Microtubular Roots Research Articles

The fine structure of the male gamete of Ectocarpus siliculosus (Ectocarpales, Phaeophyceae). II. The flagellar apparatus

The fine structure of the flagellar apparatus of male gametes of Ectocarpus siliculosus has been studied in negatively stained whole-mount preparations of isolated flagellar apparatuses and by transmission electron microscopy of embedded whole cells and flagellar apparatuses. Reconstructions from serial sections revealed a single absolute configuration and new structural details of the flagellar apparatus. A set of 5 microtubular roots is associated with the 2 flagellar basal bodies: the major anterior root consisting of 9 microtubules originates at the anterior basal body and loops through the anterior part of the cell. Cytoplasmic microtubules originate from this root at the apex of the cell and splay out in a posterior direction through the cell, thus determining cell shape. A broad microtubular band composed of a 9-membered bypassing root and a 3-membered major posterior root runs across the basal bodies. Two additional flagellar roots, the minor anterior and minor posterior rootlets, consist of a single microtubule each. The proximal end of the posterior basal body is laterally connected to the anterior basal body by a cross-striated connective (deltoid band). The basal bodies are connected to the nucleus by a small rhizoplast, and several other fibrous structures are associated with the basal bodies. The results are discussed in relation to earlier observations on flagellar apparatus organization in brown algae.

The flagellar apparatus ultrastructure in Leptosira erumpens (Deason & Bold) lukesová and its contribution to the understanding of phylogenese relationships within the microthamniales (chlorophyta)

Summary The actual spatial configuration of the flagellar apparatus in biflagellate zoospores of the terrestrial green alga Leptosira erumpens is described. The basal bodies are displaced with respect to each other in the counter clockwise direction (CCW). The two types of microtubular roots consist of 2 microtubules (R2) and 8 microtubules (R8) respectively. The latter type of microtubular root is arranged in a 7-over-1 orientation. The two basal bodies are connected to each other anteriorly by a large striated distal connecting fibre. The microtubular roots are connected to the adjacent basal body via a striated proximal fibre and linked with the lower side of the distal connecting fibre. A single striated rhizoplast runs from one basal body along a large mitochondrion towards the nuclear envelope. The distinct eyespot (stigma) is intimately associated with a series of ca. 10 microtubules originating from one of the eight-membered microtubular roots. In order to investigate the role that non-molecular characters play in the understanding of evolutionary relationships within the Microthamniales, a datamatrix was compiled for a preliminary phylogenetic analysis. Along with the present data of L. erumpens non-molecular (in great majority morphological) data of 15 more microthamnialean species and 2 selected outgroup species were included within this set. Despite incompleteness of the data, a distinct phylogenetic signal was found. This encouraged us to compare our results with a recent phylogenetic reconstruction based on molecular data.

Ultrastructure of Trimastix convexa hollande, an amitochondriate anaerobic flagellate with a previously undescribed organization

Summary Trimastix convexa is a free-living flagellate with four flagella and found in anaerobic habitats. The light microscopical appearance resembles that of Percolomonas and Tetramitus and some retortamonads, but the flagellate is shown to have an ultrastructural identity that is distinct from that of other quadriflagellate protists such as the Heterolobosea, retortamonads, diplomonads, oxymonads and trichomonads. The cell has a ventral groove bordered on the left by a microtubular root associated with a striated fibre, and on the right by a microtubular root. There are no other major non-microtubular roots or microtubular roots except a poorly developed microtubular dorsal system. The groove contains a recurrent modified flagellum with a vane reminiscent of retortamonads. The cell contains a Golgi apparatus and has hydrogenosome-like organelles but no mitochondria. On the basis of this information, we are unable to assign this species to any of the genera in which it has been previously described, and consequently classify it as Protista incertae sedis.

Ultrastructure and taxonomy of the genus Endophyton (Ulvales, Ulvophyceae)

Fine structure of vegetative cells, development and morphology of sporangia and general morphology of zoospores were studied for the first time in the genus Endophyton Gardner. Vegetative cells contained one parietal perforate chloroplast with 2–4 ulvophycean pyrenoids and transverse cell walls without plasmodesmata. Sporangial walls were formed by two distinct electron-dense and fuzzy layers which contained a contiguous electron-translucent, fibrillar material. Zoospores were naked, with an anterior dense aggregation of vesicles and a cup-shaped chloroplast with one eyespot. The flagellar apparatus showed 180° rotational symmetry and counterclockwise absolute orientation of its components. Microtubular roots had a cruciate pattern, in which d = 2 and s = 4 with 3/1 arrangement. Electron-dense bilobed terminal caps covered the proximal end of each basal body. These observations support the suggested close relationship of Endophyton to the allied genus Entocladia and other ulvalean algae, and reinforce the criteria for including the genus in the class Ulvophyceae, order Ulvales, family Ulvellaceae.

Ultrastructure and taxonomy of the genus Endophyton (Ulvales, Ulvophyceae)

Fine structure of vegetative cells, development and morphology of sporangia and general morphology of zoospores were studied for the first time in the genus Endophyton Gardner. Vegetative cells contained one parietal perforate chloroplast with 2–4 ulvophycean pyrenoids and transverse cell walls without plasmodesmata. Sporangial walls were formed by two distinct electron-dense and fuzzy layers which contained a contiguous electron-translucent, fibrillar material. Zoospores were naked, with an anterior dense aggregation of vesicles and a cup-shaped chloroplast with one eyespot. The flagellar apparatus showed 180° rotational symmetry and counterclockwise absolute orientation of its components. Microtubular roots had a cruciate pattern, in which d=2 and s=4 with 3/1 arrangement. Electron-dense bilobed terminal caps covered the proximal end of each basal body. These observations support the suggested close relationship of Endophyton to the allied genus Entocladia and other ulvalean algae, and reinforce the criteria for including the genus in the class Ulvophyceae, order Ulvales, family Ulvellaceae.

THE CYTOSKELETON OF THE NAKED GREEN FLAGELLATE SPERMATOZOPSIS SIMILIS (CHLOROPHYTA):FLAGELLAR AND BASAL BODY DEVELOPMENTAL CYCLE1

ABSTRACTFlagellar and basal body development during cell division was studied in the biflagellate green alga Spermatozopsis similis Preisig et Melkonian by light microscopy of immobilized living cells, statistical analysis of flagellar lengths during the cell cycle, and electron microscopy of cells and isolated cytoskeletons. Interphase cells display two flagella of unequal/subequal length. An eyespot located in an anterior lobe of the chloroplast is connected to the basal body bearing the shorter flagellum by means of a five‐stranded microtubular root. Until cell division, the two parental flagella attain the same length. During cell division, each cell forms two new flagella that grow to a length of 1.5 μm before they are distributed in a semiconservative fashion together with the parental flagella to the two progeny cells at cytokinesis. During the following interphase, the flagella newly formed during the preceding cell division grow to attain the same length as the parental flagella until the subsequent cell division. The shorter of the two flagella of a cell thus represents the developmentally younger flagellum, which transforms to the mature state during two consecutive cell cycles. Interphase cells display only two flagella‐bearing basal bodies; two nascent basal bodies are formed during cell division and are connected to the microtubular d‐roots of respective parental basal bodies with which the newly formed basal bodies are later distributed to the progeny cells. During segregation, basal body pairs shaft into the 11/5 o'clock direction, thus conserving the 1/7 o'clock configuration of basal body pairs of interphase cells. Prior to chloroplast and cell division, an eyespot is newly formed near the cell posterior in close association with a 1s microtubular root, while the parental eyespot is retained. During basal body segregation, eyespot‐root connections for both the old and newly formed eyespots are presumably lost, and new associations of the eyespots with the 2s roots of the newly formed basal bodies are established during cytokinesis. The significance of this “eyespot‐flagellar root developmental cycle” for the absolute orientation of the progeny cells is discussed.

Light and electron microscopical observations on Protoceratium reficulatum (Dinophyceae)

Summary Light and electron microscopical observations have been made on the gonyaulacoid dinoflagellate Protoceratium reficulatum ( Claparede et Lachmann ) Butschli collected close to the type locality of Protoceratium aceros Bergh , the type species of Protoceratium. Analysis of plate tabulation shows agreement with Gonyaulax grindleyi REINECKE and supports conspecificity between P. reticulatum, P. aceros and G. grindleyi, the first name having priority. The ultrastructure includes a branched chloroplast and a centrally located pyrenoid complex. The pusular system consists of one or two sac pusules associated with the flagellar canals. The three-dimensional arrangement of the flagellar apparatus is basically similar to that of other dinoflagellates except for the presence of a single microtubular root associated with the longitudinal basal body. Each flagellar canal is surrounded by a distinct striated fibrous collar. The collars are interconnected by two striated connectives. The plate tabulation and fine structure of Protoceratium reficulatum indicate close relationship to the genus Gonyaulax, with minor differences in plate tabulation and fine structure. It is suggested to maintain the genus Protoceratium until the fine structural diversity with Gonyaulax is better known.

The ultrastructure and systematic position of the euglenozoon Postgaardi mariagerensis, Fenchel et al.

Summary The ultrastructure of Postgaardi mariagerensis is reported for the first time. This biflagellated protist, which is covered by closely and regularly arranged rod-shaped bacteria, was obtained from the sulphidic zone of Burton Lake, Antarctica. There is a large nucleus containing apparently permanently condensed chromatin. The flagella have paraxial rods and emerge from a deep, anteriorly opening pocket. The basal bodies are associated with three asymmetrically arranged microtubular roots. Near the opening of the flagella pocket there is an MTR structure that is continuous with the microtubules of the feeding apparatus. The cell membrane is supported by a corset of evenly spaced and linked parallel microtubules subtended by an electron-dense layer. The cell has batteries of tubular, thick-walled extrusomes with a cruciform internal structure. These ultrastructural features indicate that Postgaardi mariagerensis cannot be regarded as a euglenid (as had been suggested previously) but that it is a member of the Glade Euglenozoa. On current evidence, Posfgaardi cannot be placed in any established subgroup of the Euglenozoa, nor can any subgroup be nominated as a sister taxon to Postgaardi . We therefore suggest that Postgaardi be regarded as Euglenozoa incertae sedis.

THE CYTOSKELETON OF THE NAKED GREEN FLAGELLATE SPERMATOZOPSZS SIMILIS (CHLOROPHYTA): ANALYSIS OF ISOLATED BASAL APPARATUSES IN THE PARALLEL CONFIGURATION1,2

ABSTRACTThe biflagellate green alga Spermatozopsis similis is demonstrated to be a model organism for the biochemical and functional analysis of the basal apparatus. Basal apparatuses were isolated in the presence of 10−6 M Ca2+, which induces the reorientation of the basal bodies into the parallel state. Serial thin sectioning of enriched basal apparatuses stained with tannic acid reveals several novel details of the structure of the basal bodies, the distal connecting fiber, and the striated microtubule‐associated fibers. We observed a pronounced difference in size of a striated fiber connecting the basal bodies to the five‐stranded microtubular roots depending on its association with the developmentally older or younger basal body. Instead of a proximal connecting fiber, the proximal end of each basal body is associated with a striated triangular plate; these plates appear to serve as spacers for the basal bodies in the parallel and antiparallel configurations. We suggest that the plates play a role in maintaining basal body orientation during forward and backward swimming. The results are summarized in representative drawings of the basal apparatus.

SF-assemblin in Chlamydomonas: sequence conservation and localization during the cell cycle.

Previously, SF-assemblin has been identified as the filament-forming component of the striated microtubule-associated fibers (SMAFs), which emerge from the basal bodies in several green flagellates. We have sequenced cDNAs coding for SF-assemblin from Chlalmydomonas reinhardtii and C. eugametos. Comparison of the deduced amino acid sequences with the previously described green algal SF-assemblins shows identities between 54 and 71%, indicating a strong drift in sequence. Cells of C. reinhardtii were analyzed by double immunofluorescence using polyclonal anti-SF-assemblin and anti-alpha-tubulin. In interphase cells, SF-assemblin is associated with all four microtubular flagellar roots. During mitosis the SF-assemblin-based cytoskeleton is reorganized; it divides in prophase and is reduced to two dot-like structures at each spindle pole in metaphase. During anaphase, the two dots present at each pole are connected again. In telophase we observed an asymmetrical outgrowth of new fibers. These observations suggest a role for SF-assemblin in reestablishing the microtubular root system characteristic of interphase cells after mitosis.

The organisation of Mastigamoeba schizophrenia n. sp.: More evidence of ultrastructural idiosyncrasy and simplicity in pelobiont protists

Summary A new species of pelobiont, Mastigamoeba schizophrenia n. sp., is described using light and electron microscopy. Three forms were observed: flagellates, aflagellated amoebae and cysts. This species is distinguished by having adhering paired nuclei. Most cells have a single pair of nuclei but large amoebae may have several pairs. The flagellar apparatus consists of a single basal body, connected to the most anterior nucleus by a cone of microtubules. The basal body is composed of doublets of microtubules. A single ribbonlike microtubular root originates alongside the basal body. Like other pelobionts, the cell has a comparatively simple endomembrane system which includes endoplasmic reticulum but lacks mitochondria, dictyosomes, contractile vacuole and other elements found in many eukaryotic cells. The taxonomy of the pelobionts is discussed, the genus Phreatamoeba is synonymised with Mastigamoeba, and arguments for a deep-branching position for the pelobionts among eukaryotes are discussed.

The flagellar apparatus of Cafeteria roenbergensis Fenchel & Patterson, 1988 (Bicosoecales = Bicosoecida)

Summary The flagellar apparatus of Cafeteria roenbergensis consists of two basal bodies, three microtubular roots, of which one bears secondary cytoskeletal microtubules, and a forked rhizoplast that connects the elder, posteriorly directed basal body to the nucleus. Analysis of the three-dimensional architecture of this flagellar apparatus confirms that Cafeteria belongs to the bicosoecids and suggests that there is little variation in flagellar apparatus features among bicosoecids. Feeding apparatus architecture (the “Type I feeding basket”) in bicosoecids differs from that in chromulinalean chrysophytes in several features. Nevertheless, the structures in bicosoecids and chromulinalean chrysophytes are more similar to each other than to any other stramenopiles and may share a similar mechanism based on microtubule-microtubule sliding. The feeding basket may be an ancestral motif among stramenopiles, conserved in different forms in bicosoecids and chromulinalean chrysophytes but lost from other lineages.

Phylogeny of Diplonema ambulator (Larsen and Patterson) : 2. Homologies of the feeding apparatus

Summary The ultrastructure of the flagellar apparatus of Diplonema ambulator was determined to investigate the phylogenetic placement of this genus. Two flagella insert into a pocket and bear distal and proximal transitional plates. The basal bodies are linked by a connecting fiber and are associated with three microtubular roots. A three-membered dorsal root attaches to the dorsal basal body and ascends along the flagellar pocket. An intermediate root is composed of five microtubules where it attaches to the ventral basal body (Vb). As it ascends along the flagellar pocket, two additional microtubules become associated, producing a 6 + 1 configuration. A four-membered ventral root attaches laterally to the Vb and extends into the cytoplasm. The flagellar pocket is lined on one side by an independent band of microtubules which are reminiscent of the bodonid MTR. The presence of three asymmetrically arranged microtubular roots, transitional plates similar to those in the flagella of kinetoplastids, and an MTR-like band further support the relationship of Diplonema with the Euglenozoa. However, the absence of paraxonemal rods and flagellar hairs as well as the unique mitotic chromosome behavior and peculiar mitochondrial cristae structure suggest that Diplonema may have undergone substantial independent evolution since its point of divergence.

GENERAL ULTRASTRUCTURE AND FLAGELLAR APPARATUS ARCHITECTURE OF WOLOSZYNSKIA LIMNETICA (DINOPHYCEAE)

ABSTRACT The ultrastructure of Woloszynskia limnetica Bursa was examined using serial thin section electron microscopy. Sections of W. limnetica reveal numerous chloroplast profiles without any obvious pyrenoids. The extensive pusular complex consists of a “smooth” part and a part lined with electron‐dense particles. The nucleus is located in the episome. A stigma (=eyespot) consisting of numerous electron‐dense globules is situated beneath the amphiesmal vesicles of the sulcal groove. The longitudinal microtu‐bular root extends between the stigma and the amphiesma vesicles. Subthecal fibers occur in conjunction with the microtubules and the stigma. Both flagellar exit apertures are encircled by a broad striated collar, each giving rise to a fiber that extends along the pusular canal opening. The striated collars are interconnected by the ventral ridge fiber. The basal part of the transverse flagellum has, in addition to the normal paraxonemal rod (=striated strand or fiber), a semicircular structure consisting of fibrils. The flagellar apparatus is complex but possesses components typically found in the Dinophyceae. The longitudinal mi‐crotubular root is broad and is connected to both striated collars. The transverse basal body gives rise to the transverse microtubular root, which in turn is associated with microtubules that extend to the interior of the cell and with the transverse striated root. The transitional region of both basal bodies possesses a distinctive fibrous ring attached to each microtubular triplet by short fibers that collectively appear as spokes of a wheel. Not unexpectedly, the flagellar apparatus of Woloszynskia limnetica is much like that of the related Woloszynskia sp.; however, some dif ferences were discovered. A phylogenetic relationship between Woloszynskia limnetica, W. coronata (Wolosz.) Thompson, and W. sp. is indicated based on similarities in pusule and stigma structure.

A Deviating Pattern of Cell Division in the Green Alga Microthamnion: Ultrastructure of Vegetative Cell Division and Zoosporogenesis

Summary The ultrastructure of cell division of the green alga Microthamnion is described. It involves a unique mechanism for both vegetative cell division and zoosporogenesis. It starts with migration of the counterclockwise oriented centriole pair along the nuclear surface from the polar interphase position towards a lateral prophase position. The centriole pair duplicates and the root templates of the parental basal bodies grow out distally. Two microtubules of each of the two microtubular roots per basal body form two four-stranded microtubular bundles running along the cell wall and oriented perpendicular to the cell axis. At metaphase the separated centriole pairs have migrated to positions at approximately 180° from each other apparently via a sliding mechanism along the newly formed microtubular bundles. The microtubular bundles lie in the equatorial plane and curve over the nucleus, thus indicating the future division plane. In this respect the microtubular bundles resemble the preprophase band in higher plants. At anaphase the centriole pairs seem to have slightly co-migrated with the separating chromosome halves and thus the microtubular bundles show a shifted orientation with respect to the long axis of the cell. In this stage the unilateral septum starts to develop from the previous prophase position of the centriolar complex and progresses between the two four-stranded microtubular bundles. At telophase the daughter nuclei are reformed and the septum development proceeds. After cytokinesis the centrioles either return to the interphase position awaiting a new division cycle (vegetative cell division) or the centrioles take the prophase position in the newly formed daughter cells and duplicate in order to repeat the division cycle (zoosporogenesis). The ultrastructural features of Microthamnion are compared with the other pleurastrophycean algae and some chlamydomonadalean algae. However, the systematic position of Microthamnion still remains a matter of discussion.

Ultrastructure of the flagellar apparatus of Noctiluca miliaris Suriray swarmers (Dinophyceae)

The swarmers of Noctiluca miliaris Suriray are not uniflagellate, as was assumed for more than a century, but are biflagellate cells. Their flagellar apparatus displays two distinctly heteromorphic flagella: a long flagellum extending posteriorly (=longitudinal flagellum) and a very short flagellum oriented to the cell's left side (= transverse flagellum). The transverse flagellum is situated in a groove and is therefore not visible in the light microscope. A serial section analysis of the basal apparatus of the swarmers reveals structures which are typical for dinoflagellates including a posteriorly directed micro-tubular root (pmr) and a ventral ridge with microtubular and fibrous components (vrm and vrf). Striated fibrous roots are, however, lacking. The structurally relatively simple flagellar apparatus of N. miliaris swarmers may reflect their functional specialization as (micro-)gametes.

Centrin association with the flagellar apparatus in spores ofPhytophthora cinnamomi

Antibodies raised against the calcium-binding protein centrin, were used to identify and localise centrin containing structures in the flagellar apparatus of zoospores and cysts of the oomycetePhytophthora cinnamomi. Immunoblotting of extracts from zoospores indicates that theP. cinnamomi centrin homologue is a 20 kDa protein. Immunofluorescence microscopy with anti-centrin antibodies reveals labelling in the flagella, the basal body connector and co-localisation along the microtubular R1 root (formerly called AR3) that runs from the right side of the basal body of the anterior flagellum into the anterior of the zoospore close to the ventral surface. The centrin (R1cen) and tubulin components of the R1 root split into four loops on the right hand side of the ventral groove and rejoin along the left hand side of the groove. The R1 root continues down the left hand side of the zoospore past the basal bodies and parallel to the R4 root. We propose that at least inP. cinnamomi there is no R2 root. Immunogold labelling confirms that centrin is a component of the basal body connector complex. When the zoospores become spherical during encystment, the R1cen pivots by approximately 90 ° with respect to the nucleus.

Fine structural studies on an ultraplanktonic species ofPyramimonas, P. virginica(Prasinophyceae), with a discussion of subgenera within the genusPyramimonas

The ultrastructure of Pyramimonas virginica, the only known ultraplanktonic species of Pyramimonas, has been examined in detail. This species differs from other species of Pyramimonas in the structure of the flagellar apparatus, the scaly periplast, the pyrenoid, the eyespot, location of the eyespot in the cell, and mitosis. The flagellar apparatus is simple and shows signs of reduction. A 2-2-2-2 microtubular root system is present but a rhizoplast was not found. A very short transitional helix (perhaps two rings) is present in the flagellar transition region, together with a short one-piece stellate structure. Numbering of individual flagella in different species of Pyramimonas has shown a surprising variation in the orientation of the flagellar apparatus relative to the rest of the cell. Thus the eyespot in the various species is located differently relative to the flagellar apparatus and to other organelles such as the nucleus (in Pyramimonas there is no connection between the flagellar apparatus and ...

Flagellar apparatus transformation and development in Prorocentrum micans and P. minimum (Dinophyceae)

Flagellar transformation was studied in Prorocentrum minimum (Pavillard) Schiller on free-swimming cells using low-illumination light microscopy. Cells bear two ventrally inserted, heterodynamic flagella. Prior to cell division, two new transverse flagella are produced. The parental longitudinal flagellum (1) remains unaltered during cell division, while the parental transverse flagellum (2) is either retracted or abscissed, and regrows as a longitudinal flagellum (1(2)). The basal apparatuses of both P. micans Ehrenberg and P. minimum show the typical complement of dinophycean roots at interphase. Two new daughter basal bodies (2(1) and 2(2)) are formed prior to any alteration of the parental basal apparatus. The parental longitudinal microtubular root (LMR) and basal body connective (BBC) appear to be maintained unaltered and associated with the parental longitudinal basal body (1). A new longitudinal microtubular root (Imr), basal body connective (bbc) and fibrous connectives (fc's) are produced between the transforming transverse basal body (2) and its daughter (2(2)). New striated root connectives (src's), transverse striated roots (tsr's) and their associated microtubules (tsrm's) are formed next to the new, transverse daughter basal bodies (2(1) and 2(2)). Basal body 2(1) forms its own transverse microtubular root (tmr) while the same root on basal body 2(2) might be derived from the parental transverse striated root microtubule (TSRM) as a result of root transformation and transfer. The parental transverse striated root (TSR) and transverse microtubular root (TMR) are disassembled during cell division. The pattern of flagellar root development is compared to other known root developmental processes for which we introduce a system of classification. We also introduce a flagellar root nomenclature similar to the flagellar nomenclature to indicate unaltered parental roots, newly formed roots, transferred roots and transformed parental roots that constitute another root type in the new daughter cell.

The flagellar apparatus and canal structure in Prorocentrum micans (Dinophyceae)

We have determined the three-dimensional structure of the flagellar apparatus in Prorocentrum micans Ehrenberg (Dinophyceae). The flagellar apparatus consists of two basal bodies that lie at approximately a 35° angle to one another. Each basal body bears a typical dinophycean complement of microtubular and fibrous components. Unlike the majority of Dinophyceae, the longitudinal microtubular root lies adjacent to the longitudinal basal body and abuts the transverse basal body. The transverse microtubular root is associated with an extension of microtubules that is directed posteriorly (dorsal, using proposed terminology) into the cell to a region adjacent to the nucleus. On the opposite side of the flagellar apparatus, the microtubules of the peduncle extend posteriorly (dorsal, using proposed terminology) from the flagellar canal region. The microtubules extend for several micrometers and the peduncle is closely associated with a complex striated collar that circumscribes the flagellar canal and its components. We report on the three-dimensional nature of the flagellar canal and accessory pore canal region. We conclude that the flagellar canal and accessory pore canal are not attached to each other and that the ‘Prorocentrum’ type of flagellar apparatus and cell architecture is most likely the result of episome/hyposome compression.