The Pellicle-Another Strategy of the Root Apex Protection against Mechanical Stress?

In grasses, the apical part of the root is covered by a two-layered deposit of extracellular material, the pellicle, which together with the outer periclinal wall of protodermal cells forms the three-layered epidermal surface. In this study, the effect of mechanical stress on the pellicle was examined. An experiment was performed, in which maize roots were grown in narrow diameter plastic tubes with conical endings for 24 h. Two groups of experimental roots were included in the analysis: stressed (S) roots, whose tips did not grow out of the tubes, and recovering (R) roots, whose apices grew out of the tube. Control (C) roots grew freely between the layers of moist filter paper. Scanning electron microscopy and confocal microscopy analysis revealed microdamage in all the layers of the epidermal surface of S roots, however, protodermal cells in the meristematic zone remained viable. The outermost pellicle layer was twice as thick as in C roots. In R roots, large areas of dead cells were observed between the meristematic zone and the transition zone. The pellicle was defective with a discontinuous and irregular outermost layer. In the meristematic zone the pellicle was undamaged and the protodermal cells were intact. The results lead to the conclusion that the pellicle may prevent damage to protodermal cells, thus protecting the root apical meristem from the negative effects of mechano-stress.

Detecting Brachypodium distachyon Chromosomes Bd4 and Bd5 in MH- and X-Ray-Induced Micronuclei Using mcFISH.

Micronuclei are biomarkers of genotoxic effects and chromosomal instability. They are formed when chromosome fragments or whole chromosomes fail to disjoin into daughter nuclei. We present qualitative and quantitative analyses of the involvement of specific chromosome regions of chromosomes Bd4 and Bd5 in the formation of micronuclei of Brachypodium distachyon root tip cells following maleic hydrazide (MH) treatment and X-radiation. This is visualised by cytomolecular approaches using bacterial artificial chromosome (BAC)-based multicolour fluorescence in situ hybridisation (mcFISH) in combination with 5S and 25S rDNA probes. The results showed that the long arm of submetacentric chromosome Bd4 forms micronuclei at twice the frequency of its short arm, suggesting that the former is more prone to double-strand breaks (DSBs). In contrast, no difference was observed in the frequency of micronuclei derived from the long and short arms of submetacentric chromosome Bd5. Interestingly, the proximal region of the short arm of Bd5 is more prone to DSBs than its distal part. This demonstrates that 5S rDNA and 35S rDNA loci are not "hot spots" for DNA breaks after the application of these mutagens.

Topological traits of a cellular pattern versus growth rate anisotropy in radish roots.

The topology of a cellular pattern, which means the spatial arrangement of cells, directly corresponds with cell packing, which is crucial for tissue and organ functioning. The topological features of cells that are typically analyzed are the number of their neighbors and the cell area. To date, the objects of most topological studies have been the growing cells of the surface tissues of plant and animal organs. Some of these researches also provide verification of Lewis's Law concerning the linear correlation between the number of neighboring cells and the cell area. Our aim was to analyze the cellular topology and applicability of Lewis's Law to an anisotropically growing plant organ. The object of our study was the root apex of radish. Based on the tensor description of plant organ growth, we specified the level of anisotropy in specific zones (the root proper, the columella of the cap and the lateral parts of the cap) and in specific types of both external (epidermis) and internal tissues (stele and ground tissue) of the apex. The strongest anisotropy occurred in the root proper, while both zones of the cap showed an intermediate level of anisotropy of growth. Some differences in the topology of the cellular pattern in the zones were also detected; in the root proper, six-sided cells predominated, while in the root cap columella and in the lateral parts of the cap, most cells had five neighbors. The correlation coefficient rL between the number of neighboring cells and the cell area was high in the apex as a whole as well as in all of the zones except the root proper and in all of the tissue types except the ground tissue. In general, Lewis's Law was fulfilled in the anisotropically growing radish root apex. However, the level of the applicability (rL value) of Lewis's Law was negatively correlated with the level of the anisotropy of growth, which may suggest that in plant organs in the regions of anisotropic growth, the number of neighboring cells is less dependent on the cell size.

Dissecting the chromosomal composition of mutagen-induced micronuclei in Brachypodium distachyon using multicolour FISH.

Background and Aims: Brachypodium distachyon (Brachypodium) is a model species for temperate cereals and other economically important grasses. Its favourable cytogenetic features and advanced molecular infrastructure make it a good model for understanding the mechanisms of instability of plant genomes after mutagenic treatment. The aim of this study was to qualitatively and quantitatively assess the composition and origin of micronuclei arising from genomic fracture, and to detect possible 'hot spots' for mutagen-induced DNA breaks. Methods: Seeds of Brachypodium were treated with maleic hydrazide (MH) or X-rays. The structure of mutagen-induced micronuclei was analysed in root-tip meristematic cells using multicolour fluorescence in situ hybridization (mcFISH) with various repetitive (5S rDNA, 25S rDNA, telomeric, centromeric) and low-repeat [small and large pools of bacterial artificial chromosome (BAC) clones specific for chromosome Bd1] DNA sequences. Key Results: The majority of micronuclei derive from large, acentric fragments. X-rays caused more interstitial DNA breaks than MH. Double-strand breaks rarely occurred in distal chromosome regions. Bd1 contributed to the formation of more mutagen-induced micronuclei than expected from random chromosome involvement. Conclusions: mcFISH with chromosome-specific BAC clones offers insight into micronuclei composition, in so far as it allows their origin and formation to be determined more specifically. A reliable assay for micronuclei composition is crucial for the development of modern genotoxicity tests using plant cells. The combination of mutagenic treatments and well-developed cytomolecular resources in Brachypodium make this model species very promising for plant mutagenesis research.

Morphological responses of plant roots to mechanical stress.

Background: Roots are continuously exposed to mechanical pressure and this often results in their morphological modification. Most obvious are changes in the overall form of the root system as well as in the shapes of particular roots. These changes are often accompanied by modifications of the cell pattern and cell morphology. Scope: This review focuses on the morphological responses of roots to mechanical stress. Results of early and recent experiments in which roots have been exposed to mechanical pressure are assembled, analysed and discussed. Research applying different experimental sets, obstacles, media of various compactness and structure are reviewed. An effect of the combination of mechanical stresses with other abiotic stresses on roots, and results of estimating the force exerted by the roots are briefly discussed. Possible consequences of the cell pattern rearrangements are considered. Conclusions: Several modifications in root morphology are commonly reported: (1) decreased root size, (2) radial swelling accompanied by increased radial dimension of the cortex cell layers and (3) enhanced cap cell sloughing. Nevertheless, because of differences between species and individual plants, a universal scenario for root morphological changes resulting from externally applied pressures is not possible. Thus, knowledge of the root response to mechanical impedance remains incomplete. Studies on the mechanical properties of the root as well as on possible modifications in cell wall structure and composition as the elements responsible for the mechanical properties of the plant tissue are required to understand the response of root tissue as a biomaterial.

Growth rate distribution in the forming lateral root of arabidopsis.

BACKGROUND AND AIMS: Microscopic observations of lateral roots (LRs) in Arabidopsis thaliana reveal that the cross-sectional shape of the organ changes from its basal to its apical region. The founder cells for LRs are elongated along the parent root axis, and thus from the site of initiation the base of LRs resemble an ellipse. The circumference of the apical part of LRs is usually a circle. The objective of this study was to analyse the characteristics of changes in the growth field of LRs possessing various shapes in their basal regions. METHODS: The LRs of the wild type (Col-0) and two transgenic arabidopsis lines were analysed. On the basis of measurements of the long and short diameters (DL and DS, respectively) of the ellipse-like figure representing the bases of particular LRs, their asymmetry ratios (DL/DS) were determined. Possible differences between accessions were analysed by applying statistical methods. KEY RESULTS: No significant differences between accessions were detected. Comparisons were therefore made of the maximal, minimal and mean value of the ratio of all the LRs analysed. Taking into consideration the lack of circular symmetry of the basal part, rates of growth were determined at selected points on the surface of LRs by the application of the growth tensor method, a mathematical tool previously applied only to describe organs with rotational symmetry. Maps showing the distribution of growth rates were developed for surfaces of LRs of various asymmetry ratios. CONCLUSIONS: The maps of growth rates on the surfaces of LRs having various shapes of the basal part show differences in both the geometry and the manner of growth, thus indicating that the manner of growth of the LR primordium is correlated to its shape. This is the first report of a description of growth of an asymmetric plant organ using the growth tensor method. The mathematical modelling adopted in the study provides new insights into plant organ formation and shape.

Epigenetic chromatin modifications in barley after mutagenic treatment.

In addition to their normal developmental processes, plants have evolved complex genetic and epigenetic regulatory mechanisms to cope with various environmental stresses. It has been shown that both DNA methylation and histone modifications are involved in DNA damage response to various types of stresses. In this study, we focused on the involvement of two mutagenic agents, chemical (maleic acid hydrazide; MH) and physical (gamma rays), on the global epigenetic modifications of chromatin in barley. Our results indicate that both mutagens strongly influence the level of histone methylation and acetylation. Moreover, we found that gamma irradiation, in contrast to MH, has a more robust influence on the DNA methylation level. This is the first study that brings together mutagenic treatment along with its impact at the level of epigenetic modifications examined using the immunohistochemical method.

Form matters: morphological aspects of lateral root development.

BACKGROUND: The crucial role of roots in plant nutrition, and consequently in plant productivity, is a strong motivation to study the growth and functioning of various aspects of the root system. Numerous studies on lateral roots, as a major determinant of the root system architecture, mostly focus on the physiological and molecular bases of developmental processes. Unfortunately, little attention is paid either to the morphological changes accompanying the formation of a lateral root or to morphological defects occurring in lateral root primordia. The latter are observed in some mutants and occasionally in wild-type plants, but may also result from application of external factors. SCOPE AND CONCLUSIONS: In this review various morphological aspects of lateral branching in roots are analysed. Morphological events occurring during the formation of a typical lateral root are described. This process involves dramatic changes in the geometry of the developing organ that at early stages are associated with oblique cell divisions, leading to breaking of the symmetry of the cell pattern. Several types of defects in the morphology of primordia are indicated and described. Computer simulations show that some of these defects may result from an unstable field of growth rates. Significant changes in both primary and lateral root morphology may also be a consequence of various mutations, some of which are auxin-related. Examples reported in the literature are considered. Finally, lateral root formation is discussed in terms of mechanics. In this approach the primordium is considered as a physical object undergoing deformation and is characterized by specific mechanical properties.

Principal growth directions in development of the lateral root in Arabidopsis thaliana.

BACKGROUND AND AIMS: During lateral root development a new meristem is formed within the mother root body. The main objective of this work was to simulate lateral root formation in Arabidopsis thaliana and to study a potential role of the principal directions in this process. Lateral root growth is anisotropic, so that three principal directions of growth can be distinguished within the organ. This suggests a tensorial character of growth and allows for its description by means of the growth tensor method. METHODS: First features of the cell pattern of developing lateral roots were analysed in A. thaliana and then a tensorial model for growth and division of cells for this case was specified, assuming an unsteady character of the growth field of the organ. KEY RESULTS: Microscopic observations provide evidence that the principal directions of growth are manifested at various developmental stages by oblique cell walls observed in different regions of the primordium. Other significant features observed are atypically shaped large cells at the flanks of young apices, as well as distinct boundaries between the mother root and the primordium. Simulations were performed using a model for growth. In computer-generated sequences the above-mentioned features could be identified. An attempt was made to reconstruct the virtual lateral root that included a consideration of the formation of particular tissue types based on literature data. CONCLUSIONS: In the cell pattern of the developing lateral root the principal directions of growth can be recognized through occurrence of oblique cell divisions. In simulation the role of these directions in cell pattern formation was confirmed, only when cells divide with respect to the principal directions can realistic results be obtained.

Effect of mechanical stress on Zea root apex. I. Mechanical stress leads to the switch from closed to open meristem organization.

The effect of mechanical stress on the root apical meristem (RAM) organization of Zea mays was investigated. In the experiment performed, root apices were grown through a narrowing of either circular (variant I) or elliptical (variant II) shape. This caused a mechanical impedance distributed circumferentially or from the opposite sides in variant I and II, respectively. The maximal force exerted by the growing root in response to the impedance reached the value of 0.15 N for variant I and 0.08 N for variant II. Significant morphological and anatomical changes were observed. The changes in morphology depended on the variant and concerned diminishing and/or deformation of the cross-section of the root apex, and buckling and swelling of the root. Anatomical changes, similar in both variants, concerned transformation of the meristem from closed to open, an increase in the number of the cell layers at the pole of the root proper, and atypical oblique divisions of the root cap cells. After leaving the narrowing, a return to both typical cellular organization and morphology of the apex was observed. The results are discussed in terms of three aspects: the morphological response, the RAM reorganization, and mechanical factors. Assuming that the orientation of division walls is affected by directional cues of a tensor nature, the changes mentioned may indicate that a pattern of such cues is modified when the root apex passes through the narrowing, but its primary mode is finally restored.

The tensor-based model for growth and cell divisions of the root apex. II. Lateral root formation.

In this work, the formation of the virtual lateral root (VLR) is shown. The VLR is formed using the 2D simulation model of growth and cell divisions based on the concept of growth tensor, specified for radish. Growth is generated by the field of growth rates of an unsteady type (GT field). Principal directions of growth (PDGs) are assumed to define the orientation of cell divisions. Temporal sequences of the VLR formation are a result of an application of the GT field to the polygon meshwork representing cell pattern of already initiated primordium. The computer-generated lateral root (LR) develops realistically, and its cell pattern is vivid and similar to that observed in anatomical sections. The real and virtual LRs show similar cellular organization, both originate from a small group of cells situated in two-cell layers of the pericycle and both layers are engaged in the LR development. The LR formation seems to be controlled at the tensor level and individual cells presumably detect PDGs and obey them in the course of the cell divisions. PDGs are postulated to affect the cellular organization of the LR. Using the method of computer simulations, cellular aspects of the LR morphogenesis are discussed.

Application of a changing field of growth rates to a description of root apex formation.

Based on the growth tensor method an unsteady field of growth rates for developing root apex is presented. Maps of growth rates distribution as well as simulations in which the field was applied to initially uniform grid of points are presented. In the simulations, the grid undergoes deformation that resembles new root formation in its axial plane. Four variants of field operation on the grid are shown.