A light and electron microscopic study of mitosis inBullera alba and the histochemistry of some cytoplasmic substances

Summary Mitosis in the imperfect yeast-like basidiomyceteBullera alba was studied by comparative light and electron microscopy. During mitosis the chromatin containing part of the nucleus moved into the progeny cell, and the nucleolus containing part of the nucleus remained in the parent cell. The two portions of the nucleus then separated and the nucleolar part degenerated. Metaphase and anaphase took place in the progeny cell. Subsequently one mass of chromatin returned to the parent cell, and two new nuclei were formed. The study concentrated on the nuclear envelope, nucleolus, spindle pole body, chromatin, spindle, and cytoplasmic microtubules. Mitosis inB. alba was compared with reports of mitosis in other basidiomycetes, theUredinales, and theAscomycotina and was deemed closest to the heterobasidiomycete yeasts.Histochemical evidence for the presence of lipid, glycogen, and polyphosphate in the cytoplasm was presented.     

Mitosis in the basidiomycete fungusTulasnella araneosa

Summary This is a report of a light and electron microscopic study of mitosis in the basidiomycetous fungusTulasnella araneosa. The study employs serial section analyses of nuclei preselected with fluorescence microscopy. It is the first such study of nuclear division in theTulasnellaceae and the first of conjugately dividing nuclei in basidiomycetous hyphal segments lacking clamp connections. Mitosis inT. araneosa is unusual in that the spindle pole body (SPB) develops asymmetrically; the SPB middle piece is large and transversely curved; and the nuclear envelopes of adjacent late anaphase nuclei fuse. Analyses of mitotic characteristics used for phylogenetic purposes indicate that, of the many characters available, only SPB characteristics are presently valuable. Available evidence indicates that the SPB ofT. araneosa is more different from that ofUredinales than it is from representatives of the other four orders ofBasidiomycotina that have been thoroughly studied.     

Microtubules and actin cytoskeleton in Cryptococcus neoformans compared with ascomycetous budding and fission yeasts

Actin cytoskeleton and microtubules were studied in a human fungal pathogen, the basidiomycetous yeast Cryptococcus neoformans (haploid phase of Filobasidiella neoformans), during its asexual reproduction by budding using fluorescence and electron microscopy. Staining with rhodamine-conjugated phalloidin revealed an F-actin cytoskeleton consisting of cortical patches, cables and cytokinetic ring. F-actin patches accumulated at the regions of cell wall growth, i. e. in sterigma, bud and septum. In mother cells evenly distributed F-actin patches were joined to F-actin cables, which were directed to the growing sterigma and bud. Some F-actin cables were associated with the cell nucleus. The F-actin cytokinetic ring was located in the bud neck, where the septum originated. Antitubulin TAT1 antibody revealed a microtubular cytoskeleton consisting of cytoplasmic and spindle microtubules. In interphase cells cytoplasmic microtubules pointed to the growing sterigma and bud. As the nucleus was translocated to the bud for mitosis, the cytoplasmic microtubules disassembled and were replaced by a short intranuclear spindle. Astral microtubules then emanated from the spindle poles. Elongation of the mitotic spindle from bud to mother cell preceded nuclear division, followed by cytokinesis (septum formation in the bud neck). Electron microscopy of ultrathin sections of chemically fixed and freeze-substituted cells revealed filamentous bundles directed to the cell cortex. The bundles corresponded in width to the actin microfilament cables. At the bud neck numerous ribosomes accumulated before septum synthesis. We conclude: (i) the topology of F-actin patches, cables and rings in C. neoformans resembles ascomycetous budding yeast Saccharomyces, while the arrangement of interphase and mitotic microtubules resembles ascomycetous fission yeast Schizosaccharomyces. The organization of the cytoskeleton of the mitotic nucleus, however, is characteristic of basidiomycetous yeasts. (ii) A specific feature of C. neoformans was the formation of a cylindrical sterigma, characterized by invasion of F-actin cables and microtubules, followed by accumulation of F-actin patches around its terminal region resulting in development of an isodiametrical bud.     

Microtubular and actin cytoskeletons and ultrastructural characteristics of the potentially pathogenic basidiomycetous yeast Malassezia pachydermatis

Microtubular and actin cytoskeletons were investigated in the lipophilic yeast Malassezia pachydermatis by fluorescence and electron microscopy. To detect microtubules by indirect immunofluorescence using monoclonal anti-tubulin antibody, a prolonged incubation with lysing enzymes was necessary due to its very thick cell wall. Cytoplasmic microtubules were detected in interphase and a spindle with astral microtubules was seen in M-phase. The disintegration of cytoplasmic microtubules and migration of the nucleus to the bud before mitosis were characteristic features of the basidiomycetous yeast Malassezia pachydermatis. The visualisation of F-actin structures (patches, cables and cytokinetic rings) by fluorescence microscopy using both monoclonal anti-actin antibody and rhodamine-phalloidin failed, but actin was detected by electron microscopy with immunogold labelling. Clusters of gold particles indicating actin structures were detected at the plasma membrane of cells with unique cortical ultrastructural features characteristic of the genus Malassezia. A possible association of these with the actin cytoskeleton is suggested.     

我国各类基物上的酵母菌分布及其尿素酶活性

本文讨论了2300株由我国各地分离到的酵母菌和类酵母菌与各类基物的关系。共述及23属,占Lodder(1970)系统的半数以上。在十几类基物中酵母菌的分布各有不同,叶子上有大量掷孢酵母,果实上比其它基物更能获得克勒克酵母属(Kloeckera),在咸腌食品中相对地有较多耐高渗透压的酵母如球拟酵母属(Torulopsis)与德巴利酵母属(Debaryomyces)。对部分属的定种工作也已进行,说明某些属的种在我国均有发现而且为数不少。此外还研究了600余株代表各类酵母的尿素酶活性,结果说明担子菌酵母或系统上接近担子菌的酵母都有尿素酶活性,而子囊菌酵母则无。根据作者的观察子囊菌酵母中裂殖酵母属(Schizosaccharomyces)则常常例外地有尿素酶反应,且常有辅酶Q_(10),从而可见裂殖酵母属有其特有的生物学特性,在系统上很可能与其它子囊菌酵母不同。对属于不完全菌的无孢子酵母也进行了尿素酶活性测定,结果表明它们部分与子囊菌有关,部分与担子菌有关。     

Reduction in amounts of mitochondrial DNA in the sperm cells as a mechanism for maternal inheritance in Hordeum vulgare

It is known that extranuclear organelle DNA is inherited maternally in the majority of angiosperms. The mechanisms for maternal inheritance have been well studied in plastids but not in mitochondria. In the present study we examined the mitochondrial DNA in the male reproductive cells of Hordeum vulgare L. by immunoelectron microscopy. Our results show that the number of anti-DNA gold particles on sections of sperm cell mitochondria decreased by 97% during pollen development. The reduction occurred rapidly in the generative cells and subsequently in the sperm cells, concomitant with a remarkable reduction in mitochondrial volume. It seems that the copy numbers of mitochondrial DNA were reduced in the male reproductive cells, which may be a possible mechanism by which paternal transmission is inhibited. Unlike mitochondria, plastids are excluded from the generative cells during the first pollen mitosis. These data suggest a mechanism for maternal inheritance of mitochondria in angiosperms and for independent control of inheritance of mitochondria and plastids in H. vulgare.     

Cytoskeletal structures,ultrastructural characteristics and the capsule of the basidiomycetous yeast <Emphasis Type="Italic">Cryptococcus laurentii</Emphasis>

The cytoskeleton, capsule and cell ultrastructure were studied during the cell cycle of Cryptococcus laurentii. In an encapsulated strain, cytoplasmic microtubules and a mitotic spindle were detected. Mitosis was preceded by migration of the nucleus into the bud. F-actin failed to be visualised by rhodamine-phalloidin (RhPh) in encapsulated cells and therefore an acapsular strain was used. The following actin structures were found: actin dots, actin cables and cytokinetic ring. Ultrastructural studies showed the presence of a nucleus in the bud before mitosis. A collar-shaped structure was seen at the base of bud emergence. A lamellar cell wall and a rough outer surface of the cells were detected. Cytoskeletal structures found in C. laurentii are similar to those in Cryptococcus neoformans, which is a serious human pathogen.     

Cell-cycle checkpoints that ensure coordination between nuclear and cytoplasmic events in Saccharomyces cerevisiae

Cytoskeletal organization is crucial for several aspects of cell-cycle progression but cytoskeletal elements are quite sensitive to environmental perturbations. Two novel checkpoint controls monitor the function of the actin and microtubule systems in budding yeast and operate to delay cell-cycle progression in response to cytoskeletal perturbations. In cells whose actin cytoskeleton has been perturbed, bud formation is frequently delayed and the morphogenesis checkpoint introduces a compensatory delay of nuclear division until a bud has been formed. In cells whose microtubule cytoskeleton has been perturbed, anaphase spindle elongation often occurs entirely within the mother cell, and the post-anaphase nuclear migration checkpoint introduces a compensatory delay of cytokinesis until one pole of the anaphase nucleus enters the bud. Recent studies indicate that regulators of entry into mitosis are localized to the daughter side of the mother-bud neck whereas regulators of exit from mitosis are localized to the spindle pole bodies. Thus, specific cell-cycle regulators are well-placed to monitor whether a cell has formed a bud and whether a daughter nucleus has been delivered accurately to the bud following mitosis.     

Nucleocytoplasmic protein translocation during mitosis in the social amoebozoan Dictyostelium discoideum

Mitosis is a fundamental and essential life process. It underlies the duplication and survival of all cells and, as a result, all eukaryotic organisms. Since uncontrolled mitosis is a dreaded component of many cancers, a full understanding of the process is critical. Evolution has led to the existence of three types of mitosis: closed, open, and semi‐open. The significance of these different mitotic species, how they can lead to a full understanding of the critical events that underlie the asexual duplication of all cells, and how they may generate new insights into controlling unregulated cell division remains to be determined. The eukaryotic microbe Dictyostelium discoideum has proved to be a valuable biomedical model organism. While it appears to utilize closed mitosis, a review of the literature suggests that it possesses a form of mitosis that lies in the middle between truly open and fully closed mitosis—it utilizes a form of semi‐open mitosis. Here, the nucleocytoplasmic translocation patterns of the proteins that have been studied during mitosis in the social amoebozoan D. discoideum are detailed followed by a discussion of how some of them provide support for the hypothesis of semi‐open mitosis.     

Bud morphogenesis and the actin and microtubule cytoskeletons during budding in the corn smut fungus,Ustilago maydis

Ustilago maydis is a dimorphic Basidiomycete fungus with a yeast-like form and a hyphal form. Here we present a comprehensive analysis of bud formation and the actin and microtubule cytoskeletons of the yeast-like form during the cell cycle. We show that bud morphogenesis entails a series of shape changes, initially a tubular or conical structure, culminating in a cigar-shaped cell connected to the mother cell by a narrow neck. Labelling of cells with concanavalin A demonstrated that growth occurs at bud tip. Indirect immunofluorescence studies revealed that the actin cytoskeleton consists of patches and cables that polarize to the presumptive bud site and the bud tip and an actin ring that forms at the neck region. Because the bud tip corresponds to the site of active cell wall growth, we hypothesize that actin is involved in secretion of cell wall components. The microtubule cytoskeleton has recently been shown to consist of a cytoplasmic network during interphase that disassembles at mitosis when a spindle and astral microtubules are formed. We have carried out studies of U. maydis cells synchronized by the microtubule-depolymerizing drug thiabendazole which allow us to construct a temporal sequence of steps in spindle formation and spindle elongation during the cell cycle. These studies suggest that astral microtubules may be involved in early stages of spindle orientation and migration of the nucleus into the bud and that the spindle pole bodies may be involved in reestablishment of the cytoplasmic microtubule network.     

Breaking and making of the nuclear envelope

During mitosis, a single nucleus gives rise to two nuclei that are identical to the parent nucleus. Mitosis consists of a continuous sequence of events that must be carried out once and only once. Two such important events are the disassembly of the nuclear envelope (NE) during the first stages of mitosis, and its accurate reassembly during the last stages of mitosis. NE breakdown (NEBD) is initiated when maturation-promoting factor (MPF) enters the nucleus and starts phosphorylating nuclear pore complexes (NPCs) and nuclear lamina proteins, followed by NPC and lamina breakdown. Nuclear reassembly starts when nuclear membranes assemble onto the chromatin. This article focuses on the different models of NEBD and reassembly with emphasis on recent data.     

Saccharomyces cerevisiae Cdc42p localizes to cellular membranes and clusters at sites of polarized growth

The Cdc42p GTPase controls polarized growth and cell cycle progression in eukaryotes from yeasts to mammals, and its precise subcellular localization is essential for its function. To examine the cell cycle-specific targeting of Cdc42p in living yeast cells, a green fluorescent protein (GFP)-Cdc42 fusion protein was used. In contrast to previous immunolocalization data, GFP-Cdc42p was found at the plasma membrane around the entire cell periphery and at internal vacuolar and nuclear membranes throughout the cell cycle, and it accumulated or clustered at polarized growth sites, including incipient bud sites and mother-bud neck regions. These studies also showed that C-terminal CAAX and polylysine domains were sufficient for membrane localization but not for clustering. Time-lapse fluorescence microscopy showed that GFP-Cdc42p clustered at the incipient bud site prior to bud emergence and at the mother-bud neck region postanaphase as a diffuse, single band and persisted as two distinct bands on mother and daughter cells following cytokinesis and cell separation. Initial clustering occurred immediately prior to actomyosin ring contraction and persisted postcontraction. These results suggest that Cdc42p targeting occurs through a novel mechanism of membrane localization followed by cell cycle-specific clustering at polarized growth sites.     

THE ONSET OF MITOSIS AND DNA SYNTHESIS IN ROOTS OF GERMINATING BEANS

The time of onset of mitosis and DNA synthesis has been determined in roots of germinating seeds of Vicia faba. Mitosis is not initiated in all roots simultaneously. Dividing cells are seen 36 hr from the beginning of germination, but they are present in low frequency (0.02%). Dividing cells do not become frequent, i.e., occurring as 5% or more of all cells, until 56 hr, and it is not until 66–68 hr that all roots in a sample of 10 are mitotically active. DNA synthesis shows a similar sporadic beginning. It occurs in a few cells by 28 hr, and by 40 hr all roots exposed to ³H–thymidine show active incorporation. For most cells in these germinating roots DNA synthesis precedes mitosis. In one root in 10, however, some cells are unlabeled when they enter mitosis, indicating that they had spent the dormant period in the G₂ phase of the mitotic cycle. The presence of these cells determines whether or not roots show chromatid and chromosome aberrations following irradiation during germination.     

感染环形泰勒焦虫(Theileria annulata)的牛外周血白细胞的形态观察

用光镜及扫描电镜观察了体外高代培养的含牛焦虫颗粒的牛外周血白细胞的形态及在细胞周期中细胞表面的特征性变化。这种经多年传代的含虫的牛外周血白细胞恢复了分裂和繁殖的能力,目前已成为较稳定的细胞系。细胞表面具多种伪足突起,如叶状、丝状及绒毛状。细胞周期中备期细胞表面的主要特征是:S期:细胞平扁,边缘具薄的时状伪足及丝状伪足;G_2期:细胞中部隆起,表面具少量绒毛状伪足;G_1期:绒毛状结构少或无,而出现丝状及小的叶状伪足,细胞仍保持球形;M期:细胞球形,表面密被以绒毛。作者根据扫描电镜的观察认为光镜下所观察的两类细胞,实际上是反映了一种细胞处于不同发育阶段时的特征。     

Immunofluorescence study of mitosis and cytokinesis in Acrosiphonia duriuscula (Acrosiphoniales,Chlorophyta)*

The behaviors of nuclei and microtubules (MT) in Acrosiphonia duriuscula (Ruprecht) Collins were observed in detail using fluorescence and electron microscopy. Numerous nuclei exist in cells of A. duriuscula (multinucleate cells). Cortical MT radiate from the apex of the tip cell and run parallel to its long axis. Between 30 and 40% of nuclei in the upper part of cytoplasm migrate downward to the region where cytokinesis will take place, and these numerous nuclei form a ‘nuclear ring’ before mitosis. The parallel array of the cortical MT changes to a transverse orientation at the region where cytokinesis will take place, and finally forms a characteristic circumferential band. Mitosis starts from the nuclei in the ring. Cortical MT disappear in the region of the nuclear ring and many mitotic spindles form. The band-shaped array of MT remains. Mitosis spreads in an apparent wave to the other nuclei. After mitosis, daughter nuclei that formed a nuclear ring migrate apically and repopulate the apical daughter cell. When the numerous daughter nuclei have relocated, a rearrangement of the cortical MT occurs. They are randomly arranged at first, but finally become parallel to the long axis of the cell. Cytokinesis occurs by furrowing of the cell, and the band-shaped array of MT could be detected at the leading edge of the furrow.     

Sorosporium consanguineum: Relation between variable nuclear condition and dissociation

Saprophytic development of Sorosporium consanguineum and its nuclear cycle were studied on laboratory media. During vegetative reproduction, the nuclei of the monokaryotic sporidia were shown to first migrate into developing bud cells where division occurs, one nucleus returning to the parent cell prior to completion of cell division. Following fusion of sporidia of opposite mating type and subsequent formation of infection hyphae, dikaryons eventually dissociated, giving rise to sporidia of both sex groups in the process. As a result of dissociation, shown to occur in several ways, "satellite" sporidial colonies characteristically formed in advance of the parent colony and fusions again occurred between sporidia of opposite mating type. Reports of variable nuclear condition of mycelia in other species of smut fungi in culture are discussed in light of these findings.     

An electron microscopic study of bud development in Saccharomycodes ludwigii and Saccharomyces cerevisiae

Summary Examination of sectioned cells fixed in KMnO₄ has shown that the wall of the first bud of a cell of Saccharomycodes ludwigii arises as an extension of the main wall of the parent, while in subsequent buds it develops by extension of the half-septum remaining at a previous detachment scar. Septa are formed by the deposition of wall material on each side of an electron transparent plate which develops centripetally. Structural changes occur in the marginal region of the septum prior to rupture of the main wall and the separation of cells at the surface of the septum-plate. The broken walls remain as annular rings around the scars following the successive development of buds at both apices of the cell.In Saccharomyces cerevisiae the bud wall arises as a direct extension of the parent wall or as an extension of an additional inner layer developed locally.The two types of bud origin are compared in the two yeasts and a comparison is also made with the development of buds, fission cells, conidia and germ tubes in other organisms.     

Effect of HCG on the interstitial cells and androgen production in the immature rat testis.

The effect on the interstitial cells in the immature rat testis of administration of HCG for different periods was correlated with testosterone plasma levels. Significant and progressive stimulation of mitosis was observed after 3 days of HCG treatment but stabilization occurred after 5 days. The numbers of precursor fibroblasts had increased by the 5th day and were still increasing by the 10th day of treatment. Numbers of Leydig cells were significantly greater at 5 and 10 days of treatment. Plasma testosterone showed a progressive and continuous increase in all groups. The increase in Leydig cell number is considered to be due to a combination of increased stimulation of mitoses in Leydig cells and differentiation of precursor fibroblasts. Mitosis seems to precede fibroblastic differentiation, but the latter continues when mitotic changes have stabilized. The elevation of plasma testosterone concentrations is probably due firstly to the stimulation of the existing Leydig cells and then to the increase in the number of hormone-secreting cells.     

Fission yeast cells undergo nuclear division in the absence of spindle microtubules

Mitosis in eukaryotic cells employs spindle microtubules to drive accurate chromosome segregation at cell division. Cells lacking spindle microtubules arrest in mitosis due to a spindle checkpoint that delays mitotic progression until all chromosomes have achieved stable bipolar attachment to spindle microtubules. In fission yeast, mitosis occurs within an intact nuclear membrane with the mitotic spindle elongating between the spindle pole bodies. We show here that in fission yeast interference with mitotic spindle formation delays mitosis only briefly and cells proceed to an unusual nuclear division process we term nuclear fission, during which cells perform some chromosome segregation and efficiently enter S-phase of the next cell cycle. Nuclear fission is blocked if spindle pole body maturation or sister chromatid separation cannot take place or if actin polymerization is inhibited. We suggest that this process exhibits vestiges of a primitive nuclear division process independent of spindle microtubules, possibly reflecting an evolutionary intermediate state between bacterial and Archeal chromosome segregation where the nucleoid divides without a spindle and a microtubule spindle-based eukaryotic mitosis.