scholarly journals Mg2+ modulates the activity of hyperpolarization-activated calcium currents in plant cells

2020 ◽  
Author(s):  
Fouad Lemtiri-Chlieh ◽  
Stefan T. Arold ◽  
Chris Gehring
Keyword(s):  
Plant Hormone ◽  
Plant Cells ◽  
Guard Cells ◽  
Cell Types ◽  
Calcium Currents ◽  
Binding Motif ◽  
Animal Cells ◽  
Membrane Hyperpolarization ◽  
Cyclic Nucleotide Gated Channels ◽  
Channel Opening

ABSTRACTHyperpolarization-activated calcium channels (HACCs) are found in the plasma membrane and tonoplast of many plant cell types where they have an important role in Ca2+-dependent signaling. The unusual gating properties of HACCs in plants, i.e., activation by membrane hyperpolarization rather than depolarization, dictates that HACCs are normally open at physiological hyperpolarized resting membrane potentials (the so called pump or P-state), thus, if not regulated, they would be continuously leaking Ca2+ into cells. In guard cells, HACCs are permeable to Ca2+, Ba2+ and Mg2+, activated by H2O2 and the plant hormone abscisic acid (ABA) and their activity is greatly reduced by low amounts of free cytosolic Ca2+ ([Ca2+]Cyt) and hence will close during [Ca2+]Cyt surges. Here we demonstrate that the presence of the commonly used Mg-ATP inside the cell greatly reduces HACC activity especially at voltages ≤ −200 mV and that Mg2+ causes this block. We therefore conclude, firstly, that physiological cytosolic Mg2+ levels affect HACCs gating and that channel opening requires either high negative voltages (≥ −200 mV) and/or displacement of Mg2+ away from the immediate vicinity of the channel. Secondly, based on structural comparisons with Mg2+-sensitive animal inward-rectifying K+ channel, we propose that the likely candidate HACCS described here are cyclic nucleotide gated channels (CNGCs), many of which also contain a conserved di-acidic Mg2+-binding motif within their pores. This conclusion is consistent with the electrophysiological data. Finally, we propose that Mg2+, much like in animal cells, is an important component in Ca2+ signalling and homeostasis in plants.

scholarly journals Mg2+ is a Missing Link in Plant Cell Ca2+ Signalling and Homeostasis—A Study on Vicia faba Guard Cells

2020 ◽  
Vol 21 (11) ◽  
pp. 3771
Author(s):  
Fouad Lemtiri-Chlieh ◽  
Stefan T. Arold ◽  
Chris Gehring
Keyword(s):  
Plant Cell ◽  
Guard Cells ◽  
Cell Types ◽  
K Channel ◽  
Resting Membrane Potential ◽  
Binding Motif ◽  
Animal Cells ◽  
Membrane Hyperpolarization ◽  
Cyclic Nucleotide Gated Channels ◽  
Channel Opening

Hyperpolarization-activated calcium channels (HACCs) are found in the plasma membrane and tonoplast of many plant cell types, where they have an important role in Ca2+-dependent signalling. The unusual gating properties of HACCs in plants, i.e., activation by membrane hyperpolarization rather than depolarization, dictates that HACCs are normally open in the physiological hyperpolarized resting membrane potential state (the so-called pump or P-state); thus, if not regulated, they would continuously leak Ca2+ into cells. HACCs are permeable to Ca2+, Ba2+, and Mg2+; activated by H2O2 and the plant hormone abscisic acid (ABA); and their activity in guard cells is greatly reduced by increasing amounts of free cytosolic Ca2+ ([Ca2+]Cyt), and hence closes during [Ca2+]Cyt surges. Here, we demonstrate that the presence of the commonly used Mg-ATP inside the guard cell greatly reduces HACC activity, especially at voltages ≤ −200 mV, and that Mg2+ causes this block. Therefore, we firstly conclude that physiological cytosolic Mg2+ levels affect HACC gating and that channel opening requires either high negative voltages (≥−200 mV) or displacement of Mg2+ away from the immediate vicinity of the channel. Secondly, based on structural comparisons with a Mg2+-sensitive animal inward-rectifying K+ channel, we propose that the likely candidate HACCs described here are cyclic nucleotide gated channels (CNGCs), many of which also contain a conserved diacidic Mg2+ binding motif within their pores. This conclusion is consistent with the electrophysiological data. Finally, we propose that Mg2+, much like in animal cells, is an important component in Ca2+ signalling and homeostasis in plants.

scholarly journals Arabidopsis NPF4.6 and NPF5.1 Control Leaf Stomatal Aperture by Regulating Abscisic Acid Transport

Genes ◽  
2021 ◽  
Vol 12 (6) ◽  
pp. 885
Author(s):  
Takafumi Shimizu ◽  
Yuri Kanno ◽  
Hiromi Suzuki ◽  
Shunsuke Watanabe ◽  
Mitsunori Seo
Keyword(s):  
Abscisic Acid ◽  
Plant Hormone ◽  
Leaf Surface ◽  
Stomatal Closure ◽  
Guard Cells ◽  
Cell Types ◽  
Acid Transport ◽  
Wild Type ◽  
Vascular Tissues ◽  
Abscisic Acid Transport

The plant hormone abscisic acid (ABA) is actively synthesized in vascular tissues and transported to guard cells to promote stomatal closure. Although several transmembrane ABA transporters have been identified, how the movement of ABA within plants is regulated is not fully understood. In this study, we determined that Arabidopsis NPF4.6, previously identified as an ABA transporter expressed in vascular tissues, is also present in guard cells and positively regulates stomatal closure in leaves. We also found that mutants defective in NPF5.1 had a higher leaf surface temperature compared to the wild type. Additionally, NPF5.1 mediated cellular ABA uptake when expressed in a heterologous yeast system. Promoter activities of NPF5.1 were detected in several leaf cell types. Taken together, these observations indicate that NPF5.1 negatively regulates stomatal closure by regulating the amount of ABA that can be transported from vascular tissues to guard cells.

Microtubules in plant cell division

1989 ◽  
Vol 47 ◽  
pp. 758-759
Author(s):  
S. M. Wick
Keyword(s):  
Electron Microscopy ◽  
Plant Cell ◽  
Mother Cell ◽  
Immunofluorescence Microscopy ◽  
Plant Cells ◽  
Preprophase Band ◽  
Guard Cells ◽  
Cell Types ◽  
Cell Cortex ◽  
Stomatal Complexes

Immunofluorescence microscopy has proven to be a valuable accompaniment to electron microscopy for study of the cytoskeleton of plant cells. Whereas electron microscopy provides greater resolution and details of the spatial relationships of the cytoskeleton to other cellular components, fluorescence visualization makes it possible to see the three-dimensional organization of cytoskeletal elements without laborious reconstruction of views from serial sections. An area in which immunofluorescence microscopy has been useful is the investigation of how plant cells organize and position the various microtubule arrays that are utilized during mitosis, cytokinesis and cell expansion phases. One of the earliest indications of an impending division in a meristematic plant cell is the formation of a preprophase band of microtubules in the cell cortex, at the site where the new wall will be placed at the subsequent cytokinesis. At its later stages, the band is narrower than when first identifiable. In most cells, preprophase band microtubules have the same general orientation as the preceding interphase microtubules, and so preprophase band formation here could, in theory, be achieved by lateral bundling of microtubules.Cells in which the division site and the preprophase band that marks it are not oriented parallel to interphase microtubules are found in stomatal complexes of grass leaves . Fig. 1 illustrates the arrangement of two such cell types: the guard mother cell, which divides lengthwise to form two guard cells, side-by-side, and the subsidiary mother cell, which undergoes a very asymmetric division to produce one of the pair of lens-shaped subsidiary cells that flank the guard cells. Interphase and preprophase arrangements of microtubules for each cell type are diagrammed in Figs. 2-4. In order to examine how these cell types achieve the reorientation of microtubules that is necessary to progress from interphase to preprophase, sheets of epidermis containing actively dividing stomatal complex cells were examined with immunofluorescence microscopy using antibodies to tubulin. Thin epidermal slices of leaves were fixed and glued down to a slide, whereupon cell walls were enzymatically weakened so that unwanted cell layers could be removed . Because waves of division pass along grass leaves, cells of the same type in a given file tend to be at similar stages, which facilitates deduction of the developmental pattern.

Ultrastructural distribution of a MAP kinase and transcripts in quiescent and cycling plant cells and pollen grains

1999 ◽  
Vol 112 (7) ◽  
pp. 1065-1076
Author(s):  
G. Prestamo ◽  
P.S. Testillano ◽  
O. Vicente ◽  
P. Gonzalez-Melendi ◽  
M.J. Coronado ◽  
...  
Keyword(s):  
Catalytic Domain ◽  
Polyclonal Antibodies ◽  
Plant Cells ◽  
Pollen Grains ◽  
Cell Types ◽  
Protein Distribution ◽  
Animal Cells ◽  
Proliferating Cells ◽  
Quiescent Cells

Mitogen-activated protein kinases (MAPKs) are components of a kinase module that plays a central role in the transduction of diverse extracellular stimuli, including mitogens, specific differentiation and developmental signals and stress treatments. This shows that reversible protein phosphorylation cascades play a pivotal role in signal transduction in animal cells and yeast, particularly the entry into mitosis of arrested cells. Homologues of MAPKs have been found and cloned in various plant species, but there have been no data about their in situ localization at the subcellular level and their expression in plant cells so far. In the present paper we report the first data on the ultrastructural in situ localization of MAPK and their mRNAs in various plant cells. Proliferating and quiescent meristematic plant cells were studied to evaluate whether changes in MAPK presence, distribution and expression accompany the entry into proliferation of dormant cells. Moreover, MAPK localization was analyzed in vacuolate microspores. Polyclonal antibodies against the deduced MAPK from the tobacco Ntf6 clone were able to recognize homologue epitopes by immunocytochemical techniques in the cell types studied. The pattern of protein distribution is similar in all the cases studied: it is localized in the cytoplasm and in the nucleus, mainly in the interchromatin region. The quantitative study of the density showed that MAPK labelling is more abundant in cycling than in quiescent cells, also suggesting that, in plants, MAPK pathways might play a role in cell proliferation. RNA probes for conserved regions of the catalytic domain of plant MAPK homologue genes were used to study MAPK expression in those plant cells. In situ hybridization (ISH) showed the presence of MAPK transcripts in the three plant cell types studied, but levels were very low in quiescent cells compared to those in cycling cells. The quantification of labelling density of ISH signals strongly suggests a higher level of MAPK expression in proliferating cells, but also some basal messenger presence and/or expression in the quiescent ones. Immunogold and ISH results show the presence and distribution of MAPK proteins and mRNAs in vacuolate microspores. This represents a very dynamic stage during pollen development in which the cell nucleus is being prepared for an asymmetrical mitotic division, giving rise to both the generative and the vegetative nuclei of the bicellular pollen grain. Taken together, the data indicate a role played by MAPK in the re-entry into proliferation in plant cells.

scholarly journals Guard cells control hypocotyl elongation through HXK1, HY5, and PIF4

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Gilor Kelly ◽  
Danja Brandsma ◽  
Aiman Egbaria ◽  
Ofer Stein ◽  
Adi Doron-Faigenboim ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Blue Light ◽  
Opposite Effect ◽  
Red Light ◽  
Guard Cells ◽  
Cell Types ◽  
Hypocotyl Elongation ◽  
Sugar Production ◽  
Effect Of Light

AbstractThe hypocotyls of germinating seedlings elongate in a search for light to enable autotrophic sugar production. Upon exposure to light, photoreceptors that are activated by blue and red light halt elongation by preventing the degradation of the hypocotyl-elongation inhibitor HY5 and by inhibiting the activity of the elongation-promoting transcription factors PIFs. The question of how sugar affects hypocotyl elongation and which cell types stimulate and stop that elongation remains unresolved. We found that overexpression of a sugar sensor, Arabidopsis hexokinase 1 (HXK1), in guard cells promotes hypocotyl elongation under white and blue light through PIF4. Furthermore, expression of PIF4 in guard cells is sufficient to promote hypocotyl elongation in the light, while expression of HY5 in guard cells is sufficient to inhibit the elongation of the hy5 mutant and the elongation stimulated by HXK1. HY5 exits the guard cells and inhibits hypocotyl elongation, but is degraded in the dark. We also show that the inhibition of hypocotyl elongation by guard cells’ HY5 involves auto-activation of HY5 expression in other tissues. It appears that guard cells are capable of coordinating hypocotyl elongation and that sugar and HXK1 have the opposite effect of light on hypocotyl elongation, converging at PIF4.

Cholecystokinin-gated currents in neurons of the rat solitary complex in vitro

1993 ◽  
Vol 70 (6) ◽  
pp. 2584-2595 ◽  
Author(s):  
P. Branchereau ◽  
J. Champagnat ◽  
M. Denavit-Saubie
Keyword(s):  
Voltage Clamp ◽  
Potassium Current ◽  
Reversal Potential ◽  
Input Resistance ◽  
Calcium Currents ◽  
Equilibrium Potential ◽  
Potassium Ions ◽  
Nernst Equation ◽  
Membrane Hyperpolarization ◽  
Cckb Receptor

1. Ionic conductances controlled by type A and type B cholecystokinin (CCK) receptors were studied in neurons of the rat nucleus tractus solitarius (NTS) and dorsal motor nucleus of the vagus (DMNV), using intracellular and whole-cell patch clamp recordings in current or voltage clamp configuration during bath application of agonists (CCK8, CCK4, BC 264) and antagonists. 2. CCKA receptor-related inhibition was associated with a membrane hyperpolarization and a decrease in input resistance that developed 2-6 min after the arrival of drug into the extracellular medium. These effects were induced by 5 nM CCK8 but not BC 264 and they were blocked by the CCKA antagonist, L-364,718, but not by the CCKB antagonist, L-365,260. 3. CCKA receptor-related inhibition was generated by a potassium current that reversed at a reversal potential E(rev) of -73 +/- 1 (mean +/- SE) mV with bathing potassium concentration [K+]o = 6 mM and at -88 +/- 1 with [K+]o = 3 mM, in agreement with the Nernst equation for potassium ions. 4. CCKB receptor-related excitation was associated with a membrane depolarization and an increase of the input resistance induced by the following agonists at threshold concentrations: CCK8 (0.2 nM) > or = BC 264 (0.4 nM) > CCK4 (10.9 nM). The increase of input resistance was abolished by L-365,260 and was maintained after blockade of the CCKA current by L-364,718. 5. CCKB receptor-related excitation, in the neurons (30% of cases) in which clear response reversal was observed, appeared to be generated by a decrease of a potassium conductance. Responses showed a reversal potential E(rev) of -68 +/- 4 mV with [K+]o = 6 mM and -89 +/- 1 mV with [K+]o = 3 mM, verifying predictions from the Nernst equation applied to potassium ions. However, in 70% of cases, clear reversal was not observed at membrane potentials negative to the theoretical potassium equilibrium potential EK. 6. In voltage clamp studies, CCK8 induced a 181 +/- 17 pA inward current associated with a 26 +/- 4% decrease in the instantaneous current (I(ins)) generated by hyperpolarizing voltage steps. This effect on I(ins) was demonstrated in the absence of effects on the outward noninactivating potassium current (IM) and on the inward noninactivating cationic current (IQ). 7. CCKB receptor-mediated excitation was not suppressed by cobalt, a blocker of calcium currents, and was not associated with a change of the calcium-dependent potassium current (IK(Ca)).(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Role of glutathione on cell adhesion and volume.

2021 ◽  
Author(s):  
Alain Geloen ◽  
Emmanuelle Danty
Keyword(s):  
Cell Adhesion ◽  
Cell Volume ◽  
Reduced Glutathione ◽  
Self Regulation ◽  
Cell Types ◽  
Self Control ◽  
Animal Cells ◽  
Cellular Processes ◽  
Proliferation And Apoptosis ◽  
Glutamylcysteine Synthetase

Glutathione is the most abundant thiol in animal cells. Reduced glutathione (GSH) is a major intracellular antioxidant neutralizing free radicals and detoxifying electrophiles. It plays important roles in many cellular processes, including cell differentiation, proliferation, and apoptosis. In the present study we demonstrate that extracellular concentration of reduced glutathione markedly increases cell volume within few hours, in a dose-response manner. Pre-incubation of cells with BSO, the inhibitor of 7-glutamylcysteine synthetase, responsible for the first step in intracellular glutathione synthesis did not change the effect of reduced glutathione on cell volume suggesting a mechanism limited to the interaction of extracellular reduced glutathione on cell membrane. Results show that reduced GSH decreases cell adhesion resulting in an increased cell volume. Since many cell types are able to transport of GSH out, the present results suggest that this could be a fundamental self-regulation of cell volume, giving the cells a self-control on their adhesion proteins.

scholarly journals The role of Aspergillus nidulans polo-like kinase PlkA in microtubule-organizing center control

2021 ◽  
Author(s):  
Xiaolei Gao ◽  
Saturnino Herrero ◽  
Valentin Wernet ◽  
Sylvia Erhardt ◽  
Oliver Valerius ◽  
...  
Keyword(s):  
Aspergillus Nidulans ◽  
Cell Types ◽  
Spindle Pole ◽  
Receptor Protein ◽  
Animal Cells ◽  
Microtubule Organizing Center ◽  
Spindle Pole Bodies ◽  
Organizing Center ◽  
Ring Complex ◽  
Core Components

Centrosomes are important microtubule-organizing centers (MTOC) in animal cells. In addition, non-centrosomal MTOCs (ncMTOCs) were described in many cell types. Functional analogs of centrosomes in fungi are the spindle pole bodies (SPBs). In Aspergillus nidulans additional MTOCs were discovered at septa (sMTOC). Although the core components are conserved in both MTOCs, their composition and organization are different and dynamic. Here, we show that the polo-like kinase PlkA binds the γ-tubulin ring complex (γ-TuRC) receptor protein ApsB and contributes to targeting ApsB to both MTOCs. PlkA coordinates SPB outer plaque with sMTOC activities. PlkA kinase activity was required for astral MT formation involving ApsB recruitment. PlkA also interacted with the γ-TuRC inner plaque receptor protein PcpA. Mitosis was delayed without PlkA, and the PlkA protein was required for proper mitotic spindle morphology, although this function was independent of its catalytic activity. Our results suggest polo-like kinase as a regulator of MTOC activities and as a scaffolding unit through interaction with γ-tubulin ring complex receptors.

scholarly journals Microtubule-based Endoplasmic Reticulum Motility in Xenopus laevis: Activation of Membrane-associated Kinesin during Development

1999 ◽  
Vol 10 (6) ◽  
pp. 1909-1922 ◽  
Author(s):  
Jon D. Lane ◽  
Victoria J. Allan
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Types ◽  
Cytoplasmic Dynein ◽  
Culture Cell ◽  
Tissue Culture Cell ◽  
Differential Interference Contrast Microscopy ◽  
Animal Cells ◽  
Microtubule Motor ◽  
Directed Motility ◽  
Conventional Kinesin

The endoplasmic reticulum (ER) in animal cells uses microtubule motor proteins to adopt and maintain its extended, reticular organization. Although the orientation of microtubules in many somatic cell types predicts that the ER should move toward microtubule plus ends, motor-dependent ER motility reconstituted in extracts ofXenopus laevis eggs is exclusively a minus end-directed, cytoplasmic dynein-driven process. We have used Xenopusegg, embryo, and somatic Xenopus tissue culture cell (XTC) extracts to study ER motility during embryonic development inXenopus by video-enhanced differential interference contrast microscopy. Our results demonstrate that cytoplasmic dynein is the sole motor for microtubule-based ER motility throughout the early stages of development (up to at least the fifth embryonic interphase). When egg-derived ER membranes were incubated in somatic XTC cytosol, however, ER tubules moved in both directions along microtubules. Data from directionality assays suggest that plus end-directed ER tubule extensions contribute ∼19% of the total microtubule-based ER motility under these conditions. In XTC extracts, the rate of ER tubule extensions toward microtubule plus ends is lower (∼0.4 μm/s) than minus end-directed motility (∼1.3 μm/s), and plus end-directed motility is eliminated by a function-blocking anti-conventional kinesin heavy chain antibody (SUK4). In addition, we provide evidence that the initiation of plus end-directed ER motility in somatic cytosol is likely to occur via activation of membrane-associated kinesin.

scholarly journals Engineering of Adenovirus Vectors Containing Heterologous Peptide Sequences in the C Terminus of Capsid Protein IX

2002 ◽  
Vol 76 (14) ◽  
pp. 6893-6899 ◽  
Author(s):  
Igor P. Dmitriev ◽  
Elena A. Kashentseva ◽  
David T. Curiel
Keyword(s):  
Heparan Sulfate ◽  
Capsid Protein ◽  
Minor Component ◽  
Cell Types ◽  
Binding Motif ◽  
Flag Epitope ◽  
C Terminus ◽  
Protein Ix ◽  
A Minor ◽  
Carboxy Terminal

ABSTRACT The utility of the present generation of adenovirus (Ad) vectors for gene therapy applications could be improved by restricting native viral tropism to selected cell types. In order to achieve modification of Ad tropism, we proposed to exploit a minor component of viral capsid, protein IX (pIX), for genetic incorporation of targeting ligands. Based on the proposed structure of pIX, we hypothesized that its C terminus could be used as a site for incorporation of heterologous peptide sequences. We engineered recombinant Ad vectors containing modified pIX carrying a carboxy-terminal Flag epitope along with a heparan sulfate binding motif consisting of either eight consecutive lysines or a polylysine sequence. Using an anti-Flag antibody, we have shown that modified pIXs are incorporated into virions and display Flag-containing C-terminal sequences on the capsid surface. In addition, both lysine octapeptide and polylysine ligands were accessible for binding to heparin-coated beads. In contrast to virus bearing lysine octapeptide, Ad vector displaying a polylysine was capable of recognizing cellular heparan sulfate receptors. We have demonstrated that incorporation of a polylysine motif into the pIX ectodomain results in a significant augmentation of Ad fiber knob-independent infection of CAR-deficient cell types. Our data suggest that the pIX ectodomain can serve as an alternative to the fiber knob, penton base, and hexon proteins for incorporation of targeting ligands for the purpose of Ad tropism modification.

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