The oral apparatus of Tetrahymena pyriformis, strain WH-6. IV. Observations on the organization of microtubules and filaments in the isolated oral apparatus and the differential effect of potassium chloride on the stability of oral apparatus microtubules.

This report is an ultrastructural analysis of the organization of the isolated oral apparatus of Tetrahymena pyriformis, strain WH-6, syngen 1. Attention has been focused on the organization of microtubules and filaments in oral apparatus membranelles. Oral apparatus membranellar basal bodies were characterized with respect to structural differentiations at the distal and proximal ends. The distal region of membranellar basal bodies contains the basal plate, accessory microtubules and filaments. The proximal end contains a dense material from which emanate accessory microtubules and filaments. There are at least two possibly three different arrangements of accessory structures at the proximal end of membranellar basal bodies. All membranellar basal bodies appear to have a dense material at the proximal end from which filaments emanate. Some of these basal bodies have accessory microtubules and filaments emanating from this dense material. A possible third arrangement is represented by basal bodies which have lateral projections, from the proximal end, of accessory microtubules and filaments which constitute cross or peripheral connectives. There are at least three examples of direct associations between oral apparatus microtubules and filaments: (1) filaments which form links between basal body triplet microtubules, (2) filaments which link the material of the basal plate to internal basal body microtubules, (3) filaments which link together microtubule bundles from membranellar connectives. KCl extraction of the isolated oral apparatus resulted in the selective solubilization of oral apparatus basal bodies, remnants of ciliary axonemes and fused basal plates. Based on their response to KCl extraction two distinct sets of morphologically similar micro tubules can be identified: (a) microtubules which constitute the internal structure of basal bodies and ciliary axonemes, (b) microtubules which constitute the fiber connectives between basal bodies.     

Assembly and fate of basal bodies in the colourless phytoflagellate Polytoma papillatum

Summary— The morphogenesis of basal bodies is described in the phytoflagellate Polytoma papillatum. The observations are based on the analysis of ultrathin serial sections through the flagellar apparatus of interphase, mitotic, and postmitotic cells using transmission electron microscopy. Formation of new basal bodies starts in prometaphase. Individual A-subfibres develop orthogonally to the long axis of mature basal bodies. The microtubules assemble at the surface of an annulus of amorphous material. By telophase, a complete cylinder of A-subfibres with a length of approximately 300 nm has formed. Although the proximal ends of these new probasal bodies are detached from the mature basal bodies, prominent reorientation of the probasal bodies does not occur. They remain with their proximal ends in the vicinity of mature basal bodies. In daughter cells with probasal bodies around 400 nm long, the assembly of microtubular triplets is initiated. B- and C-subfibres first show up distal from the mature basal bodies and may elongate towards them. Thus, A-subfibres on the one side and B- and C-subfibres on the other appear to growt with opposite polarity. If A-subfibres grow at their plus ends, B- and C-subfibres elongate at their minus ends. The latter is unusual in comparison with individual cytoplasmic and spindle microtubules. Possible the presence of a lateral template in the form of the A-subfibres is responsible for the deviating growth characteristics of the incomplete B- and C-subfibres. In interphase cells, the mature basal bodies extend into long flagella. The new basal bodies remain devoid of flagella and are less than 85 nm long. Thus, they have shortened relative to their precursors in mitotic and postmitotic cells. At the onset of a new division cycle, the flagellate basal badies shed their flagella. The breaking point is at the triplet-doublet transition of the flagellum.     

THE MORPHOGENESIS OF BASAL BODIES AND ACCESSORY STRUCTURES OF THE CORTEX OF THE CILIATED PROTOZOAN TETRAHYMENA PYRIFORMIS

Dividing cells of Tetrahymena pyriformis were observed by transmission electron microscopy for signs of morphogenesis of cortical structures. The earliest stage of basal body development observed was of a short cylinder of nine single tubules connected by an internal cartwheel structure. This is set perpendicular to the mature basal body at its anterior proximal surface under the transverse microtubules and next to the basal microtubules. Sequential stages show that the single tubules become triplet tubules and that the "probasal bodies" then elongate and tilt toward the organism's surface while maintaining a constant distance of 75–100 mµ with the "parent." The new basal body after it is fully extended contacts the pellicle, and then assumes a parallel orientation with and moves anterior to the parent basal body. The electron-opaque core in the lumen of the basal body and accessory structures around its outer proximal surface appear after the developing basal body has elongated. These accessory structures associating with their counterparts from other basal bodies and with the longitudinal microtubules may play a role in the final positioning of basal bodies and thus in the maintenance of cortical patterns. Observations on a second sequence of basal body formation suggest that the oral anlage arises by multiple duplication of somatic basal bodies.     

ULTRASTRUCTURAL STUDIES OF SPERMATOGENESIS IN THE ANTHOCEROTALES. I. THE BLEPHAROPLAST AND ANTERIOR MITOCHONDRION IN PHAEOCEROS LAEVIS: EARLY DEVELOPMENT

Electron microscopic examination of thin sections showed that the blepharoplast of a young spermatid of Phaeoceros consists of two side-by-side centrioles and an accumulation of osmiophilic, granular matrix at their proximal ends. Lying between these nearly parallel organelles is a dark-staining body that will later disappear at the onset of flagellogenesis. For a brief period the centrioles are oriented perpendicular to the nuclear surface so that the granular matrix at their proximal ends is confluent with the nuclear envelope; furthermore, the nucleoplasm immediately in front of the centrioles becomes densely staining. The multilayered structure (MLS) develops directly under the centrioles. It comprises a band of 12 microtubules (the S₁ stratum) and three lower strata (S_2–4) whose constitutent lamellae are oriented at an oblique angle to the S₁ axis. While the S₁ tubules grow rearward over the nucleus which forms a beak adjacent to the posterior end of the lamellar strata, the centrioles are transformed into basal bodies with the distal growth of the axonemes and the proximal growth of the central cartwheels and lowermost triplets. The proximal ends of the basal bodies and the S₁ tubules overlying the lamellar strata are invested with osmiophilic matrix that extends down to the S₂ layer and may temporarily occlude the lamellar plates. At the onset of nuclear elongation an anterior mitochondrion becomes situated close beneath the lamellar strata which extend laterally beyond the S₁ tubules.     

Stress fibers in situ in proximal tubules of the rat kidney

Actin bundles in proximal tubules of the rat kidney were examined by immunofluorescence and confocal laser microscopy with special reference to their three-dimensional distribution and identification as stress fibers. Renal tubular segments were prepared from the fresh renal cortex by simple homogenization and centrifugation, and fixed in formaldehyde for staining with fluorescent dye-labeled phalloidin. Segments of the proximal tubules could be identified easily on the bases of their diameter, the height of epithelial cells and prominent brush borders. Confocal laser microscopy clearly demonstrated the overall distribution of actin bundles in the whole-mount proximal tubular segments. Actin bundles in the basal cytoplasm of epithelial cells were observed to run parallel to each other and at a right angle to the tubular axis. In the stereo views reconstructed from serial optical sections, the basal actin bundles appeared as straight rods with both ends tapered. They varied in length and width and extended rather short distances of not more than 10 microns. Often, two or more actin bundles were longitudinally aligned in tandem. Some bundles showed irregular bandings along their length. Each bundle was composed of tightly packed actin filaments which could be decorated with heavy meromyosin subfragment-1 to display a bi-directional arrangement within the bundle. Immunostaining of cryostat sections showed that actin bundles contained myosin and vinculin. Enzymatically isolated proximal tubules contracted upon addition of Mg-ATP. These observations collectively suggest that the actin bundles at the base of renal proximal tubule epithelial cells can be listed among the examples of stress fibers in situ.     

The Fine Structure of the Paralabial Organelle in the Rumen Ciliate Ophryoscolex purkinjei Stein, 1858

The paralabial organelle of the rumen ciliate Ophryoscolex purkinjei , located on the ventral side of the ciliophor, is a highly specialized part of the somatic cortex. It consists of alternating rows of short modified cilia and thin pellicular folds which form a ridge-like structure. The central "top kinety" is composed of monokinetids which bear cilia with 9 + 2 axonemes and 2 μm in length. The top kinety is accompanied by a comb-shaped fold on its distal side and by a broad wedge-shaped fold on its proximal side. To both sides there follow two or three lateral kineties made of dikinetids. The anterior kinetosome of each pair bears a clavate cilium, only 0.5–0.7 μm in length and with a 9 + 0 axoneme while the cilium of the posterior kinetosome is even shorter. Lateral folds with numerous microtubules cover these lateral kineties and rows of barren basal bodies. The fine structure of this supposed sensory organelle show a basic pattern in four other ophryoscolecids, and its increasing complexity parallels the suggested phylogenetic line of evolution of these ciliates.     

ELECTRON MICROSCOPE OBSERVATIONS ON THE FLAGELLAR APPARATUS OF BRYOPSIS MAXIMA (CHLOROPHYCEAE)1

The flagellar apparatus in male gametes of the siphonaceous green alga, Bryopsis maxima Okamura, was studied and compared with that of other green biflagellate cells. The proximal portions of two basal bodies are connected by a single striated proximal band, unique among the biflagellate reproductive cells of green algae studied. Anterior to the flagellar bases is a pair of distal bands different from the single structure in other biflagellate cells. These bands which arise from the distal portion of each basal body, extend upward in the papilla and curve down toward the lower edges of the basal bodies. They seem to have no direct association with each other. Two pairs of distinct flagellar roots, one consisting of 3–5 microtubules and the other of a partially striated fiber of undetermined numbers of microtubules, diverge from the basal body region and extend towards the cell posterior. Their component microtubules are disorganized into single or smaller groups midway over the cell length. The uniqueness of the flagellar apparatus is briefly discussed.     

The flagellar apparatus of the golden algaSynura uvella: Four absolute orientations

Summary Flagellated vegetative cells of the colonial golden algaSynura uvella Ehr, were examined using serial sections. The two flagella are nearly parallel as they emerge from a flagellar pit near the apex of the cell. The photoreceptor is restricted to swellings on the flagella in the region where they pass through the apical pore in the scale case and the swellings are not associated with the cell membrane or an eyespot. A unique ring-like structure surrounds the axonemes of both flagella at a level just above the transitional helix. The basal bodies are interconnected by three striated, fibrous bands. Four short (<100 nm) microtubules lie between the basal bodies at their proximal ends. Two rhizoplasts extend down from the basal bodies and separate into numerous fine striated bands which lie over the nucleus. Three- and four-membered microtubular roots arise from the rhizoplasts and extend apically together. As the roots reach the cell anterior, the three-membered root bends and curves clockwise to form a large loop around the flagella; the four-membered root bends anticlockwise and terminates under the distal end of the three-membered root as it completes the loop. There are four absolute orientations, termed Types 1–4, in which the flagellar apparatus can occur. With each orientation type the positions of the Golgi body, nucleus, rhizoplasts, chloroplasts and microtubular roots change with respect to the flagella, basal bodies and photoreceptor. Two new basal bodies appear in pre-division cells, and three short microtubules appear in a dense substance adjacent to each new basal body. Based upon the positions of new pre-division basal bodies, a hypothesis is proposed to explain why there are four orientations and how they are maintained through successive cell divisions.     

Prey finding behavior and mechanosensilla of larval Toxorhynchites brevipalpis Theobald (Diptera : Culicidae)

Behavioral experiments demonstrated that starved 3rd-instar Toxorhynchites brevipalpis (Diptera : Culicidae) will attack a glass probe in response to vibrations alone. Frequencies in the range of 80–200 Hz elicited 85% of the attacks. The observation that most attacks occurred while the larva was drifting forward towards the probe, substantiates that the predatory behavior of these organisms is basically opportunistic.The main setae on the thorax and abdomen arise from setal support plates and are of 4 general morphological types: dendritic, smooth, spinulate spiniform, and unbranched aciculate setae. The numbers and lengths of the setae on 3rd- and 4th-instar larvae are given. Each seta is innervated by a bipolar neuron with a tubular body in the dendrite, a characteristic of mechanosensilla. By virtue of their abundance and location, it seems probable that one or more types of these setae sense the water vibrations capable of eliciting an attack response.The tubular bodies of the 4 types of setae are somewhat unusual in that they lack electron-dense material, except near sites of attachment of the microtubules to the dendritic membrane, yet possess a large number of complexly arranged microtubules. Among the setal types, variations were observed in the prominence of microtubular attachment sites, proportion of tubules associated with the sites, and orientation of microtubules. In spinulate spiniform setae, the arrangement of microtubules varies within the same tubular body; peripherally most of the microtubules are in one direction with little convergence and have only one distinct attachment site, whereas medially they converge considerably resulting in 2 groups, each associated with its own well-defined site.The attachment sites, which presumably through linkage with the dendritic membrane provide a means of initiating depolarization, are associated with the distal ends of almost all of the microtubules. This suggests that in these tubular bodies depolarization may be initiated through mechanical force acting along the length of the microtubules, that is, stretching and/or compression.     

Membrane/microtubule tip attachment complexes (TACs) allow the assembly dynamics of plus ends to push and pull membranes into tubulovesicular networks in interphase Xenopus egg extracts

We discovered by using high resolution video microscopy, that membranes become attached selectively to the growing plus ends of microtubules by membrane/microtubule tip attachment complexes (TACs) in interphase- arrested, undiluted, Xenopus egg extracts. Persistent plus end growth of stationary microtubules pushed the membranes into thin tubules and dragged them through the cytoplasm at the approximately 20 microns/min velocity typical of free plus ends. Membrane tubules also remained attached to plus ends when they switched to the shortening phase of dynamic instability at velocities typical of free ends, 50-60 microns/min. Over time, the membrane tubules contacted and fused with one another along their lengths, forming a polygonal network much like the distribution of ER in cells. Several components of the membrane networks formed by TACs were identified as ER by immunofluorescent staining using antibodies to ER-resident proteins. TAC motility was not inhibited by known inhibitors of microtubule motor activity, including 5 mM AMP-PNP, 250 microM orthovanadate, and ATP depletion. These results show that membrane/microtubule TACs enable polymerizing ends to push and depolymerizing ends to pull membranes into thin tubular extensions and networks at fast velocities.     

Structure of the flagellar apparatus in the bacteriotrophic flagellate Parabodo nitrophilus Skuja, 1948 (Kinetoplastea Excavata)

The structure of the flagellar apparatus in the excavate flagellate Parabodo nitrophilus Skuja has been studied. Two smooth heterodynamic flagella emerge from the bottom of the flagellar apparatus. The kinetosomes connected by their proximal ends lie under an acute angle to each other and bear against the plate on the anteior wall of kinetoplast. The dorsal and ventral rootlets emerge from the kinetosomes and are transformed into dorsal and ventral bands. The latter accompanies the posterior flagellum. The MTR band begins inside the wall of the flagellar pocket. The upper part of the cytopharynx is armored by MTR and FAS bands, cross-banded fibril and structure, and additional microtubules. The MTR band and three additional microtubules surround the bottom part of cytopharynx. The mitochondrium contains compact kinetoplast and discoid cristae. The resemblance of Parabodo nitrophilus with other free-living kinetoplastids is discussed.     

Some Observations on the Structure of the Peristomial Membranelle of Spirostomum ambiguum

SYNOPSIS. Electron-microscopic observations of Spirostomum ambiguum have demonstrated additional details of superficial and deep tubular connections with peristomial and somatic kinetosomes. The superficial peristomial tubules appear to connect adjacent rows of kinetosomes. Anatomically, they course distally from the proximal kinetosomal plate. The deep tubules run proximally from the kinetosomal plate. Those in the somatic region appear to enter the endoplasm; those in the peristomial region leave the kinetosome as bundles of either 10 or 11 tubules which steadily converge to form 2 compact rows of 10 tubular bundles. These tubules connect to 2 of the 3 rows of 10 cilia each, the rows of 3 being separated by membranous folds protruding perpendicular to the peristomial groove. The rows of bundles converge further, enter the endoplasm and fan out again into tubular sheets, some of which appear to course in an antero-posterior direction. Another set of tubules arises from each of the kinetosomes in the 3rd row of 10 kinetosomes and courses proximally at a different angle from those arising from the 2 other kinetosome rows. Terminations have not been observed for the deep somatic or peristomial tubules. Their possible role in producing the forceful longitudinal contraction of Spirostomum is discussed.     

In vitro culture of the proximal tubule of the bovine nephron

By means of a previously described method, viable pure tubules of the nephron were isolated in high yield from the outer cortex of the near-term foetal bovine kidney. The tubular suspension obtained was constituted almost exclusively of proximal segments (about 95%), whose cells were dispersed and grown as confluent primary cultures. The cultured proximal cells were shown to maintain in vitro, on glass or plastic surfaces, the same orientation as on the tubular basement membrane in vivo, with interdigitations extending from the base of the cells and along their full height. Numerous mitochondria and the typical cytoplasmic bodies of the proximal cell were retained in cells grown in vitro. A flagellum was seen in every cultured cell and was shown to be present in the proximal cell in vivo. There is a progressive change, in vitro, of the microvilli of the brush border, from a close-packed to a sparse distribution and to a decrease in height and a reduction in number. This in vitro regression to an earlier embryonic state was correlated with the ability of the proximal cells to synthesize in vitro an alpha-fetoprotein and with the loss in vitro of histiospecific antigen synthesis, confined in vivo to the brush border area. The confluent proximal cells became filled with microfilaments and microtubules, the significance of which is discussed.     

Ultrastructure of the Somatic and Buccal Cortex of the Tetrahymenine Hymenostome Glaucoma chattoni

SYNOPSIS The membranes, epiplasm, and fiber systems are described in the somatic cortex of Glaucoma chattoni strain HZ-1. Kinetodesmal fibers, postciliary and transverse microtubular ribbons, basal microtubules, transverse fibers and transverse accessory material are associated with kinetosomes. Longitudinal microtubular ribbons and mitochondria occur interkinetally. In the buccal cortex, the membranes, epiplasm and fibers of the 3 membranelles, the paroral kinety, the ribbed wall, and the cytostome are described. Comparisons between G. chattoni and other ciliates reveal ultrastructural differences of possible systematic significance. In the somatic cortex of this and other tetrahymenines. Iongitudinal microtubular ribbons and basal microtubules occur concurrently. In the buccal cortex, alveoli are absent in tetrahymenine membranelles. A table is presented of the fiber systems associated with single somatic kinetosomes of various ciliates whose cortical ultrastructure has been studied to date.     

COMPARATIVE ANALYSES OF THE DINOFLAGELLATE FLAGELLAR APPARATUS. III. FREEZE SUBSTITUTION OF AMPHIDINIUM RHYNCHOCEPHALUM1

The flagellar apparatus of the marine dinoflagellate Amphidinium rhynchocephalum Anissimowa was examined using the techniques of rapid freezing/freeze substitution and serial thin section three dimensional reconstruction. The flagellar apparatus is composed of two basal bodies that are offset from one another and lie at an angle of approximately 150° The transverse basal body is associated with two individual microtubules that extend from the proximal end of the basal body toward the flagellar opening. One of these microtubules is closely appressed to a striated fibrous root that also extends from the proximal base of the transverse basal body. The longitudinal basal body is associated with a nine member microtubular root that extends from the proximal end of the basal body toward the posterior of the cell. The longitudinal microtubular root and the transverse striated fiber are connected by a striated connective fiber. In addition to the microtubules associated with the transverse and longitudinal basal bodies, a group of microtubules originates adjacent to one of the transverse flagellar roots and extends into the cytoplasm. Vesicular channels extend from the flagellar openings to the region of the basal bodies where they expand to encompass the various connective structures of the flagellar apparatus. The possible function and evolutionary importance of these structures is discussed.     

The fine structure of the paralabial organelle in the rumen ciliate Ophryoscolex purkinjei Stein, 1858

The paralabial organelle of the rumen ciliate Ophryoscolex purkinjei, located on the ventral side of the ciliophor, is a highly specialized part of the somatic cortex. It consists of alternating rows of short modified cilia and thin pellicular folds which form a ridge-like structure. The central "top kinety" is composed of monokinetids which bear cilia with 9 + 2 axonemes and 2 microns in length. The top kinety is accompanied by a comb-shaped fold on its distal side and by a broad wedge-shaped fold on its proximal side. To both sides there follow two or three lateral kineties made of dikinetids. The anterior kinetosome of each pair bears a clavate cilium, only 0.5-0.7 micron in length and with a 9 + 0 axoneme while the cilium of the posterior kinetosome is even shorter. Lateral folds with numerous microtubules cover these lateral kineties and rows of barren basal bodies. The fine structure of this supposed sensory organelle show a basic pattern in four other ophryoscolecids, and its increasing complexity parallels the suggested phylogenetic line of evolution of these ciliates.     

Transformation of the flagella and associated flagellar components during cell division in the coccolithophoridPleurochrysis carterae

Summary Immunofluorescence microscopy, conventional and high voltage transmission electron microscopy were used to describe changes in the flagellar apparatus during cell division in the motile, coccolithbearing cells ofPleurochrysis carterae (Braarud and Fagerlund) Christensen. New basal bodies appear alongside the parental basal bodies before mitosis and at prophase the large microtubular (crystalline) roots disassemble as their component microtubules migrate to the future spindle poles. By prometaphase the crystalline roots have disappeared; the flagellar axonemes shorten and the two pairs of basal bodies (each consisting of one parental and one daughter basal body) separate so that each pair is distal to a spindle pole. By late prometaphase the pairs of basal bodies bear diminutive flagellar roots for the future daughter cells. The long flagellum of each daughter cell is derived from the parental basal bodies; thus, the basal body that produces a short flagellum in the parent produces a long flagellum in the daughter cell. We conclude that each basal body in these cells is inherently identical but that a first generation basal body generates a short flagellum and in succeeding generations it produces a long flagellum. At metaphase a fibrous band connecting the basal bodies appears and the roots and basal bodies reorient to their interphase configuration. By telophase the crystalline roots have begun to reform and the rootlet microtubules have assumed their interphase appearance by early cytokinesis.Abbreviations CR1, CR2 crystalline roots 1 and 2 - CT cytoplasmic tongue microtubules - DIC differential interference contrast light microscopy - H haptonema - HVEM high voltage transmission electron microscopy - IMF immunofluorescence microscopy - L left flagellum/basal body - M metaphase plate - MT microtubule - N nucleus - R right flagellum/basal body - R1, R2, R3 roots 1, 2, and 3 - TEM transmission electron microscopy     

FINE STRUCTURE OF THE ZOOSPORE OF OEDOCLADIUM CAROLINIANUM (CHLOROPHYTA) WITH SPECIAL REFERENCE TO THE FLAGELLAR APPARATUS1, 2

Ultrastructure of the motile zoospore has been investigated in Oedocladium catolinianum & Hoffman. An unwalled zoospore is usually produced from the contents of a terminal vegetative cell and consists of two principal regions: a small anterior dome and a larger body region; a ring of flagella marks the juncture of these two areas. Chloroplast inclusions consist of thylakoids, mature and incipient pyrenoids, starch and striated microtubules; no eyespot has been observed. Zoospores appear to possess permanent contractile vacuoles with numerous accessory vacuoles, coated vesicles and occasionally coated tubules. The cytoplasm of the dome contains numerous mitochondria ER and golgi bodies, as well as two distinct types of vesicles. The first contains an electron-dense; granular core and is surrounded by a loose, sinuate membrane. The second vesicle is electron-opaque and is found at the apex of the dome: it contains mucopolysaccharides employed during zoospore adhesion. A complex flagellar apparatus encircles the lower region of the dome. It consists of ca. 30–65 flagella, a ring-shaped fibrous band, flagella roots and additional supporting material. The flagella and roots alternate with one another beneath the fibrous band. The compound flagellar roots consist of two superimposed components: an outer ribbon-like unit composed of three microtubular elements and a single striated inner component. A band of support material lies beneath the proximal end of the basal bodies. It is a continuous fibrous band, although it often appears as three distinct, repetitive units.     

Evidence for a direct role of nascent basal bodies during spindle pole initiation in the green alga Spermatozopsis similis.

Basal body replication in the naked biflagellate green alga Spermatozopsis similis was analyzed using standard electron microscopy and immunogold localization of centrin, an ubiquitous centrosomal protein, and p210, a recently characterized basal apparatus component of S. similis. Fibrous disks representing probasal bodies appear at the proximal end of parental basal bodies at the end of interphase and development proceeds via a ring of nine singlet microtubules. Nascent basal bodies dock early to the plasma membrane but p210, usually present in basal body-membrane-linkers of S. similis, was already present on the cytosolic basal body precursors. In addition to the distal connecting fiber and the nuclear basal body connectors (NBBC) of the parental basal bodies, centrin was present on the fibrous probasal bodies, in a linker between probasal bodies and the basal apparatus, in the connecting fiber between nascent basal bodies and their corresponding parent, and, finally, a fiber linking the nascent basal bodies to the nucleus. This NBBC probably is present only in mitotic cells. During elongation a cartwheel of up to seven layers is formed, protruding from the proximal end of nascent basal bodies. Microtubules develop on the cartwheel indicating that it temporarily functions as a microtubule organizing center (MTOC). These microtubules and probably the cartwheels, touch the nuclear envelope at both sides of a nuclear projection. We propose that spindle assembly is initiated at these attachment sites. During metaphase, the spindle poles were close to thylakoid-free lobes of the chloroplast, and the basal bodies were not in the spindle axis. The role of nascent basal bodies during the initial steps of spindle assembly is discussed.     

Dynamic Behavior of Microtubules during Dynein-dependent Nuclear Migrations of Meiotic Prophase in Fission Yeast

During meiotic prophase in fission yeast, the nucleus migrates back and forth between the two ends of the cell, led by the spindle pole body (SPB). This nuclear oscillation is dependent on astral microtubules radiating from the SPB and a microtubule motor, cytoplasmic dynein. Here we have examined the dynamic behavior of astral microtubules labeled with the green fluorescent protein during meiotic prophase with the use of optical sectioning microscopy. During nuclear migrations, the SPB mostly follows the microtubules that extend toward the cell cortex. SPB migrations start when these microtubules interact with the cortex and stop when they disappear, suggesting that these microtubules drive nuclear migrations. The microtubules that are followed by the SPB often slide along the cortex and are shortened by disassembly at their ends proximal to the cortex. In dynein-mutant cells, where nuclear oscillations are absent, the SPB never migrates by following microtubules, and microtubule assembly/disassembly dynamics is significantly altered. Based on these observations, together with the frequent accumulation of dynein at a cortical site where the directing microtubules interact, we propose a model in which dynein drives nuclear oscillation by mediating cortical microtubule interactions and regulating the dynamics of microtubule disassembly at the cortex.