Zinc Transporter Research Articles

Membrane transport of rare earth metal ions N, N’-bisdioctylphosphorylmethyl-1,4-diaminobutane by the active transport mechanism

The method of membrane extraction for rare earth elements (REE) using the carrier N, N’-bisdioctylphosphorylmethyl-1,4-diaminobutane (DPDA-8) ions, leveraging an active transport mechanism, has been optimized. The distribution of DPDA-8 within the membrane pores was confirmed using scanning electron microscopy (SEM). Both light and heavy REE extractions from a simulated solution using DPDA-8 and trioctylphosphine oxide (TOPO) in 1,2-dichlorobenzene were explored. The influence of feed solution pH, carrier and anion concentration, supported liquid membrane (SLM) type and stability, and cell temperature, among other parameters, were meticulously examined. The results indicate a decrease in permeability with a decrease in feed solution pH attributed to competitive acid transport through the membrane. Additionally, the formation of H-complexes between carriers and perchloric or phosphoric acids was investigated using Fourier Transform Infrared (FTIR) spectroscopy and X-ray diffraction (XRD) techniques. Changes in the stretching bands of amino and phosphine oxide groups in DPDA-8 were monitored and characterized. XRD analysis of the crystal structures of salts DPDA-6·ClO4 and DPDA-12·H2PO4 revealed a two-dimensional layered supramolecular organization with varying anion positions relative to hydrophobic fragments. The DPDA-8 carrier demonstrated significantly higher permeability values compared to TOPO, with differences in permeability exceeding twofold for certain metals. FTIR spectroscopy identified the main coordination centers of DPDA-8 during the liquid–liquid extraction of specific REEs.

Effect of the estrous cycle on zinc transporter expression in bovine cumulus-oocyte complexes and oviduct epithelial cells.

During the luteal and follicular phases of the estrous cycle, cumulus-oocyte complexes (COC) and oviduct epithelial cells (OEC) undergo notable physiological and morphological changes. Maintaining proper zinc (Zn) homeostasis is crucial in both somatic and germinal mammalian cells. This study aimed to assess the impact of the estrous phase (luteal or follicular) on Zn transporter expression in bovine COC and OEC (BOEC). The expression of Zn transporters Slc39a6 (ZIP6), Slc39a8 (ZIP8), Slc39a14 (ZIP14), Slc30a3 (ZnT3), Slc30a7 (ZnT7), and Slc30a9 (ZnT9) was analyzed in COC and BOEC from cows during the luteal or follicular phases. Gene expression of ZIP6, ZIP14, and ZnT9 was quantified in COC and BOEC. The gene expression in the remaining transporters could not be quantified due to low mRNA levels (ZIP8 and ZnT3 in COC and BOEC; ZnT7 in BOEC) or absence of expression (ZnT7 in COC). In COC, the relative expression (RE) of all three transporters was higher in the luteal phase compared to the follicular phase (P ≤ 0.05). In BOEC, the luteal phase increased the RE of ZIP 6 (P ≤ 0.05), decreased the RE of ZnT9 (P ≤ 0.05), and did not modify the RE of ZIP14 (P > 0.05) compared to the follicular phase. In conclusion, the study reveals differences in the gene expression of ZIP6, ZIP14, and ZnT9 according to the estrous cycle phase in ex vivo samples of bovine COC and OEC.

SLC30A9: an evolutionarily conserved mitochondrial zinc transporter essential for mammalian early embryonic development

SLC30A9 (ZnT9) is a mitochondria-resident zinc transporter. Mutations in SLC30A9 have been reported in human patients with a novel cerebro-renal syndrome. Here, we show that ZnT9 is an evolutionarily highly conserved protein, with many regions extremely preserved among evolutionarily distant organisms. In Drosophila melanogaster (the fly), ZnT9 (ZnT49B) knockdown results in acutely impaired movement and drastic mitochondrial deformation. Severe Drosophila ZnT9 (dZnT9) reduction and ZnT9-null mutant flies are pupal lethal. The phenotype of dZnT9 knockdown can be partially rescued by mouse ZnT9 expression or zinc chelator TPEN, indicating the defect of dZnT9 loss is indeed a result of zinc dyshomeostasis. Interestingly, in the mouse, germline loss of Znt9 produces even more extreme phenotypes: the mutant embryos exhibit midgestational lethality with severe development abnormalities. Targeted mutagenesis of Znt9 in the mouse brain leads to serious dwarfism and physical incapacitation, followed by death shortly. Strikingly, the GH/IGF-1 signals are almost non-existent in these tissue-specific knockout mice, consistent with the medical finding in some human patients with severe mitochondrial deficiecny. ZnT9 mutations cause mitochondrial zinc dyshomeostasis, and we demonstrate mechanistically that mitochondrial zinc elevation quickly and potently inhibits the activities of respiration complexes. These results reveal the critical role of ZnT9 and mitochondrial zinc homeostasis in mammalian development. Based on our functional analyses, we finally discussed the possible nature of the so far identified human SLC30A9 mutations.

Autoantibodies against beta cells to predict early insulin requirements in pediatric patients with clinically diagnosed type 2 diabetes.

Autoimmunity has emerged as a probable disease modifier in patients with clinically diagnosed type 2 diabetes mellitus (T2DM), that is, patients who have insulin resistance, obesity, and other cardiovascular risk factors, suggesting that the presence of glutamic acid decarboxylase (anti-GAD65), islet antigen 2 (anti-IA2), and zinc transporter 8 (anti-Zn8T) antibodies could have deleterious effects on beta cell function, causing failure and earlier requirement for insulin treatment. To evaluate anti-GAD65, anti-IA2 and anti-Zn8T as predictors of early insulin requirement in adolescents with a clinical diagnosis of T2DM. This was a case-control study in patients with clinically diagnosed with T2DM (68 cases and 64 controls with and without early insulin dependence respectively), male and female, aged 12-18 years. Somatometry, blood pressure, glucose, insulin, C-peptide, glycated hemoglobin A1c, and lipid profiles were assessed. ELISA was used to measure anti-GAD65, anti-IA2, and anti-Zn8T antibodies. Descriptive statistics, Pearson's χ 2 test, Student's t test, and logistic regression was performed. P < 0.05 was considered statistically significant. There were 132 patients (53.8% female), with a mean age was 15.9 ± 1.3 years, and there was a disease evolution time of 4.49 ± 0.88 years. The presence of anti-GAD65, anti-IA2, and anti-Zn8T positivity was found in 29.5%, 18.2%, and 15.9%, respectively. Dividing the groups by early or no insulin dependence showed that the group with insulin had a higher frequency of antibody positivity: anti-GAD65 odds ratio (OR): 2.42 (1.112-5.303, P = 0.026); anti-IA2: OR: 1.55 (0.859-2.818, P = 0.105); and anti-Zn8T: OR: 7.32 (2.039-26.279, P = 0.002). Anti-GAD65 positivity was high in our study. Anti-GAD65 and anti-Zn8T positivity showed a significantly depleted beta cell reserve phenotype, leading to an increased risk of early insulin dependence.

Determination of metal ion transport rate of human ZIP4 using stable zinc isotopes

The essential microelement zinc is absorbed in the small intestine mainly by the zinc transporter ZIP4, a representative member of the Zrt/Irt-like protein (ZIP) family. ZIP4 is reportedly upregulated in many cancers, making it a promising oncology drug target. To date, there have been no reports on the turnover number of ZIP4, which is a crucial missing piece of information needed to better understand the transport mechanism. In this work, we used a non-radioactive zinc isotope, 70Zn, and inductively coupled plasma mass spectrometry (ICP-MS) to study human ZIP4 (hZIP4) expressed in HEK293 cells. Our data showed that 70Zn can replace the radioactive 65Zn as a tracer in kinetic evaluation of hZIP4 activity. This approach, combined with the quantification of the cell surface expression of hZIP4 using biotinylation or surface-bound antibody, allowed us to estimate the apparent turnover number of hZIP4 to be in the range of 0.08-0.2 s-1. The turnover numbers of the truncated hZIP4 variants are significantly smaller than that of the full-length hZIP4, confirming a crucial role for the extracellular domain in zinc transport. Using 64Zn and 70Zn, we measured zinc efflux during the cell-based transport assay and found that it has little effect on the zinc import analysis under these conditions. Finally, we demonstrated that use of laser ablation (LA) ICP-TOF-MS on samples applied to a solid substrate significantly increased the throughput of the transport assay. We envision that the approach reported here can be applied to the studies of metal transporters beyond the ZIP family.

Mesenchymal stem cell-derived extracellular vesicles mitigate neuronal damage from intracerebral hemorrhage by modulating ferroptosis

BackgroundHemorrhagic stroke is a devastating cerebrovascular event with a high rate of early mortality and long-term disability. The therapeutic potential of mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) for neurological conditions, such as intracerebral hemorrhage (ICH), has garnered considerable interest, has garnered considerable interest, though their mechanisms of action remain poorly understood.MethodsEVs were isolated from human umbilical cord MSCs, and SPECT/CT was used to track the 99mTc-labeled EVs in a mouse model of ICH. A series of comprehensive evaluations, including magnetic resonance imaging (MRI), histological study, RNA sequencing (RNA-Seq), or miRNA microarray, were performed to investigate the therapeutic action and mechanisms of MSC-EVs in both cellular and animal models of ICH.ResultsOur findings show that intravenous injection of MSC-EVs exhibits a marked affinity for the ICH-affected brain regions and cortical neurons. EV infusion alleviates the pathological changes observed in MRI due to ICH and reduces damage to ipsilateral cortical neurons. RNA-Seq analysis reveals that EV treatment modulates key pathways involved in the neuronal system and metal ion transport in mice subjected to ICH. These data were supported by the attenuation of neuronal ferroptosis in neurons treated with Hemin and in ICH mice following EV therapy. Additionally, miRNA microarray analysis depicted the EV-miRNAs targeting genes associated with ferroptosis, and miR-214-3p was identified as a regulator of neuronal ferroptosis in the ICH cellular model.ConclusionsMSC-EVs offer neuroprotective effects against ICH-induced neuronal damage by modulating ferroptosis highlighting their therapeutic potential for combating neuronal ferroptosis in brain disorders.

Zinc transporter ZnT5 is associated with epithelial mesenchymal transition via SMAD1 in breast cancer.

Zinc levels in breast cancer tissues have been reported to be higher than those in normal tissues. In addition, the expression levels of zinc transporters, including ZnT5 and ZnT6, are reportedly higher in breast cancer than in normal breast tissues. ZnT5 and ZnT6 also contribute to heterodimer formation and are involved in several biological functions. However, the functions of ZnT5 and ZnT6 heterodimers in breast cancer remain unknown. Therefore, we first investigated the immunolocalization of ZnT5 and ZnT6 in pathological breast cancer specimens and in MCF-7 and T-47D breast cancer cells. Next, we used small interfering RNA to assess cell viability and migration in ZnT5 knockdown MCF-7 and T-47D cells. Immunohistochemical analysis showed that the number of ZnT5-positive breast cancer cells was inversely correlated with the pathologic N factor status. ZnT5 knockdown had no effect on cell viability in the presence of 100 μM ZnCl2 in MCF-7 and T-47D cells. In a wound healing assay, 100 μM ZnCl2 treatment inhibited cell migration of MCF-7 and T-47D cells, whereas ZnT5 knockdown promoted cell migration, decreased E-cadherin expression and increased vimentin, slug and matrix metalloproteinase 9 expression. Antibody arrays showed that ZnT5 knockdown increased the expression of SMAD1, and that dorsomorphin treatment inhibited the promotion of migratory ability induced by ZnT5 knockdown. The results of this study revealed that both ZnT5 may be involved in less aggressive breast cancer subtypes, possibly through inhibition of cell migration.

Exploring phytoremediation potential of willow NJU513 for cadmium-contaminated soil with and without epibrassinolide treatment

There has been a growing concern over soil cadmium (Cd) pollution, underscoring the importance of finding effective remediation strategies. Willow trees have emerged as promising candidates for phytoremediation of Cd-contaminated soils. Nevertheless, the specific potential of a novel willow genotype, NJU513, in remediating Cd-polluted soil remains unexplored. Hence, the primary objectives of this study were twofold: firstly, to ascertain the suitability of the willow genotype NJU513 for remediating Cd-contaminated soil; and secondly, to elevate its remediation efficciency with the application of epibrassinolide (Brs). In the pot-culture experiment without Brs, its leaf and stem Cd concentrations were 203 mg kg−1 and 65.1 mg kg−1, with a bioaccumulation factor (BCF) of 20.8 and 6.68, respectively. In the pot-culture experiment with Brs, the corresponding Cd concentrations were 226 mg kg−1 and 59.2 mg kg−1, with a BCF of 23.1 and 6.06, respectively. In addition, the extracted Cd contents were higher in the Brs treatments (1.11–1.37 mg plant−1) than in the no-Brs treatments (0.78–0.96 mg plant−1) because Brs increased the plant biomass and leaf BCF. The mechanism underlying the Cd accumulation of NJU513 leaves with and without Brs was revealed by a transcriptome analysis. The expression levels of genes related to metal ion binding, channel activity, and transporters in leaves were up-regulated, which contributed to the high Cd accumulation and stress tolerance. Analyses of soil metabolites and bacteria in the presence and absence of Brs spraying on willow leaves indicated that soil organic compounds with carboxyl and amino groups may induce Cd activation and passivation, respectively. This study provides valuable insights for developing woody plant varieties that can be used for remediating Cd-contaminated soil.

A Possible Diagnostic Value of Zinc Transporter-8 Autoantibody for Pediatrics Type 1 Diabetic Ketoacidosis; A Systematic Review

Background: Type 1 diabetes mellitus (T1D) is a health concern all around the world. One of the most dramatic features of this disease is pediatric incidence. The most important metabolic clinical feature of T1D is ketosis and ketoacidosis. The exact role of autoantibodies in T1D pathogenesis is not completely clear yet. However, the diagnostic use for these autoantibodies is established. Zinc Transporter-8 Autoantibody (ZnT8A) is assumed to be the most specific antibody for islet beta cells. The current study aims to evaluate all relevant documents with possible risk and diagnostic value of ZnT8A for pediatrics T1D ketoacidosis (DKA) by a systematic review approach. Materials and methods: Most popular electronic databases such as PubMed, Scopus, Science Direct, and Google Scholar were used for searches the following keywords pediatrics, children, diabetic ketoacidosis, type 1 diabetes mellitus, zinc transporter-8 autoantibodies without year or any other limitations. All relevant original studies that mention ZnT8A and Ketosis or DKA in the pediatric population were evaluated and in case of being matched with the criteria extracted. Results: Primary search results led to 7233 queries. By considering the study inclusion criteria and bias assessment, six studies were included. Majority of included studies suggested ZnT8A as a risk factor for DKA in T1D pediatrics. Conclusion: The current study tried to provide an overview on the conducted studies in possible risk and diagnostic value of ZnT8A for pediatrics DKA. The majority of included studies suggest the ZnT8A as a risk factor with diagnostic association to DKA, while there are some conflicting documents. We tried to highlight the importance of this research theme for future studies to maintain a more precise conclusion with clinical relevance.

Molecular Mechanisms Underlying Fast Alkali Metal Ions Transport in Poly(ionic liquid)-in-Salt Electrolytes by Using Asymmetric Trifluoromethanesulfonyl-Cyano Anion

Molecular Mechanisms Underlying Fast Alkali Metal Ions Transport in Poly(ionic liquid)-in-Salt Electrolytes by Using Asymmetric Trifluoromethanesulfonyl-Cyano Anion

Zinc transporter 1 functions in copper uptake and cuproptosis

Copper (Cu) is a co-factor for several essential metabolic enzymes. Disruption of Cu homeostasis results in genetic diseases such as Wilson's disease. Here, we show that the zinc transporter 1 (ZnT1), known to export zinc (Zn) out of the cell, also mediates Cu2+ entry into cells and is required for Cu2+-induced cell death, cuproptosis. Structural analysis and functional characterization indicate that Cu2+ and Zn2+ share the same primary binding site, allowing Zn2+ to compete for Cu2+ uptake. Among ZnT members, ZnT1 harbors a unique inter-subunit disulfide bond that stabilizes the outward-open conformations of both protomers to facilitate efficient Cu2+ transport. Specific knockout of the ZnT1 gene in the intestinal epithelium caused the loss of Lgr5+ stem cells due to Cu deficiency. ZnT1, therefore, functions as a dual Zn2+ and Cu2+ transporter and potentially serves as a target for using Zn2+ in the treatment of Wilson's disease caused by Cu overload.

Indigenous alkaliphiles as an effective tool for bioremediation of bauxite residue (red mud)

The microorganisms thriving in ageing Bauxite residue, or red mud, have captured scientific interest for their adaptability to extreme conditions. This study investigates extremophilic microbial communities present in Indian red mud for their potential to neutralize the residue and extracting metals. These communities thrive in the highly alkaline, sodic, and metal-rich conditions of this challenging environment. The research specifically highlights alkali-halophilic species and their ability to withstand pH fluctuations (7–11) and varying NaCl levels (0–3 M). Out of the 13 isolates analyzed, all preferred a pH range of 9–10 and tolerated NaCl up to 1.5–2 M. Notably, Evansella cellulosilytica and Halalkalibacterium halodurans, showed superior tolerance index for Al3+ and Cr6+ at 2000 ppm, as well as Co2+ at 1000 ppm, followed by Sutcliffiella cohnii. However, the tolerance index for Cu2+, Te4+, and Hg2+ was relatively low for all tested strains. Additionally, Alkalihalobacillus sp. demonstrated remarkable tolerance to 10% red mud, facilitated by the production of mixed acids, neutralizing the pH within 24 h. The study proposes a potential mechanism for metal and red mud tolerance through genomic analysis using Rapid Annotation Subsystem Technology (RAST), revealing stress tolerance mechanisms, metal resistance genes, ion transporters, hydrolytic enzymes, siderophore production, and organic acid synthesis. Indigenous species like E. cellulosilytica, H. halodurans, S. cohnii, and Alkalihalobacillus sp. emerge as promising candidates for red mud bioremediation, providing insights into sustainable strategies for red mud disposal.

The Extracellular Metallometabolome: Metallophores, Metal Ionophores, and Other Chelating Agents as Natural Products

Natural products include inorganic as well as organic compounds. Living organisms face constant challenges in acquiring essential metal ions and getting rid of non-essential ones with toxic actions. They employ an extracellular biochemistry in these tasks and use it to engage in a chemical warfare against invaders and competitors by either increasing or decreasing the availability of metal ions for maintaining their welfare in aquatic or terrestrial ecological niches. To control mutualistic, cambialistic or parasitic symbiosis with other organisms they use a remarkably rich suite of secreted bioactive molecules with ligand donor atoms for metal binding. This overview discusses the interactions of these extracellular natural products with a multitude of metal ions in the periodic system of the elements. It focuses mainly on metallophores and metal ionophores secreted from bacteria, fungi, and plants, but metal-carrying cofactors and other chelating agents will also be mentioned in the context of related functions and with an intent to categorize. The intracellular fate of the metal ions and the controlled pathways for the biosynthesis, secretion, uptake, biodegradation or recycling of the secreted natural products that interact with metal ions will not be covered. Metallophores make extracellular metal ions available via delivery to specific transporters and unavailable to competing organisms, especially pathogens, though some invaders have developed ways to compete efficiently for metal ions. The classic concept of siderophores, carriers of iron(III) ions, is extended here to specific and broad-band metallophores for metal ions such as copper (chalkophores), zinc (zincophores), and yet others. Metal ionophores, in contrast, transport metal ions through biological membranes. There is a wide variety of chemical structures for either metallophores or metal ionophores. Together with physicochemical investigations of metal complexation und conditions mimicking the natural environment, “omics” mining and mapping the diversity of chemotypes is an on-going effort with analytic, genetic, and bioinformatic tools and comes together in defining the metallometabolome, which combines the metabolome and the metallome. Investigations are highly multidisciplinary, include an important, but academically infrequently crossed bridge between the biosciences (biochemistry) and the earth sciences (geochemistry), define significant applications in the pharmaceutical/medical sciences regarding immune modulation and the control of virulence at the host-pathogen interface, and have implications for the nutritional/toxicological and environmental/ecological sciences.

Genome-wide CRISPR/Cas9 screen identifies SLC39A9 and PIK3C3 as crucial entry factors for Ebola virus infection.

The Ebola virus (EBOV) has emerged as a significant global health concern, notably during the 2013-2016 outbreak in West Africa. Despite the clinical approval of two EBOV antibody drugs, there is an urgent need for more diverse and effective antiviral drugs, along with comprehensive understanding of viral-host interactions. In this study, we harnessed a biologically contained EBOVΔVP30-EGFP cell culture model which could recapitulate the entire viral life cycle, to conduct a genome-wide CRISPR/Cas9 screen. Through this, we identified PIK3C3 (phosphatidylinositide 3-kinase) and SLC39A9 (zinc transporter) as crucial host factors for EBOV infection. Genetic depletion of SLC39A9 and PIK3C3 lead to reduction of EBOV entry, but not impact viral genome replication, suggesting that SLC39A9 and PIK3C3 act as entry factors, facilitating viral entry into host cells. Moreover, PIK3C3 kinase activity is indispensable for the internalization of EBOV virions, presumably through the regulation of endocytic and autophagic membrane traffic, which has been previously recognized as essential for EBOV internalization. Notably, our study demonstrated that PIK3C3 kinase inhibitor could effectively block EBOV infection, underscoring PIK3C3 as a promising drug target. Furthermore, biochemical analysis showed that recombinant SLC39A9 protein could directly bind viral GP protein, which further promotes the interaction of viral GP protein with cellular receptor NPC1. These findings suggests that SLC39A9 plays dual roles in EBOV entry. Initially, it serves as an attachment factor during the early entry phase by engaging with the viral GP protein. Subsequently, SLC39A9 functions an adaptor protein, facilitating the interaction between virions and the NPC1 receptor during the late entry phase, prior to cathepsin cleavage on the viral GP. In summary, this study offers novel insights into virus-host interactions, contributing valuable information for the development of new therapies against EBOV infection.

Current Understanding of Androgen Signaling in Prostatitis and its Treatment: A Review.

Chronic prostatitis is a highly prevalent condition that significantly impacts the quality of life and fertility of men. Because of its heterogeneous nature, there is no definitive treatment, which requires ongoing research into its etiology. Additionally, the association between prostatitis and an elevated risk of prostate cancer highlights the importance of comprehending androgen involvement in prostatitis. This paper examines the current understanding of androgen signaling in prostatitis and explores contemporary therapeutic approaches. It was reviewed Medline articles comprehensively, using keywords such as nonbacterial prostatitis, prostatitis infertility, androgen role in prostatitis, and chronic pelvic pain. Several cellular targets are linked to androgen signaling. Notably, the major tyrosine phosphatase activity (cPAcP) in normal human prostate is influenced by androgen signaling, and its serum levels inversely correlate with prostate cancer progression. Androgens also regulate membrane-associated zinc and pyruvate transporters transduction in prostate cells, suggesting promising avenues for novel drug development aimed at inhibiting these molecules to reduce cancer tumor growth. Various therapies for prostatitis have been evaluated, including antibiotics, anti-inflammatory medications (including bioflavonoids), neuromodulators, alpha-blockers, 5α-reductase inhibitors, and androgen receptor antagonists. These therapies have demonstrated varying degrees of success in ameliorating symptoms.In conclusion, aging decreases circulating T and intraprostatic DHT, altering the proper functioning of the prostate, reducing the ability of androgens to maintain normal Zn2+ levels, and diminishing the secretion of citrate, PAcP, and other proteins into the prostatic fluid. The Zn2+-transporter decreases or is absent in prostate cancer, so the pyruvate transporter activates. Consequently, the cell ATP increases, inducing tumor growth.

Identification of Glyoxalase A in Group B Streptococcus and its contribution to methylglyoxal tolerance and virulence.

Group B Streptococcus (GBS) is a Gram-positive pathobiont that commonly colonizes the gastrointestinal and lower female genital tracts but can cause sepsis and pneumonia in newborns and is a leading cause of neonatal meningitis. Despite the resulting disease severity, the pathogenesis of GBS is not completely understood, especially during the early phases of infection. To investigate GBS factors necessary for blood stream survival, we performed a transposon (Tn) mutant screen in our bacteremia infection model using a GBS mariner transposon mutant library previously developed by our group. We identified significantly underrepresented mutations in 628 genes that contribute to survival in the blood, including those encoding known virulence factors such as capsule, the β-hemolysin, and inorganic metal ion transport systems. Most of the underrepresented genes have not been previously characterized or studied in GBS, including gloA and gloB, which are homologs for genes involved in methylglyoxal (MG) detoxification. MG is a byproduct of glycolysis and a highly reactive toxic aldehyde that is elevated in immune cells during infection. Here, we observed MG sensitivity across multiple GBS isolates and confirm that gloA contributes to MG tolerance and invasive GBS infection. We show specifically that gloA contributes to GBS survival in the presence of neutrophils and depleting neutrophils in mice abrogates the decreased survival and infection of the gloA mutant. The requirement of the glyoxalase pathway during GBS infection suggests that MG detoxification is important for bacterial survival during host-pathogen interactions.

Erg251 has complex and pleiotropic effects on sterol composition, azole susceptibility, filamentation, and stress response phenotypes.

Ergosterol is essential for fungal cell membrane integrity and growth, and numerous antifungal drugs target ergosterol. Inactivation or modification of ergosterol biosynthetic genes can lead to changes in antifungal drug susceptibility, filamentation and stress response. Here, we found that the ergosterol biosynthesis gene ERG251 is a hotspot for point mutations during adaptation to antifungal drug stress within two distinct genetic backgrounds of Candida albicans. Heterozygous point mutations led to single allele dysfunction of ERG251 and resulted in azole tolerance in both genetic backgrounds. This is the first known example of point mutations causing azole tolerance in C. albicans. Importantly, single allele dysfunction of ERG251 in combination with recurrent chromosome aneuploidies resulted in bona fide azole resistance. Homozygous deletions of ERG251 caused increased fitness in low concentrations of fluconazole and decreased fitness in rich medium, especially at low initial cell density. Homozygous deletions of ERG251 resulted in accumulation of ergosterol intermediates consistent with the fitness defect in rich medium. Dysfunction of ERG251, together with FLC exposure, resulted in decreased accumulation of the toxic sterol (14-ɑ-methylergosta-8,24(28)-dien-3β,6α-diol) and increased accumulation of non-toxic alternative sterols. The altered sterol composition of the ERG251 mutants had pleiotropic effects on transcription, filamentation, and stress responses including cell membrane, osmotic and oxidative stress. Interestingly, while dysfunction of ERG251 resulted in azole tolerance, it also led to transcriptional upregulation of ZRT2, a membrane-bound Zinc transporter, in the presence of FLC, and overexpression of ZRT2 is sufficient to increase azole tolerance in wild-type C. albicans. Finally, in a murine model of systemic infection, homozygous deletion of ERG251 resulted in decreased virulence while the heterozygous deletion mutants maintain their pathogenicity. Overall, this study demonstrates that single allele dysfunction of ERG251 is a recurrent and effective mechanism of acquired azole tolerance. We propose that altered sterol composition resulting from ERG251 dysfunction mediates azole tolerance as well as pleiotropic effects on stress response, filamentation and virulence.

A riboswitch-controlled TerC family transporter Alx tunes intracellular manganese concentration in Escherichia coli at alkaline pH.

Cells use transition metal ions as structural components of biomolecules and cofactors in enzymatic reactions, making transition metal ions integral cellular components. Organisms optimize metal ion concentration to meet cellular needs by regulating the expression of proteins that import and export that metal ion, often in a metal ion concentration-dependent manner. One such regulation mechanism is via riboswitches, which are 5'-untranslated regions of an mRNA that undergo conformational changes to promote or inhibit the expression of the downstream gene, commonly in response to a ligand. The yybP-ykoY family of bacterial riboswitches shares a conserved aptamer domain that binds manganese ions (Mn2+). In Escherichia coli, the yybP-ykoY riboswitch precedes and regulates the expression of two different genes: mntP, which based on genetic evidence encodes an Mn2+ exporter, and alx, which encodes a putative metal ion transporter whose cognate ligand is currently in question. The expression of alx is upregulated by both elevated concentrations of Mn2+ and alkaline pH. With metal ion measurements and gene expression studies, we demonstrate that the alkalinization of media increases the cytoplasmic manganese pool, which, in turn, enhances alx expression. The Alx-mediated Mn2+ export prevents the toxic buildup of the cellular manganese, with the export activity maximal at alkaline pH. We pinpoint a set of acidic residues in the predicted transmembrane segments of Alx that play a critical role in Mn2+ export. We propose that Alx-mediated Mn2+ export serves as a primary protective mechanism that fine tunes the cytoplasmic manganese content, especially during alkaline stress.IMPORTANCEBacteria use clever ways to tune gene expression upon encountering certain environmental stresses, such as alkaline pH in parts of the human gut and high concentration of a transition metal ion manganese. One way by which bacteria regulate the expression of their genes is through the 5'-untranslated regions of messenger RNA called riboswitches that bind ligands to turn expression of genes on/off. In this work, we have investigated the roles and regulation of alx and mntP, the two genes in Escherichia coli regulated by the yybP-ykoY riboswitches, in alkaline pH and high concentration of Mn2+. This work highlights the intricate ways through which bacteria adapt to their surroundings, utilizing riboregulatory mechanisms to maintain Mn2+ levels amidst varying environmental factors.

Zinc proteinate with moderate chelation strength enhances zinc absorption by upregulating the expression of zinc and amino acid transporters in primary cultured duodenal epithelial cells of broiler embryos.

Recent study showed that zinc (Zn) and amino acid transporters may be involved in enhancing Zn absorption from Zn proteinate with moderate chelation strength (Zn-Prot M) in the duodenum of broilers. However, the specific mechanisms by which Zn-Prot M promotes the above Zn absorption are unknown. Therefore, in this study, 3 experiments were conducted to investigate specific and direct effects of Zn-Prot M and Zn sulfate (ZnS) on Zn absorption and expression of related transporters in primary duodenal epithelial cells of broiler embryos so as to preliminarily address possible mechanisms. In experiment 1, cells were treated with 100 μmol Zn/L as ZnS or Zn-Prot M for 20, 40, 60, 80, 100, or 120min. Experiment 2 consisted of 3 sub-experiments. In experiment 2A, cells were treated with a Zn-unsupplemented basal medium (Control) or the basal medium supplemented with 100 or 200 μmol Zn/L as ZnS or Zn-Prot M for 60min; in experiment 2B, cells were treated with a Zn-unsupplemented basal medium (Control) or the basal medium supplemented with 200 μmol Zn/L of as the ZnS or Zn-Prot M for 120min; in experiment 2C, cells were treated with a Zn-unsupplemented basal medium (Control) or the basal medium supplemented with 400 or 800 μmol Zn/L as ZnS or Zn-Prot M for 120min. In experiment 3, cells were treated with a Zn-unsupplemented basal medium (Control) or the basal medium supplemented with 400 μmol Zn/L as ZnS or Zn-Prot M for 120min. The results of experiment 1 indicated that the minimum incubation time for saturable Zn absorption was determined to be 50.83min using the best fit line. The results in experiment 2 demonstrated that a Zn concentration of 400 μmol/L and an incubation time of 120min were suitable to increase the absorption of Zn from Zn-Prot M compared to ZnS. In experiment 3, Zn absorption across cell monolayers was significantly increased by Zn addition (P < 0.05), and was significantly greater with Zn-Prot M than with ZnS (P < 0.05). Compared to the control, Zn addition significantly decreased Zn transporter 10 and peptide-transporter 1 mRNA expression levels and increased y + L-type amino transporter 2 (y + LAT2) protein abundance (P < 0.05). Moreover, protein expression levels of zrt/irt-like protein 3 (ZIP3), zrt-irt-like protein 5 (ZIP5), and y + LAT2 were significantly greater for Zn-Prot M than for ZnS (P < 0.05). These findings suggest that Zn-Prot M promote Zn absorption by increasing ZIP3, ZIP5 and y + LAT2 protein expression levels in primary duodenal epithelial cells.

Potentials of bone marrow cells-derived from naïve or diabetic mice in autoimmune type 1 diabetes: immunomodulatory, anti-inflammatory, anti hyperglycemic, and antioxidative.

The scarcity of transplanted human islet tissue and the requirement for immunosuppressive drugs to prevent the rejection of allogeneic grafts have hindered the treatment of autoimmune type 1 diabetes mellitus (T1DM) through islet transplantation. However, there is hope in adoptively transferred bone marrow cells (BMCs) therapy, which has emerged as a propitious pathway for forthcoming medications. BMCs have the potential to significantly impact both replacement and regenerative therapies for a range of disorders, including diabetes mellitus, and have demonstrated anti-diabetic effects. The main goal of this study is to evaluate the effectiveness of adoptively transferred bone marrow cells derived from either naïve mice (nBMCs) or diabetic mice (dBMCs) in treating a T1DM mice model. Male Swiss albino mice were starved for 16 h and then injected with streptozotocin (STZ) at a dose of 40 mg/kg body weight for 5 consecutive days to induce T1DM. After 14 days, the diabetic mice were distributed into four groups. The first group served as a diabetic control treated with sodium citrate buffer, while the other three groups were treated for two weeks, respectively, with insulin (subcutaneously at a dose of 8 U/kg/day), nBMCs (intravenously at a dose of 1 × 106 cells/mouse/once), and dBMCs (intravenously at a dose of 1 × 106 cells/mouse/once). It is worth noting that administering adoptively transferred nBMCs or adoptively transferred dBMCs to STZ-induced T1DM mice resulted in a significant amelioration in glycemic condition, accompanied by a considerable reduction in the level of blood glucose and glycosylated hemoglobin % (HbA1C %), ultimately restoring serum insulin levels to their initial state in control mice. Administering nBMCs or dBMCs to STZ-induced T1DM mice led to a remarkable decrease in levels of inflammatory cytokine markers in the serum, including interferon-γ (INF-γ), tumor necrosis factor- α (TNF-α), tumor growth factor-β (TGF-β), interleukin-1 β (L-1β), interlekin-4 (IL-4), interleukin-6 (IL-6), and interleukin-10 (IL-10). Additionally, STZ-induced T1DM mice, when treated with nBMCs or dBMCs, experienced a notable rise in total immunoglobulin (Ig) level. Furthermore, there was a significant reduction in the levels of islet cell autoantibodies (ICA) and insulin autoantibodies (IAA). Furthermore, the serum of STZ-induced T1DM mice showed a significant increase in Zinc transporter 8 antigen protein (ZnT8), islet antigen 2 protein (IA-2), and glutamic acid decarboxylase antigen protein (GAD) levels. Interestingly, the administration of nBMCs or dBMCs resulted in a heightened expression of IA-2 protein in STZ-induced T1DM mice treated with nBMCs or dBMCs. Furthermore, the level of malondialdehyde (MDA) was increased, while the levels of catalase (CAT) and superoxide dismutase (SOD) were decreased in non-treated STZ-induced T1DM mice. However, when nBMCs or dBMCs were administered to STZ-induced T1DM mice, it had a significant impact on reducing oxidative stress. This was accomplished by reducing the levels of MDA in the serum and enhancing the activities of enzymatic antioxidants like CAT and SOD. STZ-induced T1DM mice displayed a significant elevation in the levels of liver enzymes ALT and AST, as well as heightened levels of creatinine and urea. Considering the crucial roles of the liver and kidney in metabolism and excretion, this research further examined the effects of administering nBMCs or dBMCs to STZ-induced T1DM mice. Notably, the administration of these cells alleviated the observed effects. The present study suggests that utilizing adoptively transferred nBMCs or adoptively transferred dBMCs in the treatment of T1DM led to noteworthy decreases in blood glucose levels, possibly attributed to their capacity to enhance insulin secretion and improve the performance of pancreatic islets. Additionally, BMCs may exert their beneficial effects on the pancreatic islets of diabetic mice through their immunomodulatory, antioxidant, anti-inflammatory, and anti-oxidative stress properties.