Biomedical subjects

Peter Nick

Publications and source records attributed to Peter Nick.

At least 19 recordsLinked to original sources

PubMed2006

Actin is bundled in activation-tagged tobacco mutants that tolerate aluminum.

A panel of aluminum-tolerant (AlRes) mutants was isolated by protoplast-based T-DNA activation tagging in the tobacco cultivar SR1. The mutants fell into two phenotypic classes: a minority of the mutants were fertile and developed similarly to the wild type (type I), the majority was male-sterile and grew as semi-dwarfs (type II). These traits, along with the aluminum tolerance, were inherited in a monogenic dominant manner. Both types of mutants were characterized by excessive bundling of actin microfilaments and by a strongly increased abundance of actin, a phenotype that could be partially phenocopied in the wild type by treatment with aluminum chloride. The actin bundles could be dissociated into finer strands by addition of exogenous auxin in both types of mutants. However, actin microfilaments and leaf expansion were sensitive to blockers of actin assembly in the wild type and in the mutants of type I, whereas they were more tolerant in the mutants of type II. The mutants of type II displayed a hypertrophic development of vasculature, manifest in form of supernumerary leaf veins and extended xylem layers in stems and petioles. Whereas mutants of type I were characterized by a normal, but aluminum-tolerant polar auxin-transport, auxin-transport was strongly promoted in the mutants of type II. The phenotype of these mutants is discussed in terms of reduced endocytosis leading, concomitantly with aluminum tolerance, to changes in polar auxin transport.

Actins↗

PubMed2005

Cholodny-Went revisited: a role for jasmonate in gravitropism of rice coleoptiles.

Gravitropism is explained by the Cholodny-Went hypothesis: the basipetal flow of auxin is diverted laterally. The resulting lateral auxin gradient triggers asymmetric growth. However, the Cholodny-Went hypothesis has been questioned repeatedly because the internal auxin gradient is too small to account for the observed growth asymmetry. Therefore, an additional gradient in indolyl-3-acetic acid (IAA) sensitivity has been suggested (Brauner and Hager in Planta 51:115-147, 1958). We challenged the Cholodny-Went hypothesis for gravitropism of rice coleoptiles (Oryza sativa L.) and found it to be essentially true. However, we observed, additionally, that the two halves of gravitropically stimulated coleoptiles responded differentially to the same amount of exogenous auxin: the auxin response is reduced in the upper flank but normal in the lower flank. This indicates that the auxin-gradient is amplified by a gradient of auxin responsiveness. Hormone contents were measured across the coleoptile by a GC-MS/MS technique and a gradient of jasmonate was detected opposing the auxin gradient. Furthermore, the total content of jasmonate increased during the gravitropic response. Jasmonate gradient and increase persist even when the lateral IAA gradient is inhibited by 1-N-naphtylphtalamic acid. Flooding with jasmonate delays the onset of gravitropic bending. Moreover, a jasmonate-deficient rice mutant bends more slowly and later than the wild type. We discuss a role of jasmonate as modulator of auxin responsiveness in gravitropism.

Acetates↗

PubMed2005

Auxin-dependent cell division and cell elongation. 1-Naphthaleneacetic acid and 2,4-dichlorophenoxyacetic acid activate different pathways.

During exponential phase, the tobacco (Nicotiana tabacum) cell line cv Virginia Bright Italia-0 divides axially to produce linear cell files of distinct polarity. This axial division is controlled by exogenous auxin. We used exponential tobacco cv Virginia Bright Italia-0 cells to dissect early auxin signaling, with cell division and cell elongation as physiological markers. Experiments with 1-naphthaleneacetic acid (NAA) and 2,4-dichlorophenoxyacetic acid (2,4-D) demonstrated that these 2 auxin species affect cell division and cell elongation differentially; NAA stimulates cell elongation at concentrations that are much lower than those required to stimulate cell division. In contrast, 2,4-D promotes cell division but not cell elongation. Pertussis toxin, a blocker of heterotrimeric G-proteins, inhibits the stimulation of cell division by 2,4-D but does not affect cell elongation. Aluminum tetrafluoride, an activator of the G-proteins, can induce cell division at NAA concentrations that are not permissive for division and even in the absence of any exogenous auxin. The data are discussed in a model where the two different auxins activate two different pathways for the control of cell division and cell elongation.

2,4-Dichlorophenoxyacetic Acid↗

PubMed2004

The jasmonate-free rice mutant hebiba is affected in the response of phyA'/phyA" pools and protochlorophyllide biosynthesis to far-red light.

Phytochrome (phy) A in its two native isoforms (phyA' and phyA") and the active (Pchlide(655)) and inactive (Pchlide(633)) protochlorophyllides were investigated by low-temperature fluorescence spectroscopy in the tips of rice (Oryza sativa L. Japonica cv Nihonmasari) coleoptiles from wild type (WT) and the jasmonate-deficient mutant hebiba. The seedlings were either grown in the dark or under pulsed (FRp) or continuous (FRc) far-red light (lambda(a) >/= 720 nm) of equal fluences. In the dark, the mutant had a long mesocotyl and a short coleoptile, whereas the situation was reversed under FR: short mesocotyl and long coleoptile, suggesting that the effect is mediated by phyA. Under these conditions the WT displayed a short coleoptile and emergence of the first leaf. In the dark, the spectroscopic and photochemical properties of phyA, its content and the proportion of its two pools, phyA' and phyA", were virtually identical between WT and hebiba. However, the total content of protochlorophyllides was higher in the mutant. Upon illumination with FRc, [phyA] declined in the WT and the ratio between phyA' and phyA" shifted towards phyA". In hebiba, the light-induced decline of [phyA] was less pronounced and the ratio between phyA' and phyA" did not shift. Moreover, in the WT, FRp stimulated the biosynthesis of Pchlide(655), whereas FRc was inhibiting. In contrast, in the mutant, both FRp and FRc stimulated the synthesis of Pchlide(655). This means that FRc caused the opposite effect in hebiba. This difference correlates with a slower photodestruction of primarily the light-labile phyA' pool in hebiba.

Cyclopentanes↗

PubMed2004

A large plant beta-tubulin family with minimal C-terminal variation but differences in expression.

Tubulins, as the major structural component of microtubules (MT), are highly conserved throughout the entire eukaryotic kingdom. They consist of alpha/beta heterodimers. Both monomers, at least in multicellular organisms, are encoded by gene families. In higher plants up to eight beta-tubulin isotypes, mostly differing in their very C-termini, have been described. These variable beta-tubulin C-termini have been discussed in the context of functional microtubule diversity. However, in plants, in contrast to vertebrates, functional isotype specificity remains yet to be demonstrated. Unlike higher plants, unicellular green algae in general do not exhibit isotypic variations. The moss Physcomitrella patens is a phylogenetic intermediate between higher plants and green algae. We isolated six beta-tubulin genes from Physcomitrella, named PpTub1 to 6. We show that the exon/intron structure, with the exception of one additional intron in PpTub6, is identical with that of higher plants, and that some members of the family are differentially expressed. Moreover, we find that all Physcomitrella isotypes are highly conserved and, most strikingly, are almost identical within their C-terminal amino acids (aa). This evolutionary ancient and large beta-tubulin gene family without significant isotypic sequence variation points to a role of differential regulation in the evolution of plant tubulin isotypes.

Amino Acid Sequence↗

PubMed2004

Arabidopsis myosin XI mutant is defective in organelle movement and polar auxin transport.

Myosins are eukaryotic molecular motors moving along actin filaments. Only a small set of myosin classes is present in plants, in which myosins have been found to play a role in cytoplasmic streaming and chloroplast movement. Whereas most studies have been done on green algae, more recent data suggest a role of higher plant myosin at the postcytokinetic cell wall. Here we characterize a loss-of-function mutation for a myosin of plant-specific class XI and demonstrate myosin functions during plant development in Arabidopsis. T-DNA insertion in MYA2 caused pleiotropic effects, including flower sterility and dwarf growth. Elongation of epidermal cells, such as in hypocotyls and anther filaments, was reduced by up to 50% of normal length. This effect on anther filaments is responsible for flower sterility. In the meristems of root tips, it was evident that cell division was delayed and that cell plates were mislocated. Like zwichel, a kinesin-related mutation causing two-branched trichomes, the mya2 knockout causes branching defects, but here the trichomes remained unbranched. Growth was also impaired in pollen tubes and root hairs, cells that are highly dependent on vesicle transport. A failure in vesicle flow could be directly confirmed, because cytoplasmic streaming of vesicles and, more so, of large endoplasmic reticulum-based organelles was slowed. The defect in vesicle trafficking was accompanied by failures in basipetal auxin transport, measured in stem segments of inflorescences. This result strongly suggests a causal link between auxin-dependent processes and the distribution of vesicles and membrane-bound molecules by plant myosin.

Arabidopsis↗

PubMed2004

Dual targeting of plastid division protein FtsZ to chloroplasts and the cytoplasm.

FtsZ is a filament-forming protein that assembles into a ring at the division site of prokaryotic cells. As FtsZ and tubulin share several biochemical and structural similarities, FtsZ is regarded as the ancestor of tubulin. Chloroplasts--the descendants of endosymbiotic bacteria within plant cells--also harbour FtsZ. In contrast to eubacteria, plants have several different FtsZ isoforms. So far, these isoforms have only been implicated with filamentous structures, rings and networks, inside chloroplasts. Here, we demonstrate that a novel FtsZ isoform in the moss Physcomitrella patens is located not only in chloroplasts but also in the cytoplasm, assembling into rings in both cell compartments. These findings comprise the first report on cytosolic localization of a eukaryotic FtsZ isoform, and indicate that this protein might connect cell and organelle division at least in moss.

Arabidopsis Proteins↗

PubMed2004

Capturing in vivo dynamics of the actin cytoskeleton stimulated by auxin or light.

We present here a transient expression system that allows the response of actin microfilaments to physiological stimuli (changes in auxin content, light) to be observed in single cells in vivo. Etiolated, intact rice seedlings are attached to glass slides, transfected biolistically with talin fused to yellow-fluorescent protein to visualize actin microfilaments, and either treated with auxin or irradiated. The talin marker labels distinct populations of actin that are differentially expressed depending on the physiological state of the coleoptile (active elongation versus ceased elongation). Whereas longitudinal transvacuolar bundles prevail in cells that have ceased to elongate, fine cortical strands are characteristic for elongating cells. The visualized actin structures remain dynamic and responsive to signals. Exogenous auxin triggers a loosening of the bundles and an extension of the cortical strands, whereas irradiation reorientates cortical strands into longitudinal arrays. These responses correspond in quality and timing to the signal responses inferred previously from fixed specimens and biochemical studies. In big advantage over those methods it is now possible to observe them directly at the single cell level. Thus, the rice coleoptile system can be used as a convenient model to study actin dynamics in vivo, in response to physiologically relevant stimuli.

Actin Cytoskeleton↗

PubMed2003

A myosin inhibitor impairs auxin-induced cell division.

The role of myosins for auxin-induced cell division was probed using the inhibitor 2,3-butanedione monoxime in the tobacco cell line VBI-0, where cell elongation and division are axially aligned under the control of auxin. A morphometric analysis revealed that cell division is blocked in a dose-dependent manner, whereas cell expansion continued. In addition, the polarity of terminal cells was impaired resulting in malformed, pear-shaped cells. Early effects of the inhibitor are aberrant features of the cytoarchitecture including a block of vesicle transport, a diffuse broadening of cross walls, and the disorganization of the actin cytoskeleton. The findings are discussed with respect to a possible role of myosins as link between vesicle flow and signal control of cell division.

Actins↗

PubMed2003

Auxin responsiveness of a novel cytochrome p450 in rice coleoptiles.

An early auxin-induced gene was isolated from rice (Oryza sativa L. subsp. japonica cv Nihonmasari) coleoptiles by a fluorescent-labeled differential display screen. The full-length gene contains conserved domains characteristic for the cytochrome p450 superfamily. This gene, designated as CYP87A3, was weakly expressed in dark-grown coleoptiles but was up-regulated rapidly and transiently when coleoptile segments were incubated in 5 microm indole-3-acetic acid. This induction by auxin could not be suppressed by cycloheximide. Depletion of segments from endogenous auxin reduced the amount of CYP87A3 transcripts. The CYP87A3 transcript level was rapidly, although transiently, up-regulated in response to light as well. The observed pattern of gene regulation might indicate a role in the suppression of auxin-induced coleoptile growth. The role of CYP87A3 is discussed with respect to auxin signaling in the regulation of coleoptile growth.

Amino Acid Sequence↗

PubMed2003

Impaired induction of the jasmonate pathway in the rice mutant hebiba.

The elongation of rice (Oryza sativa) coleoptiles is inhibited by light, and this photoinhibition was used to screen for mutants with impaired light response. In one of the isolated mutants, hebiba, coleoptile elongation was stimulated in the presence of red light, but inhibited in the dark. Light responses of endogenous indolyl-3-acetic acid and abscisic acid were identical between the wild type and the mutant. In contrast, the wild type showed a dramatic increase of jasmonate heralded by corresponding increases in the content of its precursor o-phytodienoic acid, whereas both compounds were not detectable in the mutant. The jasmonate response to wounding was also blocked in the mutant. The mutant phenotype was rescued by addition of exogenous methyl jasmonate and o-phytodienoic acid. Moreover, the expression of O. sativa 12-oxophytodienoic acid reductase, an early gene of jasmonic acid-synthesis, is induced by red light in the wild type, but not in the mutant. This evidence suggests a novel role for jasmonates in the light response of growth, and we discuss a cross-talk between jasmonate and auxin signaling. In addition, hebiba represents the first rice mutant in which the induction of the jasmonate pathway is impaired providing a valuable tool to study the role of jasmonates in Graminean development.

Cyclopentanes↗

PubMed2003

Activation-tagged tobacco mutants that are tolerant to antimicrotubular herbicides are cross-resistant to chilling stress.

T-DNA activation tagging was used to generate tobacco mutants with increased tolerance to antimicrotubular herbicides and chilling stress. After transformation, protoplast-derived calli were screened for tolerance to treatments that affect microtubule assembly. In one screen mutants with tolerance to aryl carbamates (a blocker of microtubule assembly) were selected, the second screen was targeted to chilling-tolerant mutants that could survive for several months at 3 degrees C, a third screen combined both factors. The resistance of these mutants to aryl carbamates or chilling was accompanied by resistance of microtubules to these factors. The carbamate tolerant mutants were cross-resistant to chilling stress. This was mirrored by an adaptive reorganization of microtubules and a reduction of microtubule dynamics in response to chilling. The analysis of these mutants suggests (1) that microtubule dynamics limit the tolerance to chilling and EPC, and (2) that the cold sensitivity of microtubules limits chilling tolerance in tobacco.

Cold Temperature↗

PubMed2003

Is microtubule disassembly a trigger for cold acclimation?

Cold acclimation was followed in three cultivars of winter wheat (Triticum aestivum L.) that differ in freezing tolerance, using root growth as the indicator. During acclimation (followed through 7 d at 4 degrees C), growth rate progressively recovered. The recovery was fast in the tolerant, slow in the sensitive cultivars. The development of freezing tolerance was followed by a challenging cold shock administered after various time intervals of acclimation. Acclimation proceeded faster in the tolerant cultivars. Microtubules were monitored during the acclimation period. A rapid, but transient partial disassembly in the tolerant cultivars preceded the formation of cold-stable microtubules and the recovery of growth rate. In contrast, this transient disassembly was absent in the sensitive cultivar. When a transient disassembly was artificially generated by a pulse-treatment with the antimicrotubular herbicide pronamide, this could induce freezing tolerance. The appearance of cold-stable microtubules was accompanied by a reduced abundance of type TUA1/2 alpha-tubulin isotypes. These findings are discussed with respect to a role of microtubule disassembly in the sensing of low-temperature stress.

Acclimatization↗

PubMed2003

Auxin transport synchronizes the pattern of cell division in a tobacco cell line.

The open morphogenesis of plants requires coordination of patterning by intercellular signals. The tobacco (Nicotiana tabacum cv Virginia Bright Italia) cell line VBI-0 provides a simple model system to study the role of intercellular communication in patterning. In this cell line, singular cells divide axially to produce linear cell files of distinct polarity. The trigger for this axial division is exogenous auxin. When frequency distributions of files are constructed over the number of cells per file during the exponential phase of the culture, even numbers are found to be frequent, whereas files consisting of uneven numbers of cells are rare. We can simulate these distributions with a mathematical model derived from nonlinear dynamics, which describes a chain of cell-division oscillators where elementary oscillators are coupled unidirectionally and where the number of oscillators is not conserved. The model predicts several nonintuitive properties of our experimental system. For instance, files consisting of six cells are more frequent than expected from a strictly binary division system. More centrally, the model predicts a polar transport of the coordinating signal. We therefore tested the patterns obtained after treatment with 1-N-naphthylphthalamic acid, an inhibitor of auxin efflux carriers. Using low concentrations of 1-N-naphthylphthalamic acid that leave cell division and axiality of division unaltered, we observe that the frequencies of files with even and uneven cell numbers are equalized. Our findings are discussed in the context of auxin transport as synchronizing signal in cell patterning.

Biological Transport↗

PubMed2003

Visualizing the self-assembly of tubulin with luminescent nanorods.

We succeeded in labeling tubulin with luminescent CdSe nanorods. The labeled tubulin was still able to self-assemble into microtubules, demonstrating that the protein has remained functional. The comparison with rhodamine-labeled tubulin revealed that whereas the rhodamine bleached completely within the observed time frame, the nanorods did not show any bleaching, which makes it possible to follow the key events of self-assembly at high time resolution. Preparation and derivatization of nanorods are described. Water-soluble, protein-reactive group-containing particles were covalently coupled to tubulin. The conjugate was purified by one cycle of assembly and disassembly and used to induce the formation of fluorescent microtubules. These results suggest that the use of luminescent nanorods should allow continuous confocal monitoring of dynamic biological processes.

Biocompatible Materials↗

PubMed2002

The host guides morphogenesis and stomatal targeting in the grapevine pathogen Plasmopara viticola.

The oomycete grape downy mildew (Plasmopara viticola Berk. & Curt. Ex de Bary) is a serious pathogen of grapevine and spreads by extremely efficient cycles of asexual propagation. The high efficiency must involve efficient sensing of the host. We therefore analyzed the time course and morphology of the early development of this pathogen in a host system, by infection of leaf discs of grapevine (Vitis vinifera L. cv. Müller-Thurgau), and in a host-free system. Host factors were demonstrated to influence pathogen development in the following ways: (i) the release of zoospores from mature sporangia was accelerated, (ii) the morphogenesis of the germ tube was coordinated, and (iii) the zoospores were targeted to the stomata by factors that depended on stomata closure. The findings show that the early development of P. viticola is regulated, specifically and coordinately, by factors originating from the host plant.

Morphogenesis↗

PubMed2002

OsARF1, an auxin response factor from rice, is auxin-regulated and classifies as a primary auxin responsive gene.

We screened for auxin-induced genes with an expression correlated to the auxin-induced growth response from rice coleoptiles by fluorescent differential display. A rice homologue of the auxin response factor (ARF) family of transcriptional regulators, OsARF1, was identified. An OsARF1:GFP fusion protein was localized to the nucleus. Steady-state levels of OsARF1 mRNA correlated positively with auxin-dependent differential growth: gravitropic stimulation enhanced the amount of OsARF1 transcript in the lower, faster-growing flank accompanied by a decrease in the upper flank of gravitropically stimulated rice coleoptiles. Exogenous auxin up-regulated the steady-state level of OsARF1 mRNA within 15-30 min. This up-regulation is independent of de novo protein synthesis. Thus, OsARF1 is the first ARF that classifies as an early auxin-responsive gene. The observed auxin-dependent regulation comprises a new level of regulation in auxin-induced gene expression and is discussed as a possible feedback mechanism in plant growth control.

Amino Acid Sequence↗

PubMed2002

Plant RanGAPs are localized at the nuclear envelope in interphase and associated with microtubules in mitotic cells.

In animals and yeast, the small GTP-binding protein Ran has multiple functions - it is involved in mediating (i) the directional passage of proteins and RNA through the nuclear pores in interphase cells; and (ii) the formation of spindle asters, the polymerization of microtubules, and the re-assembly of the nuclear envelope in mitotic cells. Nucleotide binding of Ran is modulated by a series of accessory proteins. For instance, the hydrolysis of RanGTP requires stimulation by the RanGTPase protein RanGAP. Here we report the complementation of the yeast RanGAP mutant rna1 with Medicago sativa and Arabidopsis thaliana cDNAs encoding RanGAP-like proteins. Confocal laser microscopy of Arabidopsis plants overexpressing chimeric constructs of GFP with AtRanGAP1 and 2 demonstrated that the fusion protein is localized to patchy areas at the nuclear envelope of interphase cells. In contrast, the cellular distribution of RanGAPs in synchronized tobacco cells undergoing mitosis is characteristically different. Double-immunofluorescence shows that RanGAPs are co-localized with spindle microtubules during anaphase, with the microtubular phragmoplast and the surface of the daughter nuclei during telophase. Co-assembly of RanGAPs with tubulin correlates with these in vivo observations. The detected localization pattern is consistent with the postulated function of plant RanGAPs in the regulation of nuclear transport during interphase, and suggests a role for these proteins in the organization of the microtubular mitotic structures.

Active Transport, Cell Nucleus↗