Nucleus-associated Organelle Research Articles

Fission yeast cdc31p is a component of the half-bridge and controls SPB duplication.

The fission yeast spindle pole body (SPB) is a nucleus-associated organelle that duplicates once each cell cycle during interphase. Duplicated SPBs serve as the poles of an intranuclear mitotic spindle after their insertion into the nuclear envelope in mitosis (Ding et al., Mol. Biol. Cell 8, 1461-1479). Here, we report the identification and characterization of Schizosaccharomyces pombe cdc31p, a member of the conserved calcium-binding centrin/CDC31 family. Immunofluorescence and immunoelectron microscopy show that cdc31p is a SPB component localized at the half-bridge structure of the SPB. cdc31 is an essential gene and Deltacdc31 cells and cdc31 conditional mutant cells arrest in mitosis with a monopolar mitotic spindle organized from a single SPB. EM analysis demonstrates that mutant cdc31 cells fail to duplicate the SPB. In addition, cdc31p exhibits genetic interactions with the SPB component sad1p and is required for sad1p localization. Finally, cdc31 mutant can undergo single or multiple rounds of septation before the exit from mitosis, suggesting that cdc31p activity or SPB duplication may be required for the proper coordination between the exit from mitosis and the initiation of septation.

The morphology of Ichthyophonus sp. in their mugilid hosts (Pisces: Teleostei) and following cultivation in vitro. A light and electron microscopy study.

The morphology of Ichthyophonus sp., a parasite of Mugil capito and Liza saliens, was investigated by light and transmission electron microscopy. The most frequent stage found in the fish hosts was the multinucleate spore, though germinating stages, hyphae, and endospores were also found. Different development patterns were observed in the media assayed for in vitro culture. Optimal growth and development were obtained in Eagle's minimum essential medium (MEM) supplemented with 10% fetal bovine serum at pH 7. Ultrastructural features of multinucleate spores, both in the fish host and in culture, were a fibrillar thick wall and an electron-lucent matrix, with large glycogen granules, some electron-dense bodies, large vacuoles, lipid inclusions, and endoplasmic reticulum mainly appearing among the nuclei. Mitochondria with scarce tubulovesicular cristae were observed in the different stages, mainly near the wall and the germinating sites. Condensed heterochromatin was rarely seen. A nucleus-associated organelle (NAO) was frequently observed, and dictyosome cisternae and vesicles appeared in its vicinity.

A role for NIMA in the nuclear localization of cyclin B in Aspergillus nidulans.

NIMA promotes entry into mitosis in late G2 by some mechanism that is after activation of the Aspergillus nidulans G2 cyclin-dependent kinase, NIMXCDC2/NIMECyclin B. Here we present two independent lines of evidence which indicate that this mechanism involves control of NIMXCDC2/NIMECyclin B localization. First, we found that NIMECyclin B localized to the nucleus and the nucleus-associated organelle, the spindle pole body, in a NIMA-dependent manner. Analysis of cells from asynchronous cultures, synchronous cultures, and cultures arrested in S or G2 showed that NIMECyclin B was predominantly nuclear during interphase, with maximal nuclear accumulation in late G2. NIMXCDC2 colocalized with NIMECyclin B in G2 cells. Although inactivation of NIMA using either the nimA1 or nimA5 temperature-sensitive mutations blocked cells in G2, NIMXCDC2/NIMECyclin B localization was predominantly cytoplasmic rather than nuclear. Second, we found that nimA interacts genetically with sonA, which is a homologue of the yeast nucleocytoplasmic transporter GLE2/RAE1. Mutations in sonA were identified as allele-specific suppressors of nimA1. The sonA1 suppressor alleviated the nuclear division and NIMECyclin B localization defects of nimA1 cells without markedly increasing NIMXCDC2 or NIMA kinase activity. These results indicate that NIMA promotes the nuclear localization of the NIMXCDC2/ NIMECyclin B complex, by a process involving SONA. This mechanism may be involved in coordinating the functions of NIMXCDC2 and NIMA in the regulation of mitosis.

Ultrastructure of vegetative organization and cell division in Glaucosphaera vacuolata Korshikov (Porphyridiales, Rhodophyta)

Abstract The ultrastructure of vegetative cells and cell division has been examined in Glaucosphaera vacuolala Korshikov. Features either not found or not denoted before include apparent intercisternal fusions within each Golgi body and fibrillar tracts possibly utilized by Golgi vesicles. Mitosis is closed except for large polar gaps and a spherical, presumptive MTOC (microtubule organizing centre) is present at prophase. The MTOC flattens by metaphase to fill the polar gap, becoming the focus of spindle microtubules. No NAO (nucleus associated organelle) was observed. All aspects of mitosis except the absence of an NAO are consistent with mitosis in red algae; most are not consistent with members of the Glaucophyta. Characteristics of mitosis and vegetative features, both those newly observed and those confirmed (especially the presence of connections between chloroplast lobes), support the removal of G. vacuolala from the Glaucophyta and its placement in the Rhodophyta. Also noted in this investigation...

Ultrastructure of cell division inAgardhiella subulata(Gigartinales, Rhodophyta)

The fine structure of cell division in Agardhiella subulata is consistent with the basic pattern observed for most multicellular red algae. Unusual features include complete microtubule-filled tunnels through the nucleus (prophase), a nucleus-associated organelle penetrated by the outer membrane of the nuclear envelope (prophase) and a metaphase-anaphase polar region that appears in thin section as two large gaps separated by a central intact fragment of nuclear envelope. The only other red alga in which the latter two features have been detected is Lomentaria baileyana (Rhodymeniales). This study is the first complete ultrastructural description of cell division in a member of the Gigartinales. In addition, it provides information on features that may be useful as taxonomic indicators for members of the Gigartinales and the Rhodymeniales.

ULTRASTRUCTURE OF VEGETATIVE ORGANIZATION AND CELL DIVISION IN THE UNICELLULAR RED ALGA DIXONIELLA GRISEA GEN. NOV. (RHODOPHYTA) AND A CONSIDERATION OF THE GENUS RHODELLA1

ABSTRACTThis study suggests that the genus Rhodella be restricted to that set of features currently observed only in Rhodella maculata Evans and Rhodella violacea (Korn‐mann) Wehrmeyer, that a new genus Dixoniella be established to accommodate the unicellular red alga, Rhodella grisea (Geitler) Fresnel, Billard, Hindák et Pekár‐ková, and that Rhodella cyanea Billard et Fresnel be further studied for probable reclassification. These conclusions are based on ultrastructural comparisons of Dixoniella grisea with published information on the genus Rhodella. The presence of thylakoids in the pyrenoid, a peripheral encircling thylakoid in the chloroplast, a dictyosome/nuclear envelope association, and the lack of a specialized pyrenoid/nucleus association in D. grisea separate this alga from the genus Rhodella. Cell division in D. grisea is not demonstrably different from that in Rhodella, although the unusually well‐defined material of the presumptive microtubule organizing center (MTOC) made it possible to follow the development and behavior of the MTOC to a greater degree than in previously studied red algal cells. The surprising amount of conformity in cell division characters between D. grisea and the genus Rhodella prompted a comparison of cell division characteristics in all red algal unicells studied to date.All unicells show a remarkable degree of similarity except for differences in interzonal spindle length, dissimilarities in size of the nucleus‐associated organelle (NAO), and the unusual NAO of Porphyridium purpureum (Bory) Drew et Ross.

Ultrastructure of vegetative organization and cell division in the freshwater red algaCompsopogon

Uniseriate filaments of the freshwater red algaCompsopogon coeruleus were examined by transmission electron microscopy for details of vegetative organization and cell division with the goal of providing useful taxonomic characters. Each cell's single, complex chloroplast contains a peripheral encircling thylakoid, and unlike the vast majority of red algae, the cis-regions of dictyosomes are not consistently juxtaposed with mitochondria. These subcellular features, which are present in all examined genera in theCompsopogonales, Erythropeltidales, andRhodochaetales, along with certain unique reproductive characteristics, unify these three orders. During mitosis in uncorticated axial cells, a small, ring-shaped nucleus associated organelle (NAO) is located at each division pole, an intranuclear spindle comes to a moderately acute focus at the flattened, fenestrated metaphase-anaphase division poles and perinuclear ER partially encloses dividing nuclei, including a well-developed interzonal midpiece. The cleavage furrow penetrates the large, central vacuolar region to separate daughter nuclei. These cell division features most closely resemble the pattern described for the orderCeramiales. Our observations of vegetative and dividing cells ofC. coeruleus supplement the growing volume of evidence in favour of uniting all red algae into a single class without subclass designations.

The ultrastructure and timing of events in the nuclear cycle of the fungus Entomophaga aulicae

ABSTRACT The ultrastructure of the mitotic nuclear division cycle of the fungus Entomophaga aulicae was studied from serial sections of hyphal tips and protoplasts. The extranuclear bar-shaped nucleus-associated organelle (NAO) remained associated with the persistent nuclear envelope throughout. Prior to spindle formation, a patch of intranuclear NAO-associated chromatin detached from the nuclear envelope to yield a chromatinfree zone containing fine filaments and a linear array of presumptive kinetochores. Early metaphase spindles less than 1 pm in length were characterized by a ‘fused’ metaphase plate consisting of kinetochore-associated chromatin and a full complement of at least 15 kinetochore microtubules per half-spindle, while most of the chromatin was remote from the intranuclear spindle. Analysis of the distribution of antiparallel spindle microtubules indicated that polar separation and concomitant spindle elongation through metaphase were not accompanied by intermicrotubule sliding. Anaphase exhibited extensive decondensation of the large patches of condensed chromatin characteristic of all other stages. In a logarithmically growing protoplast population all nuclei contained spindle microtubules, with metaphase occupying approximately 66% of the nuclear cycle time. The calculated genome size of 4-3 pg, and average DNA content per chromosome of 0-3 pg, are extremely high for fungi.

Ultrastructure of basidium and nucleus associated organelle in Filobasidiella neoformans: Its application as a taxonomic character

Filobasidiella neoformans, the teleomorph of Cryptococcus neoformans, was observed using electron microscopy to confirm its phyletic status. No septa were found in the basidium of F. neoformans. Mature basidiospores were encapsulated by a relatively thick cell wall. Basidiospores directly budded from the basidium without sterigma formation, and no hilar appendix was noted. A lateral component of a synaptonemal complex was observed. As for mitotic apparatus, a spherical nucleus associated organelle (NAO), 0.12 μm in diameter, was found in the hemispherical depressive area of the nuclear envelope. Therefore, this fungus seems to have both characteristics of the Holobasidiomycetidae and of the primitive Basidiomycotina such as the Teliomycetes.

Ultrastructure of cell division in the unicellular red alga Flintiella sanguinaria

Cell division was examined with the electron microscope in the unicellular red alga Flintiella sanguinaria. Flintiella morphologically resembles Porphyridium purpureum, the only other red algal unicell that has been examined for ultra-structural details of cell division. Both genera are in the order Porphyridiales (Bangiophyceae), an unnatural assemblage of reduced forms in great need of taxonomic revision. Before mitosis, the nucleus migrates to the cell periphery. As in all red algae examined, except Porphyridium, the nucleus-associated organelle is a polar ring. At prometaphase, one or two microtubule-containing cytoplasmic invaginations penetrate the nucleus at each pole. By metaphase an intranuclear spindle is found in association with a typical metaphase chromosomal plate. The nuclear envelope is intact except for a single, large gap at each pole. Perinuclear endoplasmic reticulum is absent. An elongated anaphase interzonal midpiece is formed which breaks down at telophase. The daughter nuclei are widely separated by the chloroplast, which is constricted at the same time as daughter cell formation occurs by a cleavage furrow associated with a finely filamentous region similar to the contractile ring found in many animal cells. Because of pronounced differences in several mitotic features, it is concluded that Flintiella is not closely related to Porphyridium and instead shows closer phylogenetic ties with other macroscopic genera.

Mitosis in the dermatophyteMicrosporum canis as revealed by freeze substitution electron microscopy

Mitosis in the dermatophyteMicrosporum canis was studied by freeze substitution and electron microscopy, and analyzed by three dimensional reconstruction from serial sections of the mitotic nuclei. The interphase nucleus has associated nucleus-associated organelle (NAO) on a portion of the outer surface of the nuclear envelope, subjacent to which there was dense intranuclear material. The NAO divided and separated on the envelope, and a spindle was formed. The spindle was composed mostly of microtubules extended between opposite NAOs. Pairing of kinetochores was observed in the spindle from an early stage of development, when chromosomes were not so condensed, and remained unchanged while chromosome condensation proceeded until metaphase. Before the completion of nuclear division, daughter nuclei were connected by a narrow spindle channel, and then the nucleolus, whose structure underwent minimal change during mitosis, was eliminated into the cytoplasm.

The Structure and Cycle of the Nucleus-Associated Organelle in Two Species of Basidiobolus

The ultrastructure ofthe nucleus-associated organelle (NAO) is described in Basidiobolus haptosporus and B. magnus. NAO morphology is the same in both species. Each NAO is composed of a cylinder of 11-12 singlet microtubules embedded in an osmiophilic matrix. A small satellite is usually associated with each NAO. Interphase cells possess two NAOs, replication apparently occurring during mitosis. The morphology of the Basidiobolus NAO is unusual since no other non-flagellated organism contains microtubules as structural com? ponents of their NAO, and all microtubule-containing NAOs (centrioles) show 9-fold symmetry.

The Structure and Cycle of the Nucleus-Associated Organelle in Two Species of Basidiobolus

The ultrastructure of the nucleus-associated organelle (NAO) is described in Basidiobolus haptosporus and B. magnus. NAO morphology is the same in both species. Each NAO is composed of a cylinder of 11–12 singlet microtubules embedded in an osmiophilic matrix. A small satellite is usually associated with each NAO. Interphase cells possess two NAOs, replication apparently occurring during mitosis. The morphology of the Basidiobolus NAO is unusual since no other non-flagellated organism contains microtubules as structural components of their NAO, and all microtubule-containing NAOs (centrioles) show 9-fold symmetry.

The taxonomic significance of the mitotic spindle and nucleus-associated organelle ofEntomophaga aulicae

Nuclei in protoplasts ofEntomophaga aulicae contain abundant condensed chromatin and a large central nucleolus. The metaphase spindle occupies a small eccentric area of the nucleus while the remainder of the nucleus is filled with condensed chromatin. Small portions of condensed chromatin are aligned along a broad metaphase plate and connected to the spindle poles by kinetochore microtubules. The nucleus associated organelle (NAO) is a solid barlike structure which lies at the spindle poles and is closely associated with the outer membrane of the nuclear envelope. Comparison of the nuclear characteristics ofE. aulicae with those of other members of theEntomophthorales supports the separation of theEntomophthoraceae from theBasidiobolaceae andAncylistaceae. Further comparison of details of nuclear division in theEntomophthoraceae, specifically NAO morphology, may be useful in helping to delineate evolutionary lines within the family.

Location of actin, myosin, and microtubular structures during directed locomotion of Dictyostelium amebae.

During their life cycle, amebae of the cellular slime mould Dictyostelium discoideum aggregate to form multicellular structures in which differentiation takes place. Aggregation depends upon the release of chemotactic signals of 3',5'-cAMP from aggregation centers. In response to the signals, aggregating amebae elongate, actively more toward the attractive source, and may be easily identified from the other cells because of their polarized appearance. To examine the role of cytoskeletal components during ameboid locomotion, immunofluorescence microscopy with antibodies to actin, myosin, and to a microtubule-associated component was used. In addition, rhodamine-labeled phallotoxin was employed. Actin and myosin display a rather uniform distribution in rounded unstretched cells. In polarized locomoting cells, actin fluorescence (due to both labeled phallotoxin and specific antibody) is prevalently concentrated in the anterior pseudopod while myosin fluorescence appears to be excluded from the pseudopod. Similarly, microtubules in locomoting cells are excluded from the leading pseudopod. The cell nucleus is attached to the microtubule network by way of a nucleus-associated organelle serving as a microtubule-organizing center and seems to be maintained in a rather fixed position by the microtubules. These findings, together with available morphological and biochemical evidences, are consistent with a mechanism in which polymerized actin is moved into the pseudopod through its interaction with myosin at the base of the pseudopod. Microtubules, apparently, do not actively participate in movement but seem to behave as anchorage structures for the nucleus and possibly other cytoplasmic organelles.

Ascospore development in the fission yeasts Schizosaccharomyces pombe and S. japonicus.

The fine structure of ascospore formation in the fission yeasts Schizosaccharomyces pombe and Schizosaccharomyces japonicus var. japonicus was studied by serial thin-sectioning and electron microscopy. The morphogenetic events were almost the same in both species. Ascospore development was initiated by the formation of the forespore membrane on the cytoplasmic side of the differentiated nucleus-associated organelle (NAO) in the interval between meiosis I and II in S. pombe, or during the post-meiotic nuclear division in S. japonicus, and the process proceeded almost synchronously through the two or four nuclei in the ascus. The forespore membrane developed by fusion of the cytoplasmic vesicles and this was clearly demonstrated in S. japonicus where the behaviour of vesicles involved in the forespore membrane development could be traced as they were marked by the presence of electron-dense granules. The staining technique, by phosphotungustic acid--chromic acid (PTA-CA) after treatment with periodic acid, was used to attempt to elucidate the origin and the nature of the forespore membrane. The method specific to plasmalemma-type membranes stained both ascus and ascospore plasmalemmas; the forespore membrane was not stained at first but developed the same affinity for stain as the plasma membrane in the course of ascospore development. The results suggest that the forespore membrane did not come directly from the ascus plasma membrane, but from another membrane system such as the endoplasmic reticulum. Spore wall material was deposited in the space between the inner and outer leaflets of the forespore membrane.

An electron microscopic study on the mating tube formation in the heterobasidiomycete Tremella mesenterica

Ultrastructure of the mating tube formed in yeast haplont of the heterobasidiomycete Tremella mesenterica was studied by electron microscopy. Cell wall of the mating tube emerged as evagination of the inner layers, rupturing outer layers of the mother cell wall. Comparison with budding cells suggested that the tube emergence place at bud scar and the process of tube emergence was the same as that of bud emergence. Electron transparent vesicles of 0.1 μm diameter were scattered in the cytoplasm of the mating tube. Nucleus-associated organelle was located at one side of the nuclear envelope which extended towards the mating tube. A few microtubules were detected in the mating tube, but their association with a nucleus was not clear. The cytoplasmic structure of the mating tube was discussed in comparison with that of hyphae of the filamentous fungi.

Ultrastructure of mitosis in the cowpea rust fungus Uromyces phaseoli var. Vignae.

Aspects of the ultrastructure of mitotic nuclei of the fungus Uromyces phaseoli var. vignae are described from both intercellular hyphae in the cowpea host and infection structures induced to differentiate in vitro. The interphase nucleus-associated organelle (NAO) consists of two trilamellar acircular disks connceted by an osmiophilic bar. The intranuclear spindle develops between these disks when they separate. The spindle contains pole to pole, interdigitating, chromosomal, and fragmentary microtubules arranged to form a central bundle along the surface of which lie the metaphase chromosomes. No metaphase plate is found. There are up to three microtubules per kinetochore and approximately 14 chromosomes on the haploid spindle. Telophase elongation appears to involve extension of pole to pole microtubules with no evidence for the remaining presence of interdigitating microtubules. Concomitantly, numerous cytoplasmic microtubules develop from each NAO disk where few or none are present in other phases. Reformation of the interphase NAO involves the formation of a sausage-shaped intermediate at late telophase. The nuclear envelope remains intact and the nucleolus persists throughtout division. Various aspects of the spindle and NAOs appear to be evolutionary intermediates between Ascomycetes and higher Basidiomycetes, thus supporting the theory of Basidiomycete evolution from the former group and demonstrating an encouraging correlation between mitotic characteristics and other phylogenetic markers.

Ultrastructure of the fungal nucleus. III. Genesis of the nucleus-associated organelle (NAO equals "KCE")

The nucleus-associated organelle (NAO) is a structure, significant for division and motion of the fungal nucleus. In the basidiomycete Polystictus (equals Trametes) versicolor it is composed during somatic interphase of two globular entities (GE) and the connecting middle plate (MP). Its genesis is followed electron optically by analysing serial sections of preselected stages. Genesis is strictly determined by time and space. It starts with synthesis of the new MP a few minutes upon karyokinesis has been completed. Synthesis is localized in the area of contact between old GE and outer membrane of the nuclear envelope. After growing of the MP to its maximal size both new GE's develop from swellings at its ends. Old Ge's are dissolved during this developmental process. The period of genesis takes 30--40 min and fills one third of the cell cycle. A GE-field exhibits characteristic alterations throughout the formation of the NAO. It is composed of GE, periglobular zone, microtubules, and both caps of endoplasmic reticulum and multivesicular bodies.

Ultrastruktur des Pilzkernes. III. Genese des Kern‐assoziierten Organells (NAO = „KCE”︁)

AbstractThe nucleus‐associated organelle (NAO) is a structure, significant for division and motion of the fungal nucleus. In the basidiomycete Polystictus (= Trametes) versicolor it is composed during somatic interphase of two globular entities (GE) and the connecting middle plate (MP). Its genesis is followed electron optically by analysing serial sections of preselected stages. Genesis is strictly determined by time and space. It starts with synthesis of the new MP a few minutes upon karyokinesis has been completed. Synthesis is localized in the area of contact between old GE and outer membrane of the nuclear envelope. After growing of the MP to its maximal size both new GE's develop from swellings at its ends. Old Ge's are dissolved during this developmental process. The period of genesis takes 30–40 min and fills one third of the cell cycle. A GE‐field exhibits characteristic alterations throughout the formation of the NAO. It is composed of GE, periglobular zone, microtubules, and both caps of endoplasmic reticulum and multivesicular bodies.