The Nuclear Apparatus of the Ditransversal Ciliate Homalozoon vermiculare (Ciliophora,Rhabdophora) during Interphase and Division. I. The Macronucleus1

ABSTRACT. The nuclear apparatus of Homalozoon vermiculare consists of a single moniliform macronucleus and about 25 micronuclei. The number of macronuclear segments depends (i) on the number of divisions of individual segments during the interphase and (ii) on the number of segments that arise prior to cytokinesis from the (temporary) filiform macronucleus. Precytokinetic changes of the macronucleus involve the fusion of individual segments followed by contraction and subsequent elongation of the entire macronucleus. The chromatin bodies uncoil into fine fibrils during macronuclear contraction. At the time when the division furrow appears, the macronucleus starts to renodulate. The interphase segment contains a more or less reticulated chromatin body partly attached to the nuclear envelope and about 30 polymorphous nucleoli. The latter consist of the pars granulosa, the pars fibrosa, and an additional fibrillar component. The nucleoli undergo drastic changes prior to division and the granular component disappears completely during macronuclear condensation. On the average, the macronucleus contains a 3,400-fold amount of DNA compared with a haploid micronucleus, but the intraspecific differences in the DNA content of the entire macronucleus are extremely large. In contrast, DNA content and size of an individual segment of the macronucleus are precisely regulated during interphase.     

Ultrastructure of the nuclear apparatus of the lower ciliateRemanella granulosa Kahl (Karyorelictida)

Summary The nuclear apparatus ofRemanella granulosa has been investigated using conventional TEM methods and Bernhard's technique of preferential RNP staining. This species has two (rarely three) macronuclei and a single micronucleus (rarely two micronuclei). The nuclei always form a single group.The macronuclei contain a fibro-granular matrix resistant to EDTA destaining, and several nucleoli and chromatin bodies. The chromatin bodies are readily bleached with EDTA and are often clustered, or even fused, forming chromocenters. The nuclei are of the compact concentric type. Some macronuclei contain nuclear bodies, as finely fibrous spheres or bundles of coarse fibers, or both. Neither type of nuclear body is destained with EDTA. The spheres are frequently associated with nucleoli. There is no evidence of any transition between the two types of nuclear bodies. The macronuclear envelope contains numerous pore complexes and is strengthened with an electron dense layer. The micronucleus is filled with spongy condensed chromatin and surrounded by an envelope with occasional pores. This nucleus lacks nucleoli and nuclear bodies.     

Ultrastructure of the nuclear apparatus of the infusorian Loxodes magnus. I. The macronuclei, macronuclear anlagen and the interphasic micronuclei]

The nuclear apparatus of Loxodes magnus Stokes (Holotricha) consists of numerous macronuclei which belong to the diploid type and never divide, and of numerous micronuclei. No nuclear groups exist; individual nuclei often lie in cytoplasmic islets surrounded by large lacunae of the smooth endoplasmic reticulum. Interphasic micronuclei have two-membraned envelopes with numerous pores, usually lined at the cytoplasmic side with a layer of vacuoles, channels, or flattened vesicles of the smooth endoplasmic reticulum. The chromatin of the micronuclei consists of anastomosing threads, 0.1--0.2 mum wide, between which several nucleolus-like bodies of microfibrillar structure occur. Adult macronuclei have a similar nuclear envelope and a similar system of vacuoles, channels, and flattened agranular cisternae outside it. The macronucleus contains a single large composite nucleolus with 3 or 4 fibrillar cores inside the common granular cortex. The fibrillar cores are pierced by channels containing nucleolar organizers in the form of strands of condensed chromatin. The peripheral zone of the macronucleus is filled with decondensed chromatin fibrils and contains a number of small chromocenters and several aggregates of RNP granules. No protein inclusions (spheres) have been observed in Loxodes macronuclei. The macronuclear anlagen, developing in the cycle of every cell division, show progressive decondensation of the chromosomes and formation of several nucleoli, each with its own organizer. Later on, the nucleoli fuse into a single nucleolus. The small chromocentres are the last to form.     

Fine structure and cytochemistry of the nuclei of the primitive ciliateTracheloraphis crassus (Karyorelictida)

Summary The nuclei ofTracheloraphis crassus were studied using light and electron microscopy combined with Bernhard's RNP staining and pronase digestion. The nuclear apparatus of this species consists of a longitudinal row of 11–43 macronuclei and 4–16 micronuclei. Like in all karyorelictids, the macronuclei are unable to divide and become segregated during cytokinesis; their number is supplemented in every cell cycle by differentiation of several new macronuclei from micronuclei.Each adult macronucleus contains a single compact endonuclear aggregate of several large chromocenters, readily destained with EDTA, and several RNP containing nucleoli. There is continuity between the material of the chromocenters and the decondensed DNP fibrils in the nuclear matrix. The nucleoli contain NORs in the form of fibrillar centers. The endonuclear aggregate includes also groups of RNP granules which are especially resistant to EDTA destaining. A microfibrillar sphere, usually localized at the periphery of the aggregate, contacts one or several nucleoli. The sphere is not bleached with EDTA, and only its periphery becomes digested with pronase. The macronuclear matrix consists of both protein fibrils and pronase-resistant fibrils, the latter being localized at the nuclear periphery.Developing macronuclear primordia contain loose strands of decondensed chromatin; only later they form chromocenters and nucleoli.The micronuclei reproduce by mitosis with typical chromosomes (2n=66). During interphase, they are filled with condensed chromatin which can be bleached with EDTA; they form no nucleoli. Ring-like lamellae, existing in the cavities of the chromatin mass, stain for RNA (after Bernhard) and are pronase-sensitive. These lamellae resemble the kinetochore material conserved during interphase in another karyorelictid ciliate,Trachelocerca geopetiti.     

The intranuclear helices of Euplotes: Their substructure,localization, and apparent migration to the cytoplasm

Study of the fine structure of the macronucleus in Euplotes eurystomus, a ciliate protozoon, during various stages of the cell division cycle has yielded new information about intranuclear helices. They are frequently observed at the periphery of chromatin bodies or next to the nuclear envelope, and they appear to be a constituent of nucleoli. The fibril that forms a helix is about 11–15 nm thick, and torus profiles of helices cut in cross section are about 35 nm in diameter. In substructure the helix is composed of a thin strand 3–5 nm thick which is coiled to form the 11–15 nm fibril; so the helix is a super-coiled structure. The intranuclear helices are present in the macronucleus throughout the cell cycle. They do not show obvious changes of relative abundance nor changes of relative localization in the nucleus, with one exception: they were never observed in the diffuse zone of replication bands. Evidence is presented indicating that nuclear helices migrate to the cytoplasm through nuclear pores. Although the chemical composition of the Euplotes intranuclear helices is unknown, information in the literature on similar helices in Amoeba indicates that they contain RNA and not DNA. The observations on Euplotes helices are consistent with a concept of “packaged” RNA for transport to the cytoplasm.     

Fine structural organization of a non-reticulate plant cell nucleus

We studied the fine structural organization of the meristematic nucleus in roots of Lycopesicon esculentum (tomato) using ultracytochemical and immunocytochemical approaches. The nucleus has a non-reticulate (i.e. low DNA content) structure whose supramolecular organization differs in some respects from that in reticulate nuclei, principally in the organization of the chromocentres associated with the nuclear envelope, with which centromeric structures appear to be associated. The main difference at the nucleolar level is found in the fibrillar centres, which have a low amount of DNA labelling and in which inclusions of condensed chromatin are present only very rarely. The distribution of nucleolar DNA amongst the nucleolar compartments is similar to that in reticulate nucleoli as demonstrated using an anti-DNA monoclonal antibody. Tomato nuclei have nucleolus-associated bodies or karyosomes, like other plant species with a low DNA content and non-reticulate nuclear organization. The nuclear ribonucleoprotein structures in the inter- and perichromatin regions, namely inter- and perichromatin fibrils and granules, show similar ultrastructural and cytochemical characteristics in both types of nuclei.Abbreviations NAC nucleolus associated chromatin - CES centromeric structures - NOR nucleolar organizing region - NAB nucleolus associated body - IG interchromatin granules - RNP ribonucleoprotein - Mab monoclonal antibody by M.F. Trendelenburg     

MITOSIS IN THE OCTAFLAGELLATE PRASINOPHYTE,PYRAMIMONAS AMYLIFERA (CHLOROPHYTA)1

Cell division is described in the octaflagellate prasinophyte Pyramimonas amylifera Conrad and is compared in related genera. Basal bodies replicate at preprophase and move toward the poles. Cells remain motile throughout division. The nuclear envelope disperses and chromosomes begin to condense at prophase. Pairs of multilayered kinetochores are evident on the chromosomes of the metaphase plate. Spindle microtubules extending from the region of the basal bodies and rhizoplasts attach to the kinetochores or extend from pole to pole. Numerous vesicles and ribosomes have entered the nuclear region and the incipient cleavage furrow invaginates. The chromosomes move toward the poles at anaphase leaving a broad interzonal spindle between the two chromosomal plates. The nuclear envelope reforms first around the chromatin on the side adjacent to the spindle poles and later on the interzonal side. The cleavage furrow progresses into the interzonal spindle at telophase. By late telophase the nucleoli have reformed and the chromosomes have decondensed. The interzonal spindle has not been observed late in telophase. As the cleavage furrow nears completion the cells begin to twist and contort, ultimately separating the two cells.     

The ultrastructure of mitosis during sporangiogenesis inWoronina pythii (Plasmodiophoromycetes)

Summary Mitotic divisions during sporangiogenous plasmodial cleavage inWoronina pythii were studied with transmission electron microscopy. We conclude that these nuclear divisions (e.g., transitional nuclear division, and sporangial mitoses) share basic similarities with the cruciform nuclear divisions inW. pythii and other plasmo-diophoraceous taxa. The major distinction appeared to be the absence of nucleoli during sporangial mitosis and the presence of nucleoli during cruciform nuclear division. The similarities were especially evident with regard to nuclear envelope breakdown and reformation. The mitotic divisions during formation of sporangia were centric, and closed with polar fenestrae, and characterized by the formation of intranuclear membranous vesicles. During metaphase, anaphase, and telophase, these vesicles appeard to bleb from the inner membrane of the original nuclear envelope and appeared to coalesce on the surface of the separating chromatin masses. By late telophase, the formation of new daughter nuclear envelopes was complete, and original nuclear envelope was fragmented. New observation pertinent to the mechanisms of mitosis in thePlasmodiophoromycetes include a evidence for the incorporation of membrane fragments of the original nuclear envelope into new daughter nuclear envelopes, and b the change in orientation of paired centrioles during sporangial mitosis.     

Ultrastructural characteristics of insect corpora allata in relation to larval diapause

Summary The ultrastructure of active and inactive corpora allata from last instar larvae of the southwestern corn borer, Diatraea grandiosella, was examined. Active glands were obtained from pre-, early, and mid-diapausing larvae; inactive ones from late and non-diapausing larvae. Each gland contains 13 to 18 cells which have the following common features: well developed smooth endoplasmic reticulum, scattered rough endoplasmic reticulum and free ribosomes, microtubules, vacuolated nucleoli, and interlocking plasma membranes. The gland contains intercellular deposits, and is supplied by regular and neurosecretory axons.Special ultrastructural features of the corpus allatum from the five groups of larvae examined were as follows: pre-diapause: extensive vesicular smooth endoplasmic reticulum, numerous cup-shaped mitochondria and Golgi bodies with stacked cisterns and vesicles, few small lipid droplets, large nuclei with dispersed chromatin, absence of lysosomes; early diapause: stacked, whorled, and vesicular smooth endoplasmic reticulum of equal abundance, numerous rod-shaped mitochondria, some Golgi bodies but without distinct stacks of cisterns, few lipid droplets and lysosomes, chromatin dispersed and also attached to the nuclear envelope; mid-diapause: similar to early diapause except for the presence of more stacked, smooth endoplasmic reticulum, chromatin in large chunks mostly attached to the nuclear envelope; late diapause: whorled smooth endoplasmic reticulum and rod-shaped mitochondria predominating, complicated Golgi bodies with stacks of cisterns and large empty sacs, few large lipid droplets, some lysosomes containing mainly whorled bodies, chromatin in large chunks attached to the nuclear envelope; non-diapause: similar to late diapause except for less extensive smooth endoplasmic reticulum, more abundant mitochondria, fewer intercellular deposits. Although these observations suggest that the smooth endoplasmic reticulum, and possibly mitochondria, and Golgi bodies are involved in juvenile hormone production, specific sites of synthesis or storage of the hormone were not revealed.Supported in part by grant no. PCM 74-18155 A01 from the National Science Foundation. Contribution from the Missouri Agricultural Experiment Station as journal series no. 8234. We thank Ms. L. Yin for her skillful assistance, and Dr. M.F. Brown of the College of Agriculture Electron Microscope Facility for his advice and the use of equipment.     

Nucleolar behaviour during division in the homobasidiomycetidae

Summary

The nucleoli of Oudemansiella mucida and Nolanea cetrata become disassociated from the parent chromatin during both mitosis and meiosis. Disassociation of the nucleoli from the post-meiotic nuclei at the onset of migration is taken, along with other evidence, to indicate that the third nuclear division is initiated in the basidium and completed in the basidiospores. It is postulated that the coincidence of migration and the third nuclear division arises from the inability of nucleoli to pass through the sterigmata. This view is supported by observations made by the authors on other members of the Homobasidiomycetidae.     

Electron Microscopy of the Nuclear Events During Binary Fission in Paramecium multimicronucleatum

SYNOPSIS. From the interphase to the early stage of binary fission in Paramecium multimicronucleatum , when the micro-nuclei are situated close to the macronucleus, the microtubules in the cytoplasm seem to connect the nuclear pores of macro- and micronuclei. During the 1st half of macronuclear division, the microtubules are formed outside the macronucleus, while during the latter half of division, numerous microtubules appear inside it. Chromatin bodies and nucleoli remain unchanged during macro-nuclear division, but the latter show temporory irregularity in shape. In late prophase of micronuclear division, spindle micro-tubules are formed, and a polar structure, composed of randomly dispersed twisting filaments, is formed at each pole of the micro-nucleus at anaphase. Spindle microtubules terminate on the surface of this structure. The nuclear envolope of the macro-and micronuclei remains intact thruout division. The envelope of the daughter micronuclei is derived from the pre-existing one.     

Ultrastructure of meristematic cells of dormant and released buds in Tradescantia paludosa

The lateral bud meristems of Tradescantia paludosa show a characteristic cytohistological zonation during dormancy. The cells comprising this so called ‘zone of inhibition’, which is located at the extreme tip of the bud apex, rarely synthesize nuclear DNA or undergo mitotic division. These nuclei are as large as prophase nuclei, yet contain only telophase (2C) amounts of DNA and significantly lower amounts of histone as compared to the 2C nuclei of the actively dividing cells.Ultrastructural observations of the nuclei in the ‘zone of inhibition’ show that a large proportion of the chromatin is organized as less condensed, diffuse, euchromatin fibrils; however, the chromatin of the actively dividing nuclei of the cells outside the ‘zone of inhibition’ or in the released bud meristems is organized to a greater extent as condensed clumps of heterochromatin. When the dormancy is released, the nuclei in the ‘zone of inhibition’ synthesize DNA and histone and undergo cell division in approx. 4 days. Striking changes in the organization of chromatin fibrils take place during this transition period. The diffuse chromatin fibrils of the nuclei in the ‘zone of inhibition’ progressively become more and more condensed as the cell prepares to undergo the first mitotic division after the release of dormancy. This change which is coupled with the synthesis of histones in the nuclei of the ‘zone of inhibition’ suggests a prominent structural role of these basic proteins in the organization of the chromatin. The large volume of 2C nuclei of the ‘zone of inhibition’ seems, therefore, to result not from a great nuclear mass, but probably from a relatively small degree of condensation of chromatin.     

MACRONUCLEAR EVENTS IN SYNCHRONOUSLY DIVIDING TETRAHYMENA PYRIFORMIS

The macronuclei of synchronously dividing mass cultures of Tetrahymena pyriformis (strain WH₆) were examined with the electron microscope for changes during two division cycles. Samples were prepared at 30-minute intervals for a period of 8½ hours which included the time required to induce synchrony by five heat shocks (4½ hours). The interphase macronucleus contains peripheral, crescent-shaped nucleoli and evenly distributed chromatin bodies. Centrally located RNA bodies, composed of fibers, appear 1 to 2 hours following the initial heat shock. They are completely destroyed with ribonuclease whereas the nucleoli are only partially so. Following the third heat shock the RNA bodies move to the periphery and disintegrate; the nucleoli aggregate and form blebs which protrude into the cytoplasm where they appear to pinch off and may contribute to the cytoplasmic ribonucleic acid. Cytokinesis does not occur at this time. Instead the nuclear events are repeated during the 4th and 5th hours, even though the heat shocks are terminated at 4½ hours. Cytokinesis takes place at about 6 hours. The second division occurs about 2½ hours later during which all the macronuclear events noted above are repeated.     

Three-dimensional analysis of the arrangement of compact chromatin in the nucleus of G0 rat lymphocytes

The arrangement of compact chromatin of G₀ lymphocytes was studied in three-dimensional reconstructions of the ensemble of the chromatin and of individual compact chromatin bodies. Rat spleen was serially cut and sections were contrasted with procedures preferential for DNA. Electron microscopy images were digitized, processed, and displayed using a commercial soft-ware package, complemented by a system for three-dimensional reconstruction and analysis developed by us on an IBM-compatible microcomputer provided with an image acquisition board. The reconstructions showed a continuous layer of compact chromatin in contact with the nuclear envelope that prevents the automatic recognition of individual chromatin clumps. The ensemble of the arrangement of compact chromatin was found to be very similar in different lymphocytes. After morphological filtering procedures, the initial mass was divided into individual bodies of compact chromatin, which were tagged. Most of these bodies contact the nuclear envelope. The number of bodies as well as the number of contacts with the envelope are similar and correspond to a haploid number of chromosomes. The largest body is always the one containing nucleolus-associated chromatin. When the cell has two nucleoli, the nucleolus-associated chromatin bodies contact the envelope in diametrically opposed areas. This feature was also described in rat liver cells. It is concluded that: (a) the individualized compact chromatin bodies do not correspond to an entire chromosome or to a pair of chromosomes; (b) the arrangement of compact chromatin is not identical in each G₀ lymphocyte, but there are patterns that are repeated with limited changes; and (c) there are common features that appear in different cell types of individuals of the same species.     

CRUCIFORM NUCLEAR DIVISION IN WORONINA PYTHII (PLASMODIOPHOROMYCETES)

Somatic nuclear divisions in sporangiogenous plasmodia of Woronina pythii Goldie-Smith were studied with transmission electron microscopy. During metaphase, each nucleus formed a cruciform configuration as chromatin became aligned at the equatorial plate perpendicular to the persistent nucleolus. Except for polar fenestrations, the original nuclear envelope remained intact throughout the mitotic division. Intranuclear membranous vesicles appeared to bleb off the inner membrane of the original nuclear envelope, adhered to the surfaces of the separating chromatin, and eventually formed new daughter nuclear envelope within the original nuclear envelope. During the first 24 hr of vegetative plasmodial growth, each telophase nucleus exhibited an obvious constriction of the original nuclear envelope in the interzonal region. Similar constrictions were not evident in telophase nuclei found in 24–36-hr-old plasmodia. This variation in the ultrastructural morphology of cruciform division appears to be related to the age and size of each sporangiogenous plasmodium, and is the first to be documented within this group of fungal pathogens.     

Effects of ultracentrifugation on the interphase nucleus of somatic cells with special reference to the nuclear envelope-chromatin relationship

Summary Ultracentrifugation of living cells from the liver of the mouse, rat, dog, frog, Necturus, follicle cells, of grasshopper testis, and meristem of the onion root tip shows evidence that the interphase chromatin is attached to the nuclear envelope. Because of its relatively high density, the bulk of the interphase chromatin, and often the nucleoli, are displaced to the centrifugal side of the nucleus and, when this occurs, the chromatin bodies attached to the centripetal side of the nucleus are drawn out into long filaments which extend across the nucleus centrifugally. They generally break before becoming detached from the envelope. Onion root tip chromosomes in early prophase also appear to be attached to the nuclear envelope. The Barr body strongly adheres to the nuclear envelope as evidenced by the high centrifugal force necessary to displace it. Nucleoli of ultracentrifuged meristematic cells of the onion root show evidence of a stratification of materials within them.Supported by Grant GM 04706 from the U.S.P.H.S.     

Ultrastructural changes in major organelles during spermatial differentiation inBangia (Rhodophyta)

Summary The major organelles within the cells of maleBangia atropurpurea (Roth) C. Ag. filaments undergo a series of ultrastructural transformations during the production of spermatia. Initially, thylakoids within the large axial chloroplast develop a reticulate pattern commencing at the central pyrenoid region. Subsequent changes involve loss of lobes and diminution of volume through division; chloroplasts in final stages contain a few dilated, distorted thylakoids and many plastoglobuli. During differentiation the large nucleolus disappears from the nucleus and four masses of chromatin aggregate near the nuclear envelope. Furrows originating from the nuclear envelope form double membranes around each of the chromatin masses and most of the nucleoplasm is eliminated. Several types of fibrillar vesicles are formed during the process and large floridean starch reserves are utilized. Multilamellar bodies and microbody-like structures occur within the cells during certain phases of spermatiogenesis.