The labile zinc pool in plant cells

2019 ◽  
Vol 46 (9) ◽  
pp. 796 ◽  
Author(s):  
Ilya E. Zlobin ◽  
Alexander V. Kartashov ◽  
Alexander V. Nosov ◽  
Artem A. Fomenkov ◽  
Vladimir V. Kuznetsov
Keyword(s):  
Plant Cells ◽  
Zinc Content ◽  
Zinc Complexes ◽  
Zinc Homeostasis ◽  
Thellungiella Salsuginea ◽  
Intracellular Zinc ◽  
Reversible Binding ◽  
Cell Extracts ◽  
First Time ◽  
Fluorimetric Titration

Zinc is the most abundant and important transition metal in plants; however, the dynamic aspects of zinc homeostasis in plant cells are poorly understood. In this study we explored the pool of labile exchangeable zinc complexes in plant cells, and the potential influence of changes in intracellular zinc availability on cellular physiology. Work was performed on cultivated cell extracts of Arabidopsis thaliana (L.) Heynh. and Thellungiella salsuginea (Pall.) O.E. Schulz grown under control (3.48 µM Zn2+), 10-fold Zn excess or Zn starvation conditions. The free and labile Zn contents in the extracts were then determined by fluorimetric titration. We observed for the first time that plant cells contain micromolar concentrations of labile zinc complexes that account for a low percentage of the total zinc content. Labile zinc is mainly protein bound. Zn starvation inhibits cell proliferation and leads to the disappearance of the labile zinc pool, whereas Zn excess drastically increases the labile zinc pool. Free Zn2+ is buffered at picomolar concentrations in the intracellular milieu, and the increase in free Zn2+ concentrations to low nanomolar values clearly modulates enzyme activity by direct reversible binding. Such increases in free Zn2+ can be achieved by the substantial influx of additional zinc or by the oxidation of zinc-binding thiols. The observed features of the labile zinc pool in plant cells suggest it has a role in intracellular zinc trafficking and zinc signalling.

scholarly journals Zinc, metallothioneins and immunosenescence

2010 ◽  
Vol 69 (3) ◽  
pp. 290-299 ◽  
Author(s):  
E. Mocchegiani ◽  
M. Malavolta ◽  
L. Costarelli ◽  
R. Giacconi ◽  
C. Cipriano ◽  
...  
Keyword(s):  
Innate Immunity ◽  
Zinc Supplementation ◽  
Killer Cell ◽  
Zinc Content ◽  
Zinc Homeostasis ◽  
Zinc Transporters ◽  
Intracellular Zinc ◽  
Natural Killer Cell Cytotoxicity ◽  
Zip Family

Ageing is an inevitable biological process with gradual and spontaneous biochemical and physiological changes and increased susceptibility to diseases. The nutritional factor, zinc, may remodel these changes with subsequent healthy ageing, because zinc improves the inflammatory/immune response as shown by in vitro and in vivo studies. The intracellular zinc homeostasis is regulated by buffering metallothioneins (MT) and zinc transporters (ZnT and ZIP families) that mediate the intracellular zinc signalling assigning to zinc a role of ‘second messenger’. In ageing, the intracellular zinc homeostasis is altered, because high MT are unable to release zinc and some zinc transporters deputed to zinc influx (ZIP family) are defective leading to low intracellular zinc content for the immune efficiency. Physiological zinc supplementation in the elderly improves these functions. However, the choice of old subjects for zinc supplementation has to be performed in relation to the specific genetic background of MT and IL-6, because the latter is involved both in MTmRNA and in intracellular zinc homeostasis. Old subjects carrying GG genotypes (C–carriers) in the IL-6–174G/C locus display high IL-6, low intracellular zinc content, impaired innate immunity and enhanced MT. Old subjects carrying GC and CC genotypes (C+carriers) display satisfactory intracellular zinc content, adequate innate immunity and are more prone to reach longevity. Zinc supplementation in old C–carriers restores natural killer cell cytotoxicity and zinc status. The genetic variations of the IL-6−174G/C locus when associated with those of the MT1A+647A/C locus are useful tools for the choice of old people for zinc supplementation.

scholarly journals Alterations in ZnT1 expression and function lead to impaired intracellular zinc homeostasis in cancer

2019 ◽  
Vol 5 (1) ◽  
Author(s):  
Adrian Israel Lehvy ◽  
Guy Horev ◽  
Yarden Golan ◽  
Fabian Glaser ◽  
Yael Shammai ◽  
...  
Keyword(s):  
Malignant Tumors ◽  
Zinc Homeostasis ◽  
Healthy Population ◽  
Missense Mutations ◽  
Protein Alignment ◽  
Loss Of Function ◽  
Intracellular Zinc ◽  
Dismal Prognosis ◽  
Zinc Levels ◽  
And Function

Abstract Zinc is vital for the structure and function of ~3000 human proteins and hence plays key physiological roles. Consequently, impaired zinc homeostasis is associated with various human diseases including cancer. Intracellular zinc levels are tightly regulated by two families of zinc transporters: ZIPs and ZnTs; ZIPs import zinc into the cytosol from the extracellular milieu, or from the lumen of organelles into the cytoplasm. In contrast, the vast majority of ZnTs compartmentalize zinc within organelles, whereas the ubiquitously expressed ZnT1 is the sole zinc exporter. Herein, we explored the hypothesis that qualitative and quantitative alterations in ZnT1 activity impair cellular zinc homeostasis in cancer. Towards this end, we first used bioinformatics to analyze inactivating mutations in ZIPs and ZNTs, catalogued in the COSMIC and gnomAD databases, representing tumor specimens and healthy population controls, respectively. ZnT1, ZnT10, ZIP8, and ZIP10 showed extremely high rates of loss of function mutations in cancer as compared to healthy controls. Analysis of the putative functional impact of missense mutations in ZnT1-ZnT10 and ZIP1-ZIP14, using homologous protein alignment and structural predictions, revealed that ZnT1 displays a markedly increased frequency of predicted functionally deleterious mutations in malignant tumors, as compared to a healthy population. Furthermore, examination of ZnT1 expression in 30 cancer types in the TCGA database revealed five tumor types with significant ZnT1 overexpression, which predicted dismal prognosis for cancer patient survival. Novel functional zinc transport assays, which allowed for the indirect measurement of cytosolic zinc levels, established that wild type ZnT1 overexpression results in low intracellular zinc levels. In contrast, overexpression of predicted deleterious ZnT1 missense mutations did not reduce intracellular zinc levels, validating eight missense mutations as loss of function (LoF) mutations. Thus, alterations in ZnT1 expression and LoF mutations in ZnT1 provide a molecular mechanism for impaired zinc homeostasis in cancer formation and/or progression.

Heat shock but not cold shock leads to disturbed intracellular zinc homeostasis

2009 ◽  
pp. n/a-n/a ◽  
Author(s):  
Elvis Pirev ◽  
Yasemin Ince ◽  
Helmut Sies ◽  
Klaus D. Kröncke
Keyword(s):  
Heat Shock ◽  
Cold Shock ◽  
Zinc Homeostasis ◽  
Intracellular Zinc

scholarly journals Intracellular zinc homeostasis and zinc signaling

2008 ◽  
Vol 99 (8) ◽  
pp. 1515-1522 ◽  
Author(s):  
Masaaki Murakami ◽  
Toshio Hirano
Keyword(s):  
Zinc Homeostasis ◽  
Intracellular Zinc ◽  
Zinc Signaling

scholarly journals Microfilaments: dynamic arrays in higher plant cells

1987 ◽  
Vol 104 (4) ◽  
pp. 995-1004 ◽  
Author(s):  
RW Seagull ◽  
MM Falconer ◽  
CA Weerdenburg
Keyword(s):  
Three Dimensional ◽  
Plant Cells ◽  
Higher Plant ◽  
Light Microscope Level ◽  
Cell Functions ◽  
Spindle Poles ◽  
Subcortical Region ◽  
Dimensional Distribution ◽  
Transvacuolar Strands ◽  
First Time

By using fluorescently labeled phalloidin we have examined, at the light microscope level, the three-dimensional distribution and reorganization of actin-like microfilaments (mfs) during plant cell cycle and differentiation. At interphase, mfs are organized into three distinct yet interconnected arrays: fine peripheral networks close to the plasma membrane; large axially oriented cables in the subcortical region; a nuclear "basket" of mfs extending into the transvacuolar strands. All these arrays, beginning with the peripheral network, disappear at the onset of mitosis and reappear, beginning with the nuclear basket, after cytokinesis. During mitotic and cytokinetic events, mfs are associated with the spindle and phragmoplast. Actin staining in the spindle is localized between the chromosomes and the spindle poles and changes in a functionally specific manner. The nuclear region appears to be the center for mf organization and/or initiation. During differentiation from rapid cell division to cell elongation, mf arrays switch from an axial to a transverse orientation, thus paralleling the microtubules. This change in orientation reflects a shift in the direction of cytoplasmic streaming. These observations show for the first time that actin-like mfs form intricate and dynamic arrays in plant cells which may be involved in many as yet undescribed cell functions.

Aerobic purification of N5,N10-methylenetetrahydromethanopterin dehydrogenase, separated from N5,N10-methenyltetrahydromethanopterin cyclohydrolase, from Methanobacterium thermoautotrophicum strain Marburg

1989 ◽  
Vol 35 (4) ◽  
pp. 499-507 ◽  
Author(s):  
Biswarup Mukhopadhyay ◽  
Lacy Daniels
Keyword(s):  
Enzyme Activity ◽  
Ammonium Sulfate ◽  
Dehydrogenase Activity ◽  
Specific Activity ◽  
Methanobacterium Thermoautotrophicum ◽  
Anaerobic Conditions ◽  
Cell Extracts ◽  
First Time ◽  
Aerobic And Anaerobic ◽  
Aerobic And Anaerobic Conditions

The N5,N10-methylenetetrahydromethanopterin dehydrogenase from Methanobacterium thermoautotrophicum strain Marburg has been purified with reasonable yield and much higher specific activity than previously reported. For the first time it has been shown that both N5,N10-methylenetetrahydromethanopterin dehydrogenase and N5,N10-methenyltetrahydromethanopterin cyclohydrolase activities were stable under air and could be purified using aerobic operations. The dehydrogenase activity from Methanobacterium thermoautotrophicum Marburg was stable in phosphate buffer with or without glycerol or ammonium sulfate under both aerobic and anaerobic conditions. However, the presence of either 2-mercaptoethanol or dithiothreitol in the enzyme solution destroyed the enzyme activity during both aerobic and anaerobic incubations. Dehydrogenase was purified 62-fold using Phenyl-Sepharose and DEAE-Sephadex chromatography in succession under air. Both of these chromatographic methods separated dehydrogenase activity from N5,N10-methenyltetrahydromethanopterin cyclohydrolase; DEAE-Sephadex provided the best separation. Phenyl-Sepharose chromatography of the supernatant of cell extracts containing ammonium sulfate at 60% of saturation provided a 4.7-fold purification and 98% recovery of cyclohydrolase; this result established the air stability of N5,N10-methenyltetrahydromethanopterin cyclohydrolase from Methanobacterium thermoautotrophicum Marburg.Key words: methylenetetrahydromethanopterin dehydrogenase, methenyltetrahydromethanopterin cyclohydrolase, Methanobacterium, aerobic purification, oxygen stability.

scholarly journals Expression of Prostaglandin G/H Synthase Type 1, but not Type 2, in Human Ovarian Adenocarcinomas

1998 ◽  
Vol 46 (1) ◽  
pp. 77-84 ◽  
Author(s):  
Monique Doré ◽  
Lucie C. Cóté ◽  
Andrew Mitchell ◽  
Jean Sirois
Keyword(s):  
Tumor Cells ◽  
Cellular Localization ◽  
Immunoblot Analysis ◽  
Cell Extracts ◽  
Prostaglandin Endoperoxide Synthase ◽  
Degree Of Malignancy ◽  
Immunoreactive Band ◽  
First Time ◽  
Ovarian Adenocarcinomas

Prostaglandin endoperoxide synthase (PGHS) is a key rate-limiting enzyme in prostaglandin biosynthesis. PGHS has recently been shown to be expressed in human colorectal cancers and in experimental cutaneous papillomas and carcinomas. However, PGHS expression has not been investigated in ovarian cancers. The objectives of this study were to determine whether PGHS isoenzymes are expressed in human ovarian cancer and, if so, to identify which isoform is involved (PGHS-1 and/or PGHS-2) and to characterize its cellular localization. Sixteen human ovarian adenocarcinomas were studied by immunohistochemistry using specific antibodies recognizing PGHS-1 or PGHS-2. Results showed that all adenocarcinomas demonstrated the presence of tumor cells expressing PGHS-1 but not PGHS-2. Patterns of staining of tumor cells varied among different types of adenocarcinomas, with cells presenting either a mostly diffuse cytoplasmic immunoreactivity or, alternatively, a staining mainly concentrated around the nucleus. No correlation between the intensity of the immunostaining and the degree of malignancy of tumors could be established ( r −0.03: p>0.05). Immunoblot analysis with PGHS-1-selective antibodies of cell extracts from adenocarcinomas revealed the presence of a characteristic 72,000 Mr immunoreactive band. Therefore, these results show for the first time that PGHS-1 is expressed in human ovarian adenocarcinomas.

scholarly journals The influence of dietary zinc source and coccidial vaccine exposure on intracellular zinc homeostasis and immune status in broiler chickens

2015 ◽  
Vol 114 (2) ◽  
pp. 202-212 ◽  
Author(s):  
Catalina Troche ◽  
Susan D. Eicher ◽  
Todd J. Applegate
Keyword(s):  
Broiler Chickens ◽  
Immune Status ◽  
Basal Diet ◽  
Zinc Homeostasis ◽  
Phagocytic Capacity ◽  
Intracellular Zinc ◽  
Mucosal Tissues ◽  
Zn Status ◽  
Zip Transporters ◽  
Transporter Expression

Coccidia are protozoal parasites which compromise mucosal integrity of the intestine, potentiating poultry morbidity. The host's Zn status influences the course of infection. Therefore, two experiments were designed to determine how supplemental Zn regimens impacted jejunal and caecal immune status and Zn transporter expression. Coccivac®-B was administered weekly at ten times the recommended dose as a mild coccidial challenge (10CV). Zn was provided through a basal diet, supplemental zinc sulfate (ZnSO4), or a supplemental 1:1 blend of ZnSO4 and Availa®-Zn (Blend). Mucosal jejunum (Expt 1) and caecal tonsils (Expt 2) were evaluated for intracellular Zn concentrations and phagocytic capacity. Messenger expression of Zn transporters ZnT5, ZnT7, Zip9 and Zip13 were investigated to determine Zn trafficking. With 10CV, phagocytic capacity was decreased in jejunal cells by 2 %. In the caecal tonsils, however, phagocytic capacity increased with challenge, with the magnitude of increase being more pronounced with higher dietary Zn (10CV × Zn interaction; P= 0·04). Intracellular Zn within caecal tonsils was found significantly reduced with 10CV (27 %, P= 0·0001). 10CV also resulted in an overall increase in the ratio of Zip:ZnT transporters. With the exception of Zip13 transporter expression, dietary Zn source had little impact on any of the measured cellular parameters. Thus, intestinal mucosal tissues had reductions in intracellular free Zn during coccidial challenge, which was coupled with an upregulation of measured Zip transporters. This suggests that under coccidial challenge, intestinal cells attempt to compensate for the drop in intracellular Zn.

scholarly journals Lack of ZnT8 protects pancreatic islets from hypoxia- and cytokine induced cell death

2022 ◽  
Author(s):  
Maria Karsai ◽  
Richard A Zuellig ◽  
Roger Lehmann ◽  
Federica Cuozzo ◽  
Daniela Nasteska ◽  
...  
Keyword(s):  
Cell Death ◽  
Islet Cell ◽  
Islet Cells ◽  
Zinc Content ◽  
Zinc Homeostasis ◽  
Wild Type ◽  
Zinc Chelator ◽  
Zinc Concentrations ◽  
Processing And Storage

Pancreatic β-cells depend on the well-balanced regulation of cytosolic zinc concentrations, providing sufficient zinc ions for the processing and storage of insulin, but avoiding toxic effects. The zinc transporter ZnT8, encoded by SLC30A8, is a key player regarding islet cell zinc homeostasis, and polymorphisms in this gene are associated with altered type 2 diabetes susceptibility in man. The objective of this study was to investigate the role of ZnT8 and zinc in situations of cellular stress as hypoxia or inflammation. Isolated islets of wild-type and global ZnT8-/- mice were exposed to hypoxia or cytokines and cell death was measured. To explore the role of changing intracellular Zn2+ concentrations, wild-type islets were exposed to different zinc concentrations using zinc chloride or the zinc chelator N,N,N′,N′-tetrakis(2-pyridinylmethyl)-1,2-ethanediamine (TPEN). Hypoxia or cytokine (TNFα, IFNγ, IL1β) treatment induced islet cell death, but to a lesser extent in islets from ZnT8-/- mice, which were shown to have a reduced zinc content. Similarly, chelation of zinc with TPEN reduced cell death in wild-type islets treated with hypoxia or cytokines, whereas increased zinc concentrations aggravated the effects of these stressors. This study demonstrates a reduced rate of cell death in islets from ZnT8-/- mice as compared to wild-type islets when exposed to two distinct cellular stressors, hypoxia or cytotoxic cytokines. This protection from cell death is, in part, mediated by a reduced zinc content in islet cells of ZnT8-/- mice. These findings may be relevant for altered diabetes burden in carriers of risk SLC30A8 alleles in man.

scholarly journals Fast and easy single-molecule pulldown assay based on agarose microbeads

2020 ◽  
Author(s):  
Qirui Zhao ◽  
Yusheng Shen ◽  
Xiaofen Li ◽  
Fang Tian ◽  
Xiaojie Yu ◽  
...  
Keyword(s):  
Single Molecule ◽  
Signal To Noise Ratio ◽  
Original Method ◽  
Auditory Hair Cells ◽  
Single Molecule Level ◽  
Cell Extracts ◽  
Binding Partners ◽  
Pulldown Assay ◽  
Ultrahigh Sensitivity ◽  
First Time

SUMMARYThe recently developed single-molecule pulldown (SiMPull) assay by Jain and colleagues is a highly innovative technique but its wide application is hindered by the high technical barrier and time consumption. We report an innovative, agarose microbead-based approach for SiMPull. We used commercially available, pre-surface-functionalized agarose microbeads to capture the protein of interest together with its binding partners specifically from cell extracts and observed these interactions under a microscope at the single-molecule level. Relative to the original method, microbead-based SiMPull is considerably faster, easier to use, and more reproducible and yet provides similar sensitivity and signal-to-noise ratio; specifically, with the new method, sample-preparation time is substantially decreased (from ∼10 to ∼3 h). These crucial features should facilitate wide application of powerful and versatile SiMPull in common biological and clinical laboratories. Notably, by exploiting the simplicity and ultrahigh sensitivity of microbead-based SiMPull, we used this method in the study of rare auditory hair cells for the first time.

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