Cluster root development in Grevillea robusta (Proteaceae). II. The development of the endodermis in a determinate root and in an indeterminate, lateral root

Light, fluorescence and electron microscopy were employed to follow the development of the endodermis in cluster roots and lateral roots of Grevillea robusta A. Cunn. ex R. Br. Endodermal cells had three different origins: rootlet endodermis arose from the rootlet meristem; endodermis covering the primordium shortly after initiation came from division of parental endodermis; cells at the junction between parent and rootlet endodermis developed from re-differentiated rootlet cortical cells. In the cluster root, the Casparian band formed in three ways, and was not initially present opposite the two sets of single xylem elements in the rootlet stele. A new clearing technique was developed that allowed visualization of xylem, suberized endodermis, Casparian band formation and phenolic compounds. In lateral roots, endodermal differentiation was asynchronous, but was related to position relative to protoxylem poles. However, the observed delay began before these poles had differentiated. At the tip of mature rootlets, which are determinate, the endodermis terminates in a 'dome' of cells, with the initial cell differentiating as an endodermal cell. Results are discussed in terms of determinate development in roots and the spatial and temporal contexts within which this development takes place.     

Effects of growing conditions and development of the underlying exodermis on the vitality of the onion root epidermis

Bulbs of Allium cepa L., which had developed short, adventitious roots, were transferred to various conditions, i.e. vermiculite watered to saturation, vermiculite watered to half saturation, immersed in hydroculture, and immersed in hydroculture except for the proximal 20 mm which was continuously exposed to air. The development of the exodermis occurred in a patchy fashion in many roots but was not influenced by the growing conditions. The vitality of the epidermis declined under all conditions, the rate of decline being inversely related to the ambient moisture level. The differences between the treatments were most evident at the oldest region sampled (120 mm from the root tip) where 4% of the epidermal cells were dead in roots grown in hydroponics. This compared with 62% dead cells in saturated vermiculite, 78% in half-saturated vermiculite and 92% in roots exposed to air. Death of the epidermal cells was not accelerated by the maturation of the underlying exodermis. Epidermal cells which did not overlie a short cell of the exodermis (i.e. were only in contact with long cells) died earlier than the others: this trend was evident even prior to the maturation of the exodermis. These results suggest that the epidermal cells are not well connected symplasmically to the long cells or to the neighbouring epidermal cells. The large majority of epidermal cells (98% of the total) were in contact with a short cell of the exodermis. These epidermal cells tended to die off slowly, even under very favourable ambient conditions. Since these cells provide the major site for ion uptake in roots with a mature exodermis. their death may reduce the efficiency of the root for this activity.     

The ciliary rootlet maintains long-term stability of sensory cilia

The striated ciliary rootlet is a prominent cytoskeleton originating from basal bodies of ciliated cells. Although a familiar structure in cell biology, its function has remained unresolved. In this study, we carried out targeted disruption in mice of the gene for rootletin, a component of the rootlet. In the mutant, ciliated cells are devoid of rootlets. Phototransduction and ciliary beating in sensory and motile cilia initially exhibit no apparent functional deficits. However, photoreceptors degenerate over time, and mutant lungs appear prone to pathological changes consistent with insufficient mucociliary clearance. Further analyses revealed a striking fragility at the ciliary base in photoreceptors lacking rootlets. In vitro assays suggest that the rootlet is among the least dynamic of all cytoskeletons and interacts with actin filaments. Thus, a primary function of the rootlet is to provide structural support for the cilium. Inasmuch as photoreceptors elaborate an exceptionally enlarged sensory cilium, they are especially dependent on the rootlet for structural integrity and long-term survival.     

The formation,morphology and anatomy of cluster root of Lupinus albus L. as dependent on soil type and phosphorus supply

The effect of phosphorus supply on the formation, morphology and anatomy of cluster roots of Lupinus albus L. cv Ultra grown in a loam and two sandy soils was examined relative to its effect on total root length, shoot weight and the phosphorus concentration of the shoots. The loam soil was most conducive to the formation of cluster roots. Cluster roots growing in the sandy soils developed to a lesser extent on plants of an equivalent phosphorus status, suggesting that some biotic or abiotic factors independent of phosphorus supply were also operating. The presence of mature cluster rootlets on a length of lateral root increased the root surface area by 14–22 times of an equal length of lateral roots not bearing cluster rootlets. The application of phosphorus decreased cluster-root length, whereas total root length showed a steady increase. There was an inverse relationship between cluster-root production and phosphorus concentration in shoots ranging from 2 to 8.5 mg g^–1 with the critical phosphorus level for maximum shoot growth being around 2.5 mg g^–1. Cluster roots formed in solution culture were not well developed in comparison with those grown in the loam soil or nutrient solution with added loam soil. The organisation of the cluster rootlet was similar to that of the lateral roots. Mature rootlets lacked an apical meristem and a vascular cambium with a reduced root cap and cortical tissue.     

Comparative ultrastructure of the root system in rhizocephalan barnacles (Crustacea: Cirripedia: Rhizocephala).

Rhizocephalan barnacles are parasites of Crustacea. They lack even the rudiments of an alimentary canal, but infiltrate their hosts with a nutrient-absorbing system of rootlets. We review the ultrastructure of the rootlets using light microscopy, SEM, and TEM in nine species from five families, representing both suborders of the Rhizocephala: from the Kentrogonida Peltogaster paguri, P. curvatus, Peltogasterella sulcata, Cyphosaccus norvegicus (Peltogastridae); Lernaeodiscus porcellanae (Lernaeodiscidae); and Sacculina carcini (Sacculinidae); and from the Akentrogonida Clistosaccus paguri (Clistosaccidae); Chthamalophilus delagei, and Boschmaella japonica (Chthamalophilidae). With the exception of Chthamalophilus delagei, the root system of the investigated species shares numerous apomorphies at the ultrastructural level and displays at all levels specializations that maximize the surface area. The rootlets consist of a cuticle, an epidermis and a subjacent layer of axial cells that often, but not always surround, a central lumen. The rootlets are at all times enclosed in a less than 0.5 microm thick cuticle, which is never molted. The cuticle consists of an inner homogeneous layer with a slightly fibrous structure and an outer, less than 15-nm thick electron-dense layer, from which numerous microcuticular projections extend into the hemolymphatic space of the host. The microcuticular projections consist of the outer electron-dense layer and sometimes a core of the more translucent homogeneous layer. They vary among the species from being simple in Sacculina carcini to exhibiting complex branching patterns in Peltogasterella sulcata and Cyphosaccus norvegicus. Beneath the cuticle the epidermal plasma membrane is thrown into irregularly shaped projections. The epidermal cells are joined by long septate junctions and exhibit the characteristics of a transporting epithelium. Experiments with acid phosphatase revealed activity both in the epidermis and among the microcuticular projections. The projections may therefore form a domain that is important in absorption and extracellular digestion of nutrients from the host. The axial cells contain abundant endoplasmic reticulum and seem to convert absorbed carbohydrates into lipid, which is stored in large droplets. Subepidermal muscle cells cause sinuous movements of the rootlets, but it remains unknown how nutrients are transported along the rootlets towards the external reproductive body. In C. delagei the single, bladder-shaped rootlet lacks both the apical projections in the epidermis, the electron-dense cuticle layer, and the microcuticular projections. We review previous studies on the rhizocephalan root system and discuss functional and phylogenetic aspects of the morphology.     

Isolation, ultrastructure, and protein composition of the flagellar rootlet of Naegleria gruberi

Attached to the basal bodies of Naegleria gruberi flagellates is a striated rootlet or rhizoplast. The rootlet-basal body complex has been isolated by Triton X-100 lysis of deflagellated cells and differential centrifugation through a 25% glycerol medium. Rootlets isolated from mature flagellates are approximately 13 micrometers long but vary from 8 to 15 micrometers in length: they taper at both ends from a maximum width of approximately 0.25 micrometers in the vicinity of the basal bodies. They are highly stable during isolation but can be solubilized by urea, high salt, low pH, or detergent (Sarkosyl). Partial dissociation of rootlets with 1 M urea reveals that they are composed of filaments, approximately 5 nm diameter, associated in a linear fashion to yield the characteristic 21-nm cross-banded appearance. Differential solubilization of rootlets and their associated contaminants allowed identification of a major rootlet protein, comprising at least 50% of any purified rootlet preparation, with an apparent subunit molecular weight of 170,000. The localization of rootlets in situ by indirect immunofluorescence using a specific antibody directed against the purified rootlet protein demonstrated unequivocally that this 170,000-dalton protein is an organelle component.     

Cortical morphogenesis in Paramecium: a transcellular wave of protein phosphorylation involved in ciliary rootlet disassembly

In Paramecium, the morphogenesis of the cortex at cell division, which assures reconstruction of shape and surface pattern, has been shown to involve transcellular signals which spread across the cortex like a wave, originating principally from the oral apparatus. One of the events these signals control is the reorganization of the ciliary rootlets through a cycle of regression and regrowth. The ciliary rootlets are nucleated on the ciliary basal bodies and form a scaffold extending over the entire cell surface that is important in aligning the basal bodies and the unit territories organized around them in longitudinal rows. We present evidence that the mechanism underlying their reorganization is cell-cycle-dependent phosphorylation of the structural proteins which compose the ciliary rootlets. We have isolated the rootlets and prepared a polyclonal antibody against them. In situ immunofluorescence of dividing cells with the anti rootlet antibody, and with the monoclonal antibody MPM-2 specific for phosphoproteins shows that a wave of phosphorylation of the ciliary rootlets spreads across the cell at division and just precedes their regression. Two-dimensional Western blot analysis of cytoskeleton and isolated rootlets along with alkaline phosphatase treatment demonstrates that the rootlets are composed of phosphoproteins, while experiments with interphase and dividing cells provide direct evidence that hyperphosphorylation of these proteins at division brings about disassembly of the structure.     

Rootletin,a novel coiled-coil protein,is a structural component of the ciliary rootlet

The ciliary rootlet, first recognized over a century ago, is a prominent structure originating from the basal body at the proximal end of a cilium. Despite being the largest cytoskeleton, its structural composition has remained unknown. Here, we report a novel 220-kD protein, designated rootletin, found in the rootlets of ciliated cells. Recombinant rootletin forms detergent-insoluble filaments radiating from the centrioles and resembling rootlets found in vivo. An mAb widely used as a marker for vertebrate rootlets recognizes an epitope in rootletin. Rootletin has a globular head domain and a tail domain consisting of extended coiled-coil structures. Rootletin forms parallel in register homodimers and elongated higher order polymers mediated by the tail domain alone. The head domain may be required for targeting to the basal body and binding to a kinesin light chain. In retinal photoreceptors where rootlets appear particularly robust, rootlets extend from the basal bodies to the synaptic terminals and anchor ER membranes along their length. Our data indicate that rootlets are composed of homopolymeric rootletin protofilaments bundled into variably shaped thick filaments. Thus, rootletin is the long-sought structural component of the ciliary rootlet.     

Early development of rat ventral root transitional zone: An immunohistochemical and morphometric study

Bundles of ventral motoneuron axons cross the white matter of the spinal cord, emerge through the cord surface at the CNS-PNS transitional zone (TZ) and continue in the PNS as ventral rootlets. This study identifies immunohistochemical and morphometric changes which characterise the key events in early TZ formation in the rat. E18 is a landmark stage, since it is then that the major events of TZ differentiation are initiated. In the glial processes associated with the TZ, vimentin expression decreases, while that of GFAP increases. In the proximal rootlets the transient expression of CNS markers such as GFAP and of neural adhesion molecules such as HNK-1/N-CAM begin to decrease. Their resulting differential expression clearly defines the CNS-PNS interface. These changes coincide with the arrival of glial nuclei at the TZ. Cell clusters which appear on proximal ventral rootlet surfaces shortly after their emergence from the cord, have by E18 formed an extensive matrix of processes which segregates the axon bundle. This comprises the earliest of two well-defined barriers across the axon bundle. An important function may be to prevent Schwann cell invasion of the cord. Cluster cells display some immunohistochemical features in common with Schwann cells. The second barrier becomes fully established only at P2 and forms the definitive CNS-PNS interface. It consists of processes arising from astrocytes surrounding the TZ. Changes in the nuclear density of the cell types correspond closely to their segregating activity. The immunohistochemical and ultrastructural changes complement one another to deepen and enhance understanding of TZ development.     

Asymmetric properties of the Chlamydomonas reinhardtii cytoskeleton direct rhodopsin photoreceptor localization

The eyespot of the unicellular green alga Chlamydomonas reinhardtii is a photoreceptive organelle required for phototaxis. Relative to the anterior flagella, the eyespot is asymmetrically positioned adjacent to the daughter four-membered rootlet (D4), a unique bundle of acetylated microtubules extending from the daughter basal body toward the posterior of the cell. Here, we detail the relationship between the rhodopsin eyespot photoreceptor Channelrhodopsin 1 (ChR1) and acetylated microtubules. In wild-type cells, ChR1 was observed in an equatorial patch adjacent to D4 near the end of the acetylated microtubules and along the D4 rootlet. In cells with cytoskeletal protein mutations, supernumerary ChR1 patches remained adjacent to acetylated microtubules. In mlt1 (multieyed) mutant cells, supernumerary photoreceptor patches were not restricted to the D4 rootlet, and more anterior eyespots correlated with shorter acetylated microtubule rootlets. The data suggest a model in which photoreceptor localization is dependent on microtubule-based trafficking selective for the D4 rootlet, which is perturbed in mlt1 mutant cells.     

ULTRASTRUCTURE OF THE FLAGELLAR APPARATUS OF PYROBOTRYS (CHLOROPHYCEAE)1

The flagellar apparatus of Pyrobotrys has a number of features that are typical of the Chlorophyceae, but others that are unusual for this class. The two flagella are inserted at the apex, but they extend to the side of the cell toward the outside of the colony, here designated as the ventral side. Four basal bodies are present, two of which extend into flagella. Four microtubular rootlets alternate between the functional and accessory basal bodies. In each cell, the two ventral rootlets are nearly parallel, but the dorsal rootlets are more widely divergent. The rootlets alternate between two and four microtubules each. A striated distal fiber connects the two functional basal bodies in the plane of the flagella. Two additional, apparently nonstriated, fibers connect the basal bodies proximal to the distal fiber. Another striated fiber is associated with each four-membered rootlet near its insertion into the flagellar apparatus. A fine periodic component is associated with each two-membered rootlet. A rhizoplast-like structure extends into the cell from each of the functional basal bodies. The arrangement of these components does not reflect the 180° rotational symmetry that is usually present in the Chlorophyceae, but appears to be derived from a more symmetrical ancestor. It is suggested that the form of the flagellar apparatus is associated with the unusual colony structure of Pyrobotrys.     

Three‐dimensional organization of flagellar basal apparatus in Ectocarpus gametes

The flagellar basal apparatus of the brown alga Ectocarpus siliculosus was re‐investigated in details using transmission electron microscopy and electron tomography. As a result, three‐dimensional structures with spatial arrangement of bands and microtubular flagellar rootlets were observed. Fibrous structures linking the anterior flagellar basal body to the major anterior rootlet (R3) or the bypassing rootlet was newly discovered in this study. A direct attachment from the minor anterior rootlet (R4) to the anterior and posterior basal bodies was also discovered, as were attachments from the minor posterior rootlet (R1) to the deltoid striated band and from the major posterior rootlet (R2) to the posterior fibrous band. The microtubular flagellar rootlets were connected to the bands and to the anterior or posterior basal body. These bands may have a role in maintaining the spatial arrangement of the anterior and posterior flagellar basal bodies and the microtubular flagellar rootlets. A numbering system of the basal body triplets was established by tracing axonemal doublets in the serial sections. From these observations, the precise position of two flagellar basal bodies, bands, and flagellar rootlets was determined.     

Two ways to skin a plant: The analysis of root and shoot epidermal development in Arabidopsis

The post-embryonic architecture of higher plants is derived from the activity of two meristems that are formed in the embryo: the shoot meristem and the root meristem. The epidermis of the shoot is derived from the outermost layer of cells covering the shoot meristem through repeated anticlinal divisions. By contrast, the epidermis of the root is derived from an internal ring of cells, located at the centre of the root meristem, by a precise series of both periclinal and anticlinal divisions. Each epidermis has an independent origin. In Arabidopsis the mature shoot epidermis is composed of a small number of cell types: hair cells (trichomes), stomatal guard cells and other epidermal cells. In shoots, hairs take the form of branched trichomes that are surrounded at their base by a ring of accessory cells in a sheet of epidermal cells. The root epidermis is composed of two cell types: trichoblasts that form root hair cells and atrichoblasts that form non-hair cells. Mutations affecting both the patterning and the morphogenesis of cells in both shoot and root epidermis have recently been described. Most of these mutations affect development in a single epidermis, but at least one, ttg, is involved in development in both epidermal systems.     

Identification of a 195 kDa protein in the striated rootlet: its expression in ciliated and ciliogenic cells

Little is known about the molecular composition of the ciliary rootlet. We raised monoclonal antibodies to a crude preparation of striated rootlets isolated from the human oviduct, and obtained a clone (R4109) that specifically labeled the ciliary rootlets. Rootlets associated with the solitary cilium in secretory cells and fibroblasts were also labeled. R4109 identified a 195-kDa protein by immunoblotting. Ciliogenic cells in the oviduct epithelium of young mice were labeled in the globular and/or granular pattern by R4109 by immunofluorescence microscopy. Immunoelectron microscopy showed that they corresponded to fibrogranular complex and dense granule, respectively. The result demonstrated that the 195-kDa protein is a component common to the striated rootlet and dense granule, and thus suggested that dense granules are involved in the rootlet formation.     

Fine structure of the zoospore ofUlothrix belkae with emphasis on the flagellar apparatus

Summary The anterior end of the zoospore ofUlothrix belkae has been examined in detail and is compared toStigeoclonium and other filamentous green algae. The nature of the symmetry of green algal motile cells is discussed and the term, 180° rotational symmetry, is proposed to describe the type of arrangement of anterior end components seen inU. belkae, including the four basal bodies, rootlets and striated fibers. The four microtubular rootlets are cruciately arranged. A striated microtubule-associated component (SMAC) has a periodicity of 6.4 nm and extends with each 2-membered rootlet posteriorly into the cell. One 5-membered rootlet passes very near the eyespot. Phylogeny in green algal motile cells is discussed.     

Developmental changes in peanut root structure during root growth and root-structure modification by nodulation

Background and Aims

Basic information about the root and root nodule structure of leguminous crop plants is incomplete, with many aspects remaining unresolved. Peanut (Arachis hypogaea) forms root nodules in a unique process. Structures of various peanut root types were studied with emphasis on insufficiently characterized lateral roots, changes in roots during their ontogenesis and root modification by nodule formation.

Methods

Peanut plants were grown in the field, in vermiculite or in filter paper. The taproot, first-order and second-order lateral roots and root nodules were analysed using bright-field and fluorescence microscopy with hand sections and resin sections.

Key Results

Three root categories were recognized. The primary seminal root was thick, exhibiting early and intensive secondary thickening mainly on its base. It was tetrarch and contained broad pith. First-order lateral roots were long and thin, with limited secondary thickening; they contained no pith. Particularly different were second- and higher-order lateral roots, which were anatomically simple and thin, with little or no secondary growth. Unusual wall ingrowths were visible in the cells of the central part of the cortex in the first-order and second-order lateral roots. The nodule body was formed at the junction of the primary and lateral roots by the activity of proliferating cells derived originally from the pericycle.

Conclusions

Two morphologically and anatomically distinct types of lateral roots were recognized: long, first-order lateral roots, forming the skeleton of the root system, and thin and short second- and higher-order lateral roots, with an incomplete second state of endodermal development, which might be classified as peanut ‘feeder roots’. Formation of root nodules at the base of the lateral roots was the result of proliferating cell divisions derived originally from the pericycle.Key words: Endodermis, lateral root structure, nodule structure, peanut, Arachis hypogaea, primary root structure     

Microtubules and the flagellar apparatus in zoospores and cysts of the fungusPhytophthora cinnamomi

Summary A correlated immunofluorescence and ultrastructural study of the microtubular cytoskeleton has been made in zoospores and young cysts ofPhytophthora cinnamomi. Labelling of microtubules using antibodies directed towards tubulin has revealed new details of the arrangement of the flagellar rootlets in these cells, and of the variability that occurs from cell to cell. Most of the variation exists at the distal ends of the rootlets, and may be correlated with differences in cell shape in these regions. The rootlets have the same right and left configuration in all zoospores. The arrangement of the rootlet microtubules at the anterior end of the zoospores raises the possibility that the microtubules on the left hand side of the groove may not comprise an independent rootlet which arises at the basal bodies.The absolute configuration of the flagellar apparatus has been determined from ultrastructural observations of serial sections. In the vicinity of the basal bodies, there is little, if any, variation between individuals, and the structure of the flagellar apparatus is similar to that described for related species of fungi. Two ribbon-like coils surround the central pair of microtubules at the distal tip of the whiplash flagellum, and clusters of intramembranous particles, similar to ciliary plaques, have been found at the bases of both flagella. There are two arrays of microtubules associated with the nucleus in the zoospores. One array lies next to the outer surface of the nuclear envelope, and probably functions in the shaping and positioning of the apex of the nucleus. The nuclear pores in this region are aligned in rows alongside these microtubules. The second array is formed by kinetochore microtubules which extend into a collar-like arrangement of chromatin material around the narrow end of the (interphase) nucleus. During encystment, all flagellar rootlets are internalized when the flagella are detached at the terminal plate. The rootlets arrays are no longer recognizable 5–10 minutes after the commencement of encystment.     

The seminal root primordia in barley and the participation of their non-meristematic cells in root construction

In addition to the primary seminal primordium, the so-called secondary seminal root primordia are also initiated in a barley embryo. The primary root primordium is developmentally most advanced. It is formed by root meristem covered with the root cap, and by a histologically determined region with completed cell division. On germination, the restoration of growth processes begins in this non-meristematic region of root primordium by cell elongation, with the exception of the zone adjacent to the scutellar node, the cells of which do not elongate but continue differentiating. In the root primordia initiated later, the zone with completed cell division is relatively shorter, in the youngest primordia the non-meristematic cells may be lacking. The root meristem is reactivated after the primary root primordium has broken through the sheath-like coleorrhiza and emerges from the caryopsis as the primary root. The character of root meristem indicates a reduced water content at the embryonic development of root primordium. With progressing growth the root apex becomes thinner, the meristematic region becomes longer, and the differences in the extent of cell division between individual cell types increase. — The primary root base is formed of cells pre-existing in the seminal root primordium. Upon desiccation of caryopsis in maturation, and subsequent quiescent period, their development was temporarily broken, proceeding with the onset of germination. The length of this postembryonically non-dividing basal zone is different in individual cell types. The column of central metaxylem characteristic of the smallest number of cell cycles, has, under the given conditions, a mean length of about 22 mm, whereas the pericycle, as the tissue with most prolonged cell division, has a mean length of about 6 mm. In the seminal root primordia initiated later the non-dividing areas are relatively shorter. The basal region of seminal roots thus differs in its ontogenesis from the increase which is formed “de novo” by the action of root meristem upon seed germination.