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Geoffrey O Wasteneys

Publications and source records attributed to Geoffrey O Wasteneys.

17 recordsLinked to original sources
PubMed2006

Cellulose synthesis is required for deposition of reticulate wall ingrowths in transfer cells.

Despite the recognized physiological importance of transfer cells, little is known about how these specialized cells achieve localized deposition of cell wall material, leading to amplification of plasma membrane surface area and enhanced membrane transport capacity. This study establishes that cellulose synthesis is a key early factor in the construction of 'reticulate' wall ingrowths, an elaborate but common form of localized wall deposition characteristic of most transfer cells. Using field emission scanning electron microscopy, wall ingrowths were first visible in epidermal transfer cells of Faba bean cotyledons as raised 'patches' of disorganized and tangled cellulosic material, and, from these structures, ingrowths emerged via further deposition of wall material. The cellulose biosynthesis inhibitors 2,6-dichlorobenzonitrile and isoxaben both caused dramatic reductions in the number of cells depositing wall ingrowths, altered wall ingrowth morphology and visibly disrupted microfibril structure. The restriction of cellulose deposition to discrete patches suggests a novel mechanism for cellulose synthesis in this circumstance. Overall, these results implicate a central role for cellulose synthesis in reticulate wall ingrowth morphology, especially at the initial stage of ingrowth formation, possibly by providing a template for the self-assembly of wall polymers.

Benzamides↗
PubMed2006

Hypersensitivity to cytoskeletal antagonists demonstrates microtubule-microfilament cross-talk in the control of root elongation in Arabidopsis thaliana.

Elongation of diffusely expanding plant cells is thought to be mainly under the control of cortical microtubules. Drug treatments that disrupt actin microfilaments, however, can reduce elongation and induce radial swelling. To understand how microfilaments assist growth anisotropy, we explored their functional interactions with microtubules by measuring how microtubule disruption affects the sensitivity of cells to microfilament-targeted drugs. We assessed the sensitivity to actin-targeted drugs by measuring the lengths and diameters of expanding roots and by analysing microtubule and microfilament patterns in the temperature-sensitive Arabidopsis thaliana mutant microtubule organization 1 (mor1-1), along with other mutants that constitutively alter microtubule arrays. At the restrictive temperature of mor1-1, root expansion was hypersensitive to the microfilament-disrupting drugs latrunculin B and cytochalasin D, while immunofluorescence microscopy showed that low doses of latrunculin B exacerbated microtubule disruption. Root expansion studies also showed that the botero and spiral1 mutants were hypersensitive to latrunculin B. Hypersensitivity to actin-targeted drugs is a direct consequence of altered microtubule polymer status, demonstrating that cross-talk between microfilaments and microtubules is critical for regulating anisotropic cell expansion.

Actin Cytoskeleton↗
PubMed2005

MICROTUBULE ORGANIZATION 1 regulates structure and function of microtubule arrays during mitosis and cytokinesis in the Arabidopsis root.

MICROTUBULE ORGANIZATION 1 (MOR1) is a plant member of the highly conserved MAP215/Dis1 family of microtubule-associated proteins. Prior studies with the temperature-sensitive mor1 mutants of Arabidopsis (Arabidopsis thaliana), which harbor single amino acid substitutions in an N-terminal HEAT repeat, proved that MOR1 regulates cortical microtubule organization and function. Here we demonstrate by use of live cell imaging and immunolabeling that the mor1-1 mutation generates specific defects in the microtubule arrays of dividing vegetative cells. Unlike the universal cortical microtubule disorganization in elongating mor1-1 cells, disruption of mitotic and cytokinetic microtubule arrays was not detected in all dividing cells. Nevertheless, quantitative analysis identified distinct defects in preprophase bands (PPBs), spindles, and phragmoplasts. In nearly one-half of dividing cells at the restrictive temperature of 30 degrees C, PPBs were not detected prior to spindle formation, and those that did form were often disrupted. mor1-1 spindles and phragmoplasts were short and abnormally organized and persisted for longer times than in wild-type cells. The reduced length of these arrays predicts that the component microtubule lengths are also reduced, suggesting that microtubule length is a critical determinant of spindle and phragmoplast structure, orientation, and function. Microtubule organizational defects led to aberrant chromosomal arrangements, misaligned or incomplete cell plates, and multinucleate cells. Antiserum raised against an N-terminal MOR1 sequence labeled the full length of microtubules in interphase arrays, PPBs, spindles, and phragmoplasts. Continued immunolabeling of the disorganized and short microtubules of mor1-1 at the restrictive temperature demonstrated that the mutant mor1-1(L174F) protein loses function without dissociating from microtubules, providing important insight into the mechanism by which MOR1 may regulate microtubule length.

Arabidopsis↗
PubMed2005

Establishing and maintaining axial growth: wall mechanical properties and the cytoskeleton.

Organ morphology depends on cell placement and directional cell expansion. Microtubules are involved in both of these processes so genetic approaches to understand the role microtubules play in organ expansion are not straightforward. Our use of the temperature-sensitive mor1-1 mutants led to the surprising discovery that Arabidopsis thaliana (L.) Heynh. root cells can establish and maintain transverse cellulose texture without well organized microtubule arrays. This work also demonstrated that cells can lose the ability to expand anisotropically without losing transversely oriented cellulose microfibrils. We suggest that microtubule disruption affects the cell's ability to generate long cellulose microfibrils, which may be essential for achieving growth anisotropy. Thus organ shape may depend not only on the orientation but also on the relative length of cellulose microfibrils during axis establishment and growth. More recent work has shown an important correlation between microtubule organization and the deposition patterns of the glycosylphosphatidylinositol (GPI)-anchored wall protein COBRA. Loss of microtubule organization is associated with the dissipation of transverse banding patterns of COBRA, suggesting that COBRA's function in maintaining anisotropic expansion may be microtubule-dependent.

Anisotropy↗
PubMed2005

COBRA, an Arabidopsis extracellular glycosyl-phosphatidyl inositol-anchored protein, specifically controls highly anisotropic expansion through its involvement in cellulose microfibril orientation.

The orientation of cell expansion is a process at the heart of plant morphogenesis. Cellulose microfibrils are the primary anisotropic material in the cell wall and thus are likely to be the main determinant of the orientation of cell expansion. COBRA (COB) has been identified previously as a potential regulator of cellulose biogenesis. In this study, characterization of a null allele, cob-4, establishes the key role of COB in controlling anisotropic expansion in most developing organs. Quantitative polarized-light and field-emission scanning electron microscopy reveal that loss of anisotropic expansion in cob mutants is accompanied by disorganization of the orientation of cellulose microfibrils and subsequent reduction of crystalline cellulose. Analyses of the conditional cob-1 allele suggested that COB is primarily implicated in microfibril deposition during rapid elongation. Immunodetection analysis in elongating root cells revealed that, in agreement with its substitution by a glycosylphosphatidylinositol anchor, COB was polarly targeted to both the plasma membrane and the longitudinal cell walls and was distributed in a banding pattern perpendicular to the longitudinal axis via a microtubule-dependent mechanism. Our observations suggest that COB, through its involvement in cellulose microfibril orientation, is an essential factor in highly anisotropic expansion during plant morphogenesis.

Arabidopsis↗
PubMed2004

Transient exposure to ethylene stimulates cell division and alters the fate and polarity of hypocotyl epidermal cells.

After transient exposure to the gaseous hormone ethylene, dark-grown cucumber (Cucumis sativus) hypocotyls developed unusual features. Upon ethylene's removal, the developing epidermis showed significant increases in cell division rates, producing an abundance of guard cells and trichomes. These responses to ethylene depended on the stage of development at the time of ethylene exposure. In the upper region of the hypocotyl, where cells were least differentiated at the onset of ethylene treatment, complex, multicellular protuberances formed. Further down the hypocotyl, where stomata and trichomes were beginning to develop at the onset of ethylene exposure, an increase in the number of stomata and trichomes was observed. Stomatal complexes developing after the ethylene treatment had a significant increase in the number of stomatal subsidiary cells and the number of cells per trichome increased. Analysis of division patterns in stomatal complexes indicated that exposure to ethylene either suspended or altered cell fate. Ethylene also altered cell division polarity, resulting in aberrant stomatal complexes and branched trichomes. To our knowledge, the results of this study demonstrate for the first time that transient treatment with physiological concentrations of ethylene can alter cell fate and increase the propensity of cells to divide.

Cell Differentiation↗
PubMed2004

Progress in understanding the role of microtubules in plant cells.

Microtubules have long been known to play a key role in plant cell morphogenesis, but just how they fulfill this function is unclear. Transverse microtubules have been thought to constrain the movement of cellulose synthase complexes in order to generate transverse microfibrils that are essential for elongation growth. Surprisingly, some recent studies demonstrate that organized cortical microtubules are not essential for maintaining or re-establishing transversely oriented cellulose microfibrils in expanding cells. At the same time, however, there is strong evidence that microtubules are intimately associated with cellulose synthesis activity, especially during secondary wall deposition. These apparently conflicting results provide important clues as to what microtubules do at the interface between the cell and its wall. I hypothesize that cellulose microfibril length is an important parameter of wall mechanics and suggest ways in which microtubule organization may influence microfibril length. This concept is in line with current evidence that links cellulose synthesis levels and microfibril orientation. Furthermore, in light of new evidence showing that a wide variety of proteins bind to microtubules, I raise the broader question of whether a major function of plant microtubules is in modulating signaling pathways as plants respond to sensory inputs from the environment.

Cell Division↗
PubMed2003

Ethylene stimulates endoreduplication but inhibits cytokinesis in cucumber hypocotyl epidermis.

The effects of ethylene on cell division are generally considered inhibitory. In this study, we demonstrate that transient ethylene exposure, while suppressing cytokinesis, stimulates DNA synthesis. We monitored DNA synthesis and cytokinesis in the epidermis of cucumber (Cucumis sativus) hypocotyls, an organ whose post-germination development involves strictly limited cell division. During exposure to ethylene, DNA synthesis, assessed by the incorporation of the thymidine homolog 5-bromo-2'-deoxyuridine, was detected in 20% of the epidermal cells, whereas DNA synthesis was nearly undetectable in normal air. Cytofluorometric analysis of nuclei in affected cells showed an up to 8-fold increase in DNA content. During this time, new cell plate formation was not detected. However, shortly after ethylene was removed, DNA content was rapidly restored to 2C (diploid) levels in all cells, and new cell plate formation dramatically increased. These results demonstrate that ethylene promotes DNA synthesis and its endoreduplication but inhibits cytokinesis, thereby maintaining some cells in G2 phase.

Cell Division↗
PubMed2003

Cellulose microfibril alignment recovers from DCB-induced disruption despite microtubule disorganization.

Cellulose microfibril deposition patterns define the direction of plant cell expansion. To better understand how microfibril alignment is controlled, we examined microfibril orientation during cortical microtubule disruption using the temperature-sensitive mutant of Arabidopsis thaliana, mor1-1. In a previous study, it was shown that at restrictive temperature for mor1-1, cortical microtubules lose transverse orientation and cells lose growth anisotropy without any change in the parallel arrangement of cellulose microfibrils. In this study, we investigated whether a pre-existing template of well-ordered microfibrils or the presence of well-organized cortical microtubules was essential for the cell to resume deposition of parallel microfibrils. We first transiently disrupted the parallel order of microfibrils in mor1-1 using a brief treatment with the cellulose synthesis inhibitor 2,6-dichlorobenzonitrile (DCB). We then analysed the alignment of recently deposited cellulose microfibrils (by field emission scanning electron microscopy) as cellulose synthesis recovered and microtubules remained disrupted at the mor1-1 mutant's non-permissive culture temperature. Despite the disordered cortical microtubules and an initially randomized wall texture, new cellulose microfibrils were deposited with parallel, transverse orientation. These results show that transverse cellulose microfibril deposition requires neither accurately transverse cortical microtubules nor a pre-existing template of well-ordered microfibrils. We also demonstrated that DCB treatments reduced the ability of cortical microtubules to form transverse arrays, supporting a role for cellulose microfibrils in influencing cortical microtubule organization.

Arabidopsis↗
PubMed2003

Ethylene modulates root-wave responses in Arabidopsis.

When stimulated to bend downward by being held at 45 degrees off vertical but unable to penetrate into agar-based media, Arabidopsis roots develop waving and looping growth patterns. Here, we demonstrate that ethylene modulates these responses. We determined that agar-containing plates sealed with low-porosity film generate abiotic ethylene concentrations of 0.1 to 0.3 microL L(-1), whereas in plates wrapped with porous tape, ethylene remains at trace levels. We demonstrate that exogenous ethylene at concentrations as low as a few nanoliters per liter modulates root waving, root growth direction, and looping but through partly different mechanisms. Nutrients and Suc modify the effects of ethylene on root waving. Thus, ethylene had little effect on temporal wave frequency when nutrients were omitted but reduced it significantly on nutrient-supplemented agar. Suc masked the ethylene response. Ethylene consistently suppressed the normal tendency for roots of Landsberg erecta to skew to the right as they grow against hard-agar surfaces and also generated righthanded petiole twisting. Furthermore, ethylene suppressed root looping, a gravity-dependent growth response that was enhanced by high nutrient and Suc availability. Our work demonstrates that cell file twisting is not essential for root waving or skewing to occur. Differential flank growth accounted for both the extreme root waving on zero-nutrient plates and for root skewing. Root twisting was nutrient-dependent and was thus strongly associated with the looping response. The possible role of auxin transport in these responses and the involvement of circadian rhythms are discussed.

Arabidopsis↗
PubMed2003

Mutation or drug-dependent microtubule disruption causes radial swelling without altering parallel cellulose microfibril deposition in Arabidopsis root cells.

As critical determinants of growth anisotropy in plants, cortical microtubules are thought to constrain the movement of cellulose synthase complexes and thus align newly deposited cellulose microfibrils. We tested this cellulose synthase constraint model using the temperature-sensitive mor1-1 mutant of Arabidopsis. Contrary to predictions, the disruption of cortical microtubules in mor1-1 root epidermal cells led to left-handed root twisting and radial swelling but did not alter the transverse orientation of cellulose microfibrils. We also found that drug-dependent disassembly or hyperstabilization of cortical microtubules did not alter the parallel order of cellulose microfibrils. By measuring cellulose content in mor1-1 seedlings, we verified that cellulose synthesis is not reduced at the restrictive temperature. The independence of cortical microtubule organization and cellulose microfibril alignment was supported by the observation that double mutants of mor1-1 and rsw1-1, the cellulose-deficient mutant with misaligned microfibrils, had additive phenotypes. Our results suggest that cortical microtubules regulate growth anisotropy by some mechanism other than cellulose microfibril alignment or synthesis.

Arabidopsis↗
PubMed2003

Remodeling the cytoskeleton for growth and form: an overview with some new views.

The cytoskeleton coordinates all aspects of growth in plant cells, including exocytosis of membrane and wall components during cell expansion. This review seeks to integrate current information about cytoskeletal components in plants and the role they play in generating cell form. Advances in genome analysis have fundamentally changed the nature of research strategies and generated an explosion of new information on the cytoskeleton-associated proteins, their regulation, and their role in signaling to the cytoskeleton. Some of these proteins appear novel to plants, but many have close homologues in other eukaryotic systems. It is becoming clear that the mechanisms behind cell growth are essentially similar across the growth continuum, which ranges from tip growth to diffuse expansion. Remodeling of the actin cytoskeleton at sites of exocytosis is an especially critical feature of polarized and may also contribute to axial growth. We evaluate the most recent work on the signaling mechanisms that continually remodel the actin cytoskeleton via the activation of actin-binding proteins (ABPs) and consider the role the microtubule cytoskeleton plays in this process.

Actins↗
PubMed2002

Microtubule organization in the green kingdom: chaos or self-order?

Plant microtubule arrays differ fundamentally from their animal, fungal and protistan counterparts. These differences largely reflect the requirements of plant composite polymer cell walls and probably also relate to the acquisition of chloroplasts. Plant microtubules are usually dispersed and lack conspicuous organizing centres. The key to understanding this dispersed nature is the identification of proteins that interact with and regulate the spatial and dynamic properties of microtubules. Over the past decade, a number of these proteins have been uncovered, including numerous kinesin-related proteins and a 65 kDa class of structural microtubule-associated proteins that appear to be unique to plants. Mutational analysis has identified MOR1, a probable stabilizer of microtubules that is a homologue of the TOGp-XMAP215 class of high-molecular-weight microtubule-associated proteins, and a katanin p60 subunit homologue implicated in the severing of microtubules. The identification of these two proteins provides new insights into the mechanisms controlling microtubule assembly and dynamics, particularly in the dispersed cortical array found in highly polarized plant cells.

Adenosine Triphosphatases↗
PubMed2002

Mutant alleles of Arabidopsis RADIALLY SWOLLEN 4 and 7 reduce growth anisotropy without altering the transverse orientation of cortical microtubules or cellulose microfibrils.

The anisotropic growth of plant cells depends on cell walls having anisotropic mechanical properties, which are hypothesized to arise from aligned cellulose microfibrils. To test this hypothesis and to identify genes involved in controlling plant shape, we isolated mutants in Arabidopsis thaliana in which the degree of anisotropic expansion of the root is reduced. We report here the characterization of mutants at two new loci, RADIALLY SWOLLEN 4 (RSW4) and RSW7. The radial swelling phenotype is temperature sensitive, being moderate (rsw7) or negligible (rsw4) at the permissive temperature, 19 degrees C, and pronounced at the restrictive temperature, 30 degrees C. After transfer to 30 degrees C, the primary root's elongation rate decreases and diameter increases, with all tissues swelling radially. Swelling is accompanied by ectopic cell production but swelling is not reduced when the extra cell production is eliminated chemically. A double mutant was generated, whose roots swell constitutively and more than either parent. Based on analytical determination of acid-insoluble glucose, the amount of cellulose was normal in rsw4 and slightly elevated in rsw7. The orientation of cortical microtubules was examined with immunofluorescence in whole mounts and in semi-thin plastic sections, and the orientation of microfibrils was examined with field-emission scanning electron microscopy and quantitative polarized-light microscopy. In the swollen regions of both mutants, cortical microtubules and cellulose microfibrils are neither depleted nor disoriented. Thus, oriented microtubules and microfibrils themselves are insufficient to limit radial expansion; to build a wall with high mechanical anisotropy, additional factors are required, supplied in part by RSW4 and RSW7.

Anisotropy↗