Transverse Microtubules Research Articles

Suppression of Cortical Microtubule Reorientation and Stimulation of Cell Elongation in Arabidopsis Hypocotyls under Microgravity Conditions in Space.

How microgravity in space influences plant cell growth is an important issue for plant cell biology as well as space biology. We investigated the role of cortical microtubules in the stimulation of elongation growth in Arabidopsis (Arabidopsis thaliana) hypocotyls under microgravity conditions with the Resist Tubule space experiment. The epidermal cells in the lower half of the hypocotyls of wild-type Columbia were longer in microgravity than at on-orbit 1 g, which precipitated an increase in the entire hypocotyl length. In the apical region, cortical microtubules adjacent to the outer tangential wall were predominantly transverse to the long axis of the cell, whereas longitudinal microtubules were predominant in the basal region. In the 9th to 12th epidermal cells (1 to 3 mm) from the tip, where the modification of microtubule orientation from transverse to longitudinal directions (reorientation) occurred, cells with transverse microtubules increased, whereas those with longitudinal microtubules decreased in microgravity, and the average angle with respect to the transverse cell axis decreased, indicating that the reorientation was suppressed in microgravity. The expression of tubulin genes was suppressed in microgravity. These results suggest that under microgravity conditions, the expression of genes related to microtubule formation was downregulated, which may cause the suppression of microtubule reorientation from transverse to longitudinal directions, thereby stimulating cell elongation in Arabidopsis hypocotyls.

Microgravity modelling by two-axial clinorotation leads to scattered organisation of cytoskeleton in Arabidopsis seedlings.

Proper plant development in a closed ecosystem under weightlessness will be crucial for the success of future space missions. To supplement spaceflight experiments, such conditions of microgravity are modelled on Earth using a two-axial (2A) clinorotation, and in several fundamental studies resulted in the data on proteome and metabolome adjustments, embryo development, cell cycle regulation, etc. Nevertheless, our understanding of the cytoskeleton responses to the microgravity is still limited. In the present work, we study the adjustment of actin microfilaments (MFs) and microtubules (MTs) in Arabidopsis thaliana (L.) Heynh. seedlings under 2A clinorotation. Modelled microgravity resulted in not only the alteration of seedlings phenotype, but also a transient increase of the hydrogen peroxide level and in the cytoskeleton adjustment. Using GFP-fABD2 and Lifeact-Venus transgenic lines, we demonstrate that MFs became 'scattered' in elongating root and hypocotyl cells under 2A clinorotation. In addition, in GFP-MAP4 and GFP-TUA6 lines the tubulin cytoskeleton had higher fractions of transverse MTs under 2A clinorotation. Remarkably, the first static gravistimulation of continuously clinorotated seedlings reverted MF organisation to a longitudinal one in roots within 30 min. Our data suggest that the 'scattered' organisation of MFs in microgravity can serve as a good basis for the rapid cytoskeleton conversion to a 'longitudinal' structure under the gravity force.

Rapid, redox-mediated mechanical susceptibility of the cortical microtubule lattice in skeletal muscle

The highly ordered cortical microtubule lattice of skeletal muscle is disorganized in dystrophin-deficient mdx mice. Implicated mechanisms include loss of dystrophin binding, altered α-tubulin posttranslational modification, expression of a β-tubulin involved in regeneration, and reactive oxygen species (ROS). Here we show that the transverse microtubules in mdx muscle expressing miniaturized dystrophins are rapidly lost after eccentric contraction. Analysis of mdx lines expressing different dystrophin constructs demonstrate that spectrin-like repeats R4-15 and R20-23 were required for mechanically stable microtubules. Microtubule loss was prevented by the non-specific antioxidant N-acetylcysteine while inhibition of NADPH oxidase 2 had only a partial effect, suggesting that ROS from multiple sources mediate the rapid loss of transverse microtubules after eccentric contraction. Finally, ablation of α-dystrobrevin, β- or γ-cytoplasmic actin phenocopied the transverse microtubule instability of miniaturized dystrophins. Our data demonstrate that multiple dystrophin domains, α-dystrobrevin and cytoplasmic actins are necessary for mechanically stable microtubules.

Maize ZmRPH1 encodes a microtubule-associated protein that controls plant and ear height.

Maize ZmRPH1 encodes a microtubule-associated protein that controls plant and ear height.

Exogenous Auxin Induces Transverse Microtubule Arrays Through TRANSPORT INHIBITOR RESPONSE1/AUXIN SIGNALING F-BOX Receptors.

Auxin plays a central role in controlling plant cell growth and morphogenesis. Application of auxin to light-grown seedlings elicits both axial growth and transverse patterning of the cortical microtubule cytoskeleton in hypocotyl cells. Microtubules respond to exogenous auxin within 5 min, although repatterning of the array does not initiate until 30 min after application and is complete by 2 h. To examine the requirements for auxin-induced microtubule array patterning, we used an Arabidopsis (Arabidopsis thaliana) double auxin f-box (afb) receptor mutant, afb4-8 afb5-5, that responds to conventional auxin (indole-3-acetic acid) but has a strongly diminished response to the auxin analog, picloram. We show that 5 µm picloram induces immediate changes to microtubule density and later transverse microtubule patterning in wild-type plants, but does not cause microtubule array reorganization in the afb4-8 afb5-5 mutant. Additionally, a dominant mutant (axr2-1) for the auxin coreceptor AUXIN RESPONSIVE2 (AXR2) was strongly suppressed for auxin-induced microtubule array reorganization, providing additional evidence that auxin functions through a transcriptional pathway for transverse patterning. We observed that brassinosteroid application mimicked the auxin response, showing both early and late microtubule array effects, and induced transverse patterning in the axr2-1 mutant. Application of auxin to the brassinosteroid synthesis mutant, diminuto1, induced transverse array patterning but did not produce significant axial growth. Thus, exogenous auxin induces transverse microtubule patterning through the TRANSPORT INHIBITOR 1/AUXIN F-BOX (TIR1/AFB) transcriptional pathway and can act independently of brassinosteroids.

The Effect of Naphthazarin on the Growth, Electrogenicity, Oxidative Stress, and Microtubule Array in Z. mays Coleoptile Cells Treated With IAA.

Naphthazarin (5,8-dihydroxy-1,4-naphthoquinone, DHNQ) is a naturally occurring 1,4-naphthoquinone derivative. In this study, we focused on elucidating the toxic effect of this secondary metabolite on the growth of plant cells. The dose–response curves that were obtained for the effects of DHNQ on endogenous and IAA-induced growth in maize coleoptile segments differ in shape; in the first case, it is linear, while in the presence of auxin it is bell-shaped with the maximum at 1 μM. It was found that DHNQ at almost all concentrations studied, when added to the incubation medium inhibited endogenous growth (excluding naphthazarin at 0.001 μM) as well as growth in the presence of IAA. Simultaneous measurements of the growth and external medium pH of coleoptile segments indicated that DHNQ diminished or eliminated proton extrusion at all of the concentrations that were used. Interestingly, the oxidative stress in maize coleoptile cells, which was measured as hydrogen peroxide (H2O2) production, catalase activity, redox activity and malondialdehyde (MDA) content, increased at the lower concentrations of DHNQ (<1 μM), thus suggesting a specific character of its action. It was also found that naphthazarin at concentration higher than 0.1 μM caused the depolarization of the membrane potential (Em). An analysis of the organization and anisotropy of the cortical microtubules showed that naphthazarin at all of the concentrations that were studied changed the IAA-induced transverse microtubule reorientation to an oblique reorientation. Our results indicate that naphthazarin diminished the growth of maize coleoptile cells by a broad spectrum of its toxic effects, thereby suggesting that naphthazarin might be a hypothetical component of new bioherbicides and biopesticides.

A Cycloheximide-Sensitive Step in Transverse Microtubule Array Patterning.

The growth properties of individual cells within a tissue determine plant morphology, and the organization of the cytoskeleton, particularly the microtubule arrays, determines cellular growth properties. We investigated the mechanisms governing the formation of transverse microtubule array patterns in axially growing Arabidopsis (Arabidopsis thaliana) epidermal hypocotyl cells. Using quantitative imaging approaches, we mapped the transition of the cortical microtubule arrays into a transverse coaligned pattern after induction with auxin and gibberellic acid. Hormone induction led to an early loss of microtubule plus end density and a rotation toward oblique patterns. Beginning 30 min after induction, transverse microtubules appeared at the cell's midzone concurrently with the loss of longitudinal polymers, eventually progressing apically and basally to remodel the array pattern. Based on the timing and known hormone-signaling pathways, we tested the hypothesis that the later events require de novo gene expression and, thus, constitute a level of genetic control over transverse patterning. We found that the presence of the translation inhibitor cycloheximide (CHX) resulted in a selective and reversible loss of transverse patterns that were replaced with radial-like pinwheel arrays exhibiting a split bipolar architecture centered at the cell's midzone. Experiments using hormone induction and CHX revealed that pinwheel arrays occur when transverse microtubules increase at the midzone but longitudinal microtubules in the split bipolar architecture are not suppressed. We propose that a key regulatory mechanism for creating the transverse microtubule coalignment in axially growing hypocotyls involves the expression of a CHX-sensitive factor that acts to suppress the nucleation of the longitudinally oriented polymers.

Cortical microtubule orientation in Arabidopsis thaliana root meristematic zone depends on cell division and requires severing by katanin

BackgroundTransverse cortical microtubule orientation, critical for anisotropic cell expansion, is established in the meristematic root zone. Intending to elucidate the possible prerequisites for this establishment and factors that are involved, microtubule organization was studied in roots of Arabidopsis thaliana, wild-type and the p60-katanin mutants fra2, ktn1-2 and lue1. Transverse cortical microtubule orientation in the meristematic root zone has proven to persist under several regimes inhibiting root elongation. This persistence was attributed to the constant moderate elongation of meristematic cells, prior to mitotic division. Therefore, A. thaliana wild-type seedlings were treated with aphidicolin, in order to prevent mitosis and inhibit premitotic cell elongation.ResultsIn roots treated with aphidicolin for 12 h, cell divisions still occurred and microtubules were transverse. After 24 and 48 h of treatment, meristematic cell divisions and the prerequisite elongation ceased, while microtubule orientation became random. In meristematic cells of the p60-katanin mutants, apart from a general transverse microtubule pattern, cortical microtubules with random orientation were observed, also converging at several cortical sites, in contrast to the uniform transverse pattern of wild-type cells.ConclusionTaken together, these observations reveal that transverse cortical microtubule orientation in the meristematic zone of A. thaliana root is cell division-dependent and requires severing by katanin.

Growth and cortical microtubule dynamics in shoot organs under microgravity and hypergravity conditions

ABSTRACTThe body shape of plants varied in proportion to the logarithm of the magnitude of gravity in the range from microgravity to hypergravity to resist the gravitational force. Here we discuss the roles of cortical microtubule and 65 kDa microtubule-associated protein-1 (MAP65-1) in gravity-induced modification of growth anisotropy. Microgravity stimulated elongation growth and suppressed lateral expansion in shoot organs, such as hypocotyls and epicotyls. On the other hand, hypergravity inhibited elongation growth and promoted lateral expansion in shoot organs. The number of cells with transverse microtubules was increased by microgravity, but decreased by hypergravity. Furthermore, the levels of MAP65-1, which is involved in the maintenance of the transverse microtubule orientation, were increased by microgravity, but decreased by hypergravity. Therefore, the regulation of orientation of cortical microtubules via changes in the levels of MAP65-1 may contribute to the modification of the body shape of plants to resist the gravitational force.

Modification of growth anisotropy and cortical microtubule dynamics in Arabidopsis hypocotyls grown under microgravity conditions in space.

We carried out a space experiment, denoted as Aniso Tubule, to examine the effects of microgravity on the growth anisotropy and cortical microtubule dynamics in Arabidopsis hypocotyls, using lines in which microtubules are visualized by labeling tubulin or microtubule-associated proteins (MAPs) with green fluorescent protein (GFP). In all lines, GFP-tubulin6 (TUB6)-, basic proline-rich protein1 (BPP1)-GFP- and spira1-like3 (SP1L3)-GFP-expressing using a constitutive promoter, and spiral2 (SPR2)-GFP- and GFP-65 kDa MAP-1 (MAP65-1)-expressing using a native promoter, the length of hypocotyls grown under microgravity conditions in space was longer than that grown at 1 g conditions on the ground. In contrast, the diameter of hypocotyls grown under microgravity conditions was smaller than that of the hypocotyls grown at 1 g. The percentage of cells with transverse microtubules was increased under microgravity conditions, irrespective of the lines. Also, the average angle of the microtubules with respect to the transverse cell axis was decreased in hypocotyls grown under microgravity conditions. When GFP fluorescence was quantified in hypocotyls of GFP-MAP65-1 and SPR2-GFP lines, microgravity increased the levels of MAP65-1, which appears to be involved in the maintenance of transverse microtubule orientation. However, the levels of SPR2 under microgravity conditions were comparable to those at 1 g. These results suggest that the microgravity-induced increase in the levels of MAP65-1 is involved in increase in the transverse microtubules, which may lead to modification of growth anisotropy, thereby developing longer and thinner hypocotyls under microgravity conditions in space.

Fine Structure and Molecular Phylogeny of Parametopidium circumlabens (Ciliophora: Armophorea), Endocommensal of Sea Urchins.

Metopid armophoreans are ciliates commonly found in anaerobic environments worldwide; however, very little is known of their fine structure. In this study, the metopid Parametopidium circumlabens (Biggar and Wenrich 1932) Aescht, 1980, a common endocommensal of sea urchins, is investigated for the first time with emphasis on transmission electron microscopy, revealing several previously unknown elements of its morphology. Somatic dikinetids of P. circumlabens have a typical ribbon of transverse microtubules, an isolated microtubule near triplets 4 and 5 of the anterior kinetosome, plus two other microtubules between anterior and posterior kinetosomes, a short kinetodesmal striated fiber and long postciliary microtubules. In the dikinetids of the perizonal stripe, the kinetodesmal fiber is very pronounced, and there is a conspicuous microfibrillar network system associated with the kinetosomes. A new structure, shaped as a dense, roughly cylindrical mass surrounded by microtubules, is found associated with the posterior kinetosome of perizonal dikinetids. The paroral membrane is diplostichomonad and the adoral membranelles are of the "paramembranelle" type. Bayesian inference and maximum-likelihood analysis of the 18S-rDNA gene unambiguously placed P. circumlabens as sister group of the cluster formed by ((Atopospira galeata, Atopospira violacea) Metopus laminarius) + Clevelandellida, corroborating its classification within the Metopida.

Purification and Characterization of Novel Microtubule-Associated Proteins from Arabidopsis Cell Suspension Cultures

Plant microtubules (MTs) play essential roles in cell division, anisotropic cell expansion, and overall organ morphology. Microtubule-associated proteins (MAPs) bind to MTs and regulate their dynamics, stability, and organization. Identifying the full set of MAPs in plants would greatly enhance our understanding of how diverse MT arrays are formed and function; however, few proteomics studies have characterized plant MAPs. Using liquid chromatography-tandem mass spectrometry, we identified hundreds of proteins from MAP-enriched preparations derived from cell suspension cultures of Arabidopsis (Arabidopsis thaliana). Previously reported MAPs, MT regulators, kinesins, dynamins, peroxisome-resident enzymes, and proteins implicated in replication, transcription, and translation were highly enriched. Dozens of proteins of unknown function were identified, among which 12 were tagged with green fluorescent protein (GFP) and examined for their ability to colocalize with MTs when transiently expressed in plant cells. Six proteins did indeed colocalize with cortical MTs in planta. We further characterized one of these MAPs, designated as BASIC PROLINE-RICH PROTEIN1 (BPP1), which belongs to a seven-member family in Arabidopsis. BPP1-GFP decorated interphase and mitotic MT arrays in transgenic Arabidopsis plants. A highly basic, conserved region was responsible for the in vivo MT association. Overexpression of BPP1-GFP stabilized MTs, caused right-handed helical growth in rapidly elongating tissues, promoted the formation of transverse MT arrays, and resulted in the outgrowth of epidermal cells in light-grown hypocotyls. Our high-quality proteome database of Arabidopsis MAP-enriched preparations is a useful resource for identifying novel MT regulators and evaluating potential MT associations of proteins known to have other cellular functions.

ULTRASTRUCTURAL STUDY OF BALANTIDIUM POLYVACUOLUM LI, 1963 (CLASS: LITOSTOMATEA) THAT INHABITS IN XENOCYPRINAE FISH

Balantidium polyvacuolum Li,1963,an obligatory intestinal symbiont of Xenocyprinae fish,mainly inhabits in the luminal contents of the middle and hindgut.Though the description and some revisions are given at both light and scanning electronic microscopic levels,the morphological data are still incomplete since fine structural data with taxonomic values are still unavailable.This study examined the ultrastructural anatomy of this Chinese balantidium that inhabits in the intestines of Xenocypris davidi collected from the Jialing River in Sichuan province during fish parasite surveys in August 2011.All fish samples were dissected,and intestines were cut to collect the luminal contents into petridishes for further examination with the aid of a dissecting microscope.The luminal contents of the hindgut containing numerous B.polyvacuolum were selected and fixed directly in 2.5% glutaraldehyde in 0.2 M phosphate buffered saline(PBS,PH 7.4) for 2h at 4 ℃,then postfixed in 1%(v/v) osmium tetroxide in PBS for 2h at 4 ℃,this step was followed by a series of dehydration in acetone and embedded in Araldite.Ultrathin sections were cut on a Leica Ultracut R ultramicrotome,stained with uranyl acetate and lead citrate before being observed in a Philips Tecnai G2 20 Transmission Electron Microscope(Netherlands).Its somatic cortex,vestibulum,nuclei,cytoplasmic organelles and the centroventral concave platelet were well described here.Our results indicated that both somatic cortex and vestibulum were composed of pellicle(the outermost cell membranes,pellicular alveoli and the layer of microtubules) and infraciliature(kinetosomes and associated fibrils such as kinetodesmal fibril,postciliary microtubules,transverse microtubules and nematodesmata).However,the nematodesmata of vestibular kinetids were much longer than those of somatic kinetids and usually extended deeply into the endoplasm.As for the nuclei,the macronucleus was club-shaped with several nucleoli and heterochromatin occupying the karyoplasm,whereas the spherical micronucleus was homoeomerous with diffuse chromatin distributed over the karyoplasm.In addition,large amounts of amylopectin granules were observed within cytoplasm of the centroventral concave platelet.We suspected that this region may act as a storage pool of amylose to provide energy for the organism.Besides,the components of the cortex were compared among the related Balantidium species and their possible functions were also discussed.

Membrane Microdomains and Cytoskeleton Organization Shape and Regulate the IL-7 Receptor Signalosome in Human CD4 T-cells

Interleukin (IL)-7 is the main homeostatic regulator of CD4 T-lymphocytes (helper) at both central and peripheral levels. Upon activation by IL-7, several signaling pathways, mainly JAK/STAT, PI3K/Akt and MAPK, induce the expression of genes involved in T-cell differentiation, activation, and proliferation. We have analyzed the early events of CD4 T-cell activation by IL-7. We have shown that IL-7 in the first few min induces the formation of cholesterol-enriched membrane microdomains that compartmentalize its activated receptor and initiate its anchoring to the cytoskeleton, supporting the formation of the signaling complex, the signalosome, on the IL-7 receptor cytoplasmic domains. Here we describe by stimulated emission depletion microscopy the key roles played by membrane microdomains and cytoskeleton transient organization in the IL-7-regulated JAK/STAT signaling pathway. We image phospho-STAT5 and cytoskeleton components along IL-7 activation kinetics using appropriate inhibitors. We show that lipid raft inhibitors delay and reduce IL-7-induced JAK1 and JAK3 phosphorylation. Drug-induced disassembly of the cytoskeleton inhibits phospho-STAT5 formation, transport, and translocation into the nucleus that controls the transcription of genes involved in T-cell activation and proliferation. We fit together the results of these quantitative analyses and propose the following mechanism. Activated IL-7 receptors embedded in membrane microdomains induce actin-microfilament meshwork formation, anchoring microtubules that grow radially from rafted receptors to the nuclear membrane. STAT5 phosphorylated by signalosomes are loaded on kinesins and glide along the microtubules across the cytoplasm to reach the nucleus 2 min after IL-7 stimulation. Radial microtubules disappear 15 min later, while transversal microtubules, independent of phospho-STAT5 transport, begin to bud from the microtubule organization center.

Membrane Microdomains and Cytoskeleton Organization Shape and Regulate the IL7-Receptor Signalosome in Human CD4 T-Cells

Event Abstract Back to Event Membrane Microdomains and Cytoskeleton Organization Shape and Regulate the IL7-Receptor Signalosome in Human CD4 T-Cells BLANCHE TAMARIT1 and THIERRY ROSE1* 1 Institut Pasteur, Infection and Epidemiology/Immunology, France Interleukin(IL)-7 is the main homeostatic regulator of CD4 T-lymphocytes (helper) at both central and peripheral levels. Upon activation by IL-7, several signalling pathways, mainly Jak/STAT, PI3K/Akt and MAPK, induce the expression of genes involved in T-cell differentiation, activation and proliferation. We have analyzed the early events of CD4 T-cell activation by IL-7. We have shown that IL-7 in the first few minutes induces the formation of cholesterol-enriched membrane microdomains that compartmentalize its activated receptor and initiate its anchoring to the cytoskeleton supporting the formation of the signalosome on the IL-7-receptor cytoplasmic-domains [1]. We describe by stimulated emission depletion (STED) microscopy, the key roles played by membrane microdomains and cytoskeleton transient organization in the IL-7-regulated Jak/STAT signalling pathway [2]. We image phospho-STAT5 and cytoskeleton components along IL-7-activation kinetics using appropriate inhibitors. Lipid raft inhibitors delay and reduce IL-7-induced Jak1 and Jak3 phosphorylation. Drug-induced disassembly of cytoskeleton inhibits phospho-STAT5 formation, transport and translocation into the nucleus that controls the transcription of genes involved in T-cell activation and proliferation. We fit together the results of these quantitative analyses and propose the following mechanism sketched in the figure bellow with the corresponding STED images [2]: activated IL-7-receptors embedded in membrane microdomains induce actin-microfilament meshwork formation, anchoring microtubules that grow radially from rafted receptors to the nuclear membrane. STAT5 phosphorylated by signalosomes are loaded on kinesins and glide along the microtubules across the cytoplasm to reach the nucleus two minutes after IL-7-stimulation. Radial microtubules disappear 15 minutes later while transversal microtubules, independent of phospho-STAT5 transport, begin to bud from the microtubule-organization-center. Figure 1 References [1] T. Rose, A. H. Pillet, V. Lavergne, B. Tamarit, P. Lenormand, J.C. Rousselle, A. Namane, J. Thèze, Interleukin-7 compartmentalizes its receptor signaling complex to initiate CD4 T lymphocyte response, J Biol Chem, 285, 14898, 2010. [2] B. Tamarit, F. Bugault, A. H. Pillet, V. Lavergne, P. Bochet, N. Garin, U. Schwarz, J. Thèze, T. Rose, Membrane microdomains and cytoskeleton organization shape and regulate the IL7-receptor signalosome in human CD4 T-cells, J Biol Chem, 288 Keywords: lipid microdomain, Cytoskeleton, Interleukin-7, JAK/STAT signaling pathway, human CD4 T-cell, Kinesin Conference: 15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013. Presentation Type: Abstract Topic: Immune receptors and signaling Citation: TAMARIT B and ROSE T (2013). Membrane Microdomains and Cytoskeleton Organization Shape and Regulate the IL7-Receptor Signalosome in Human CD4 T-Cells. Front. Immunol. Conference Abstract: 15th International Congress of Immunology (ICI). doi: 10.3389/conf.fimmu.2013.02.00028 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 08 Mar 2013; Published Online: 22 Aug 2013. * Correspondence: Dr. THIERRY ROSE, Institut Pasteur, Infection and Epidemiology/Immunology, Paris, 75015, France, rose@pasteur.fr Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers BLANCHE TAMARIT THIERRY ROSE Google BLANCHE TAMARIT THIERRY ROSE Google Scholar BLANCHE TAMARIT THIERRY ROSE PubMed BLANCHE TAMARIT THIERRY ROSE Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Dissecting the mechanism of abscisic acid-induced dynamic microtubule reorientation using live cell imaging.

Abscisic acid (ABA) is involved in plant development and responses to environmental stress including the formation of longitudinal microtubule arrays in elongating cells, although the underlying mechanism for this is unknown. We explored ABA-induced microtubule reorientation in leek (Allium porrum L.) leaf epidermal cells transiently expressing a GFP-MBD microtubule reporter. After 14-18h incubation with ABA, the frequency of cells with longitudinal arrays of cortical microtubules along the outer epidermal wall increased with dose-dependency until saturation at 20μM. Time-course imaging of individual cells revealed a gradual increase in the occurrence of discordant, dynamic microtubules deviating from the normal transverse microtubule array within 2-4h of exposure to ABA, followed by reorientation into a completely longitudinal array within 5-8h. Approximately one-half of the ABA-induced reorientation occurred independently of cytoplasmic streaming following the application of cytochalasin D. Reorientation occurred also in the elongation zone of Arabidopsis root tips. Transient expression of AtEB1b-GFP reporter and analysis of 'comet' velocities in Allium revealed that the microtubule growth rate increased by 55% within 3h of exposure to ABA. ABA also increased the sensitivity of microtubules to depolymerisation by oryzalin and exacerbated oryzalin-induced radial swelling of Arabidopsis root tips. The swelling was further aggravated in AtPLDδ-null mutant, suggesting PLDδ plays a role in microtubule stability. We propose that ABA-induced reorientation of transverse microtubule array initially involves destabilisation of the array combined with the formation of dynamic, discordant microtubules.

The Ultrastructure of Placus striatus and a Revision of the Family Placidae (Ciliophora)

The first ultrastructural description of the genus Placus is presented. Each somatic kinetosome has a cone-shaped axosomal plate, nonoverlapping postciliary microtubules, an anteriorly directed kinetodesmal fiber, and a radial ribbon of transverse microtubules, which extend laterally under the ciliary furrow and insert in the cortical ridge. A closed ring of paired kinetosomes encircles the cytostome. A brosse begins adjacent to the oral pairs and extends posteriorly for one-fourth to one-half the cell's length. Autapomorphies for Placus include bowling pin-shaped toxicysts extruded onto a distinct area of the cell surface immediately posterior to the brosse, and a brosse kinety consisting of a single row of paired cilia. Placus and its sister taxon Spathidiopsis both have spiraling kineties composed of single cilia inserted into the side of the ciliary furrow. Spathidiopsis can be distinguished from Placus because it has a brosse consisting of two kineties (i.e. one composed of paired cilia, the other of single cilia), a row of rod-shaped toxicyst-bearing palps extending around one side of the oral area and along the length of the brosse, and a mid-cell cortical inpocketing containing toxicysts and a segment of the brosse. A revised listing of species assigned to the family Placidae is given.

Changes in the transcript levels of microtubule-associated protein MAP65-1 during reorientation of cortical microtubules in azuki bean epicotyls

The relation between orientation of cortical microtubules and the transcript levels of a 65 kDa microtubule-associated protein (VaMAP65-1) was investigated along epicotyls of azuki bean (Vigna angularis) seedlings. In addition, the effects of 1-aminocyclopropane-1-carboxylic acid (ACC), the immediate precursor of ethylene, and hypergravity, a gravitational force exceeding 1g, on the relation were examined. The percentage of cells with longitudinal microtubule increased, whereas that with transverse microtubules decreased from the apical to the basal regions of epicotyls. The transcript levels of VaMAP65-1 decreased toward the basal region. ACC induced reorientation of cortical microtubules from transverse to longitudinal directions and down-regulation of VaMAP65-1 expression. Hypergravity also induced reorientation of cortical microtubules and down-regulation of the expression. Strong correlations were observed between the percentage of cells with longitudinal or transverse microtubules and the transcript levels of VaMAP65-1. These results suggest that down-regulation of VaMAP65-1 expression is at least partly involved in the regulation of the orientation of cortical microtubules in azuki bean epicotyls.

Differential regulation of cellulose orientation at the inner and outer face of epidermal cells in the Arabidopsis hypocotyl.

It is generally believed that cell elongation is regulated by cortical microtubules, which guide the movement of cellulose synthase complexes as they secrete cellulose microfibrils into the periplasmic space. Transversely oriented microtubules are predicted to direct the deposition of a parallel array of microfibrils, thus generating a mechanically anisotropic cell wall that will favor elongation and prevent radial swelling. Thus far, support for this model has been most convincingly demonstrated in filamentous algae. We found that in etiolated Arabidopsis thaliana hypocotyls, microtubules and cellulose synthase trajectories are transversely oriented on the outer surface of the epidermis for only a short period during growth and that anisotropic growth continues after this transverse organization is lost. Our data support previous findings that the outer epidermal wall is polylamellate in structure, with little or no anisotropy. By contrast, we observed perfectly transverse microtubules and microfibrils at the inner face of the epidermis during all stages of cell expansion. Experimental perturbation of cortical microtubule organization preferentially at the inner face led to increased radial swelling. Our study highlights the previously underestimated complexity of cortical microtubule organization in the shoot epidermis and underscores a role for the inner tissues in the regulation of growth anisotropy.

Microtubules and CESA tracks at the inner epidermal wall align independently of those on the outer wall of light-grownArabidopsishypocotyls

Microtubules are classically described as being transverse, which is perpendicular to the direction of cell elongation. However, fixation studies have indicated that microtubules can be variably aligned across the epidermis of elongating shoots. In addition, microtubules are reported to have different orientations on inner and outer epidermal surfaces, undermining the idea of hoop-reinforcement. Here, long-term movies of Arabidopsis seedlings expressing GFP-TUA6 allowed microtubule alignment to be directly correlated with the rate of elongation within individual growing cells. We also investigated whether microtubule alignment at the inner or the outer epidermal wall better reflected the growth rate. Movies confirmed that transverse microtubules form on the inner wall throughout elongation, but orientation of microtubules is variable at the outer wall, where they tend to become transverse only during episodes of accelerated growth. Because this appears to contradict the concept that circumferential arrays of transverse microtubules or microfibrils are essential for cell elongation, we checked the organisation of cellulose synthase tracks using GFP-CESA3 and found a similar mismatch between trajectories on inner and outer epidermal surfaces. We conclude that microtubule alignment on the inner wall appears to be a more stable predictor of growth anisotropy, whereas outer-wall alignment is more sensitive to the elongation rate.