobulin action against bacterial infections.
Subject: Chemistry, Organic Chemistry Keywords: Type 2 diabetes; glycogen phosphorylase; anomeric spironucleosides; 1,6-dioxa-4-azaspiro[4.5]decane; [1,5]-radical translocation
Online: 18 June 2019 (10:26:30 CEST)
In the case of type 2 diabetes, inhibitors of glycogen phosphorylase (GP) might prevent unwanted glycogenolysis under high glucose conditions and thus aim at the reduction of excessive glucose production by the liver. Anomeric spironucleosides, such as hydantocidin, present a rich synthetic chemistry and important biological function, e.g., inhibition of GP. Herein, the Suárez radical methodology is successfully applied to synthesize the first example of a 1,6-dioxa-4-azaspiro[4.5]decane system, not been previously constructed via a radical pathway, starting from 6-hydroxymethyl-b-D-glucopyranosyluracil. It is shown that in the rigid pyranosyl conformation the required [1,5]-radical translocation is a minor process. The stereochemistry of the spirocycles obtained was unequivocally determined based on the chemical shifts of key sugar protons in the 1H NMR spectra. The two spirocycles were found to be modest inhibitors of RMGPb.
ARTICLE | doi:10.20944/preprints202008.0687.v1
Subject: Biology, Other Keywords: gross chromosomal rearrangements; non-homologous end joining; translocation; Illumina MiSeq; Oxford Nanopore; kluyveromyces marxianus; saccharomyces cerevisiae; URA3 gene
Online: 31 August 2020 (02:54:09 CEST)
Kluyveromyces marxianus (K. marxianus) is a newly emerging industrially relevant yeast. It is known to possess a highly efficient Non-Homologous End Joining (NHEJ) pathway that promotes random integration of non-homologous DNA fragments into its genome. The nature of the integration events was traditionally analyzed by Southern blot hybridization. However, the precise DNA sequence at the insertion sites were not fully explored. We transformed a PCR product of the Saccharomyces cerevisiae URA3 gene (ScURA3) into an uracil auxotroph K. marxianus wildtype strain and picked 24 stable Ura+ transformants for sequencing analysis. We took advantage of rapid advances in DNA sequencing technologies and developed a method using a combination of Illumina MiSeq and Oxford Nanopore sequencing. This approach enables us to uncover the Gross Chromosomal Rearrangements (GCRs) that are associated with the ScURA3 random integration. Moreover, it will shine a light on understanding DNA repair mechanisms in Eukaryotes, which could potentially provide insights for cancer research.
REVIEW | doi:10.20944/preprints201901.0081.v1
Subject: Life Sciences, Molecular Biology Keywords: gene regulation; translation; mRNA; IRES; ITAF; hnRNP; chaperone; stress; nucleocytoplasmic translocation; ribosome; lncRNA; translation initiation factor; P-bodies
Online: 9 January 2019 (08:49:45 CET)
The cellular stress response corresponds to the molecular changes that cell undergoes in response to various environmental stimuli. It induces drastic changes in the regulation of gene expression, at transcriptional and post-transcriptional levels. Actually, translation is strongly affected with a blockade of the classical cap-dependent mechanism, whereas alternative mechanisms are activated to support translation of specific mRNAs. One of the major mechanisms involved in stress-activated translation is the internal ribosome entry site (IRES)-driven initiation. IRESs, first discovered in viral mRNAs, are present in cellular mRNAs coding for master regulators of cell responses, whose expression must be tightly controlled. IRESs allow translation of these mRNAs in response to different stresses, including DNA damage, amino-acid starvation, hypoxia or endoplasmic reticulum stress, as well as to physiological stimuli such as cell differentiation or synapse network formation. Importantly, cellular mRNA IRESs are regulated by IRES trans-acting factor (ITAFs), exerting their action by at least nine different mechanisms. This review presents an update of the reported ITAFs regulating cellular mRNA translation and of the different mechanisms allowing them to control translation initiation in specific conditions. The impact of ITAFs on coordinated expression of mRNA families and consequences in cell physiology and diseases are also highlighted.
REVIEW | doi:10.20944/preprints201711.0007.v1
Subject: Life Sciences, Virology Keywords: human papillomavirus; HPV16; L2; subcellular trafficking; mitosis; transmembrane domain; translocation; membrane penetration; toxin; fusion peptide; gamma secretase; retromer
Online: 1 November 2017 (05:05:31 CET)
Beginning in 2012, our understanding of human papillomavirus (HPV) subcellular trafficking has undergone a drastic paradigm shift. Work from multiple laboratories has revealed that HPV has evolved a unique means to deliver its viral genome (vDNA) to the cell nucleus, relying on a myriad of host cell proteins and processes. The major breakthrough finding from these recent endeavors was the realization of L2-dependent utilization of cellular sorting factors for the retrograde transport of vDNA away from degradative endo/lysosomal compartments to the Golgi, prior to mitosis-dependent nuclear accumulation of L2/vDNA. An overview of current models of HPV entry, subcellular trafficking, and the role of L2 during initial infection is provided below, highlighting unresolved questions and gaps in knowledge.
ARTICLE | doi:10.20944/preprints202111.0414.v1
Subject: Biology, Physiology Keywords: endoplasmic reticulum; lipid droplets; peroxisomes; PEX3; protein targeting; membrane protein insertion; protein translocation; label-free quantitative mass spectrometry; differential protein abundance analysis; Zellweger syndrome
Online: 23 November 2021 (09:23:16 CET)
Protein import into the endoplasmic reticulum (ER) is the first step in the biogenesis of about 10,000 different soluble and membrane proteins in humans. It involves co- or post-translational targeting of precursor polypeptides to the ER and their subsequent membrane insertion or translocation. So far, three pathways for ER targeting of precursor polypeptides plus four pathways for ER targeting of mRNAs were described. Typically, these pathways deliver their substrates to the Sec61 polypeptide-conducting channel in the ER membrane. Next, the precursor polypeptides are inserted into the ER membrane or translocated into the ER lumen, which may involve auxiliary translocation components, such as the TRAP and Sec62/Sec63 complexes, or auxiliary membrane protein insertases, such as EMC and the TMCO1 complex. Recently, the PEX19/PEX3-dependent pathway, which has a well-known function in targeting and inserting various peroxisomal membrane proteins into pre-existent peroxisomal membranes, was also found to act in targeting and, putatively, inserting monotopic hairpin proteins into the ER. These either remain in the ER as resident ER membrane proteins or are pinched off from the ER as components of new lipid droplets. Therefore, the question arose if this pathway may play a more general role in ER protein targeting, i.e. represents a fourth pathway for ER targeting of precursor polypeptides. Thus, we addressed the client spectrum of the PEX19/PEX3-dependent pathway in both PEX3-depleted HeLa cells and PEX3-deficient Zellweger patient fibroblasts by an established approach, which involves label-free quantitative mass spectrometry of the total proteome of depleted or deficient cells and differential protein abundance analysis. The negatively affected proteins included twelve peroxisomal proteins and two hairpin proteins of the ER, thus confirming two previously identified classes of putative PEX19/PEX3-clients in human cells. Interestingly, fourteen collagen-related proteins with signal peptides or N-terminal transmembrane helices and belonging to the secretory pathway were also negatively affected by PEX3-deficiency, which may suggest compromised collagen biogenesis as a hitherto unknown contributor to organ failures in the respective Zellweger patients.