Occurrence of Mitochondria in the Nuclei of Tobacco Sperm Cells

Tobacco sperm cells contain intact mitochondria within their nuclei with a frequency of 0.35 [plusmn] 0.13 per cell. These inclusions appear to originate from mitochondria found among chromatids in the highly elongated metaphase plate of the dividing generative cell. These organelles are apparently captured during the reconstitution of the nuclear envelope. Only sperm cells were observed to contain these nuclear mitochondria; generative cells, vegetative pollen cells, transmitting tissue cells, unfertilized egg cells, and central cells lacked them. Nuclear mitochondria were also seen in the nuclei of the egg and central cell after fusion with sperm nuclei, suggesting that nuclear mitochondria are transmitted into the zygote and primary endosperm cells during double fertilization. Organellar inclusions in the sperm nucleus provide a potential mechanism for transmitting organellar DNA into the next generation and could potentially facilitate the transfer of genetic material between the nucleus and other organelles.     

FROM PROTOPLASM TO SWARMER: REGENERATION OF PROTOPLASTS FROM DISINTEGRATED CELLS OF THE MULTICELLULAR MARINE GREEN ALGA MICRODICTYON UMBILICATUM (CHLOROPHYTA)1

Protoplast regeneration from extruded cytoplasm of the multicellular marine green alga Microdictyon umbilicatum (Velley) Zanardini (Cladophorales, Anadyomenaceae) was investigated. The early process of protoplast formation is comprised of two steps: agglutination of cell organelles into protoplasmic masses followed by generation of a temporary enclosing envelope around them. Agglutination of cell organelles was mediated by a lectin–carbohydrate complementary system. Three sugars, D‐galactosamine, D‐glucosamine, and α‐D‐mannose, inhibited the agglutination process, and three complementary lectins for the above sugars, peanut agglutinin, Ricinus communis agglutinin, and concanavalin A, bound to the surfaces of chloroplasts. Agglutination assay using human erythrocytes showed the presence of lectins specific for the above sugars in the algal vacuolar sap. A fluorescent probe 1‐(4‐trimethylammoniumphenyl)‐6‐phenyl‐a, 3,5‐hexatriene revealed that the envelope initially surrounding protoplasts was not a lipid‐based cell membrane. However, this developed several hours later. Simultaneous fluorescein diacetate and propidium iodide staining showed that the primary envelope had some characteristics of cell membranes, such as semipermeability and selective transport of materials. Also, fluorescein diacetate staining showed esterase activity in the protoplast and relocation of cell organelles and compartmentalization of cytoplasm during the process of regeneration. Both pH 7–9 and salinity 400–500 mM were found to be essentially important for the development of the protoplast envelope. When the basic regeneration process was accomplished, two alternative pathways of development were seen; about 70% of one‐celled protoplasts transformed into reproductive cells within 2 weeks after wounding, whereas others began cell division and grew into typical Microdictyon thalli. Quadriflagellate swarmers were liberated from the reproductive cells, and they germinated into mature individuals. It is therefore suggested that this species may use the wound response as a method of propagation and dispersal.     

On the mechanism of rapid plasma membrane and chloroplast envelope expansion in Dunaliella salina exposed to hypoosmotic shock

Dunaliella salina cells rapidly diluted from their normal 1.71 M NaCl-containing growth medium into medium containing 0.86 M NaCl swelled within 2--4 min to an average volume 1.76 X larger and a surface area 1.53 X larger than found in control cells. Morphometric analysis of thin section electron micrographs revealed that certain organelles, including the chloroplast, nucleus, and some types of vacuoles, also expanded in surface area as much or more than did the entire cell. It is likely that glycerol, the most important osmotically active intracellular solute, was present in high concentration within these organelles as well as in the cytoplasm itself. Thin section and freeze-fracture electron microscopy were utilized to trace the origin of membrane material whose addition permitted the large increase in plasma membrane surface area and the equally large growth of the chloroplast outer envelope. The findings indicated that the plasma membrane's expansion resulted from its selective fusion with numerous small (less than or equal to 0.25 micron diam) vesicles prevalent throughout the cytoplasm. In contrast, new membrane added to the chloroplast outer envelope was drawn from an entirely different source, namely, elements of the endoplasmic reticulum.     

Plastids and stromules interact with the nucleus and cell membrane in vascular plants

The various metabolic activities of plastids require continuous exchange of reactants and products with other organelles of the plant cell. Physical interactions between plastids and other organelles might therefore enhance the efficiency of plant metabolism. We have observed a close apposition of plastids and nuclei in various organs of Nicotiana tabacum and Arabidopsis thaliana. In hypocotyl epidermal cells, plastids and stromules, stroma-filled tubular extensions of the plastid envelope membrane, were observed to reside in grooves and infoldings of the nuclear envelope, indicating a high level of contact between the two organelle membranes. In a number of non-green tissues, including suspension-cultured cells, perinuclear plastids were frequently associated with long stromules that extended from the cell center to the cell membrane. In cotyledon petioles, cells lying adjacent to one another frequently contained stromules that met on either side of the shared cell wall, suggesting a means of intercellular communication. Our results therefore suggest that stromules have diverse roles within plant cells, perhaps serving as pathways between nuclei and more distant regions of the cell and possibly even other cells.     

The regulated secretory pathway in CD4(+) T cells contributes to human immunodeficiency virus type-1 cell-to-cell spread at the virological synapse

Direct cell-cell spread of Human Immunodeficiency Virus type-1 (HIV-1) at the virological synapse (VS) is an efficient mode of dissemination between CD4(+) T cells but the mechanisms by which HIV-1 proteins are directed towards intercellular contacts is unclear. We have used confocal microscopy and electron tomography coupled with functional virology and cell biology of primary CD4(+) T cells from normal individuals and patients with Chediak-Higashi Syndrome and report that the HIV-1 VS displays a regulated secretion phenotype that shares features with polarized secretion at the T cell immunological synapse (IS). Cell-cell contact at the VS re-orientates the microtubule organizing center (MTOC) and organelles within the HIV-1-infected T cell towards the engaged target T cell, concomitant with polarization of viral proteins. Directed secretion of proteins at the T cell IS requires specialized organelles termed secretory lysosomes (SL) and we show that the HIV-1 envelope glycoprotein (Env) localizes with CTLA-4 and FasL in SL-related compartments and at the VS. Finally, CD4(+) T cells that are disabled for regulated secretion are less able to support productive cell-to-cell HIV-1 spread. We propose that HIV-1 hijacks the regulated secretory pathway of CD4(+) T cells to enhance its dissemination.     

南方鲶卵巢滤泡细胞和卵膜生成的组织学研究

南方鲶的卵巢滤泡细胞源于卵巢基质细胞,从发生到退化分为零散卵泡膜细胞期、单层扁平泡膜细胞期、多层扁平卵泡膜细胞期、立方形颗粒细胞期柱状颗粒细胞期、颗粒细胞分泌期和颗粒细胞退化期。精孔细胞中发育中滤泡细胞分化形成。初级卵精源于卵母细胞,次级卵膜由晚期滤泡细胞分泌形成。本文还对滤泡细胞和卵膜的作用进行了阐述。     

Nuclear migration: rock and roll facilitated by dynein and kinesin

The nucleus encounters other organelles as well as high cytoplasmic pressures during its migration within the cell. A new study describes how the action of kinesin and dynein motors is coordinated at the nuclear envelope to rock and roll the nucleus in Caenorhabditis elegans.     

AN ELECTRON MICROSCOPE STUDY OF VITALLY STAINED SINGLE CELLS IRRADIATED WITH A RUBY LASER MICROBEAM

An electron microscope study has been made of vitally stained single cells whose cytoplasm has been subjected to a localized ruby laser microbeam. Light and moderate laser absorption (the resultant of stain concentration and laser energy density) produced restricted selective damage of mitochondria in cells stained with Janus green B; heavy laser absorption resulted in mitochondrial damage, as well as in nonselective interaction with other cell structures. With four other basic vital stains, the polysomes, ergastoplasm, mitochondria and other organelles at the irradiated site were uniformly damaged. Unstained cells showed no morphological alterations. With light primary damage (that restricted to the irradiation site), no secondary effects of the incident radiation were observed. With moderate primary damage, however, secondary damage of the mitochondria in the unirradiated cell portions was produced, which was reversible within 4 hr after irradiation. Heavy primary lesions caused severe secondary alteration of all cell structures that was irreversible and cell death occurred within 2 hr. Surviving cells examined 24 hr after light and moderate irradiation could not be distinguished from unirradiated controls. The possible mechanisms involved in the production of laser-induced cellular alterations are discussed.     

Assembly of the protoplasm of Codium fragile (Bryopsidales, Chlorophyta) into new protoplasts

The cell organelles of the coenocytic alga Codium fragile (Sun) Hariot aggregated rapidly and protoplasts were formed when its protoplasm was extruded out in seawater. Continuous observation showed that there were long and gelatinous threads connecting the cell organelles. The threads contracted, and thus the cell organelles aggregated into protoplasmic masses. The enzyme digestion experiments and Coomassie Brilliant Blue and Anthrone stainings showed that the long and gelatinous threads involved in the formation of the protoplasts might Include protein and saccharides as structure components. Nile Red staining Indicated that the protoplast primary envelope was non-lipid at first, and then lipid materials Integrated Into its surface gradually. The fluorescent brightener staining Indicated that the cell wall did not regenerate in the newly formed protoplasts and they all disintegrated within 72 h after formation. Transmission electron microscopy of the cell wall of wild C. fragile showed electron-dense material embedded in the whole cell wall at regular intervals. The experiments indicated that C. fragile would be a suitable model alga for studying the formation of protoplasts.     

Cloning and characterization of human immunodeficiency virus type 1 variants diminished in the ability to induce syncytium-independent cytolysis.

The phenomenon of interference was exploited to isolate low-abundance noncytopathic human immunodeficiency virus type 1 (HIV-1) variants from a primary HIV-1 isolate from an asymptomatic HIV-1-seropositive hemophiliac. Successive rounds of virus infection of a cytolysis-susceptible CD4+ cell line and isolation of surviving cells resulted in selective amplification of an HIV-1 variant reduced in the ability to induce cytolysis. The presence of a PvuII polymorphism facilitated subsequent amplification and cloning of cytopathic and noncytopathic HIV-1 variants from the primary isolate. Cloned virus stocks from cytopathic and noncytopathic variants exhibited similar replication kinetics, infectivity, and syncytium induction in susceptible host cells. The noncytopathic HIV-1 variant was unable, however, to induce single-cell killing in susceptible host cells. Construction of viral hybrids in which regions of cytopathic and noncytopathic variants were exchanged indicated that determinants for the noncytopathic phenotype map to the envelope glycoprotein. Sequence analysis of the envelope coding regions indicated the absence of two highly conserved N-linked glycosylation sites in the noncytopathic HIV-1 variant, which accompanied differences in processing of precursor gp160 envelope glycoprotein. These results demonstrate that determinants for syncytium-independent single-cell killing are located within the envelope glycoprotein and suggest that single-cell killing is profoundly influenced by alterations in envelope sequence which affect posttranslational processing of HIV-1 envelope glycoprotein within the infected cell.     

进化细胞生物学的提出及其任务

作者提出应创建一门源于进化生物学与细胞生物学两者的交叉学科一进化细胞生物学(细胞的进化生物学)。其根本任务在于用进化的观点考察真核细胞的一切方面,从它们的起源和演化来认识它们的现在。文中列举了其具体的研究内容,并分析了其研究方法上的特点,指出在这里需要把进化生物学的综合性分析与细胞生物学的实验研究最紧密地结合起来。文中还论述了真核细胞的细胞器的“不进化”现象,指出其根本原因在于进化焦点的转移。     

Programmed Cell Death During the Vessel Element Differentiation of the Secondary Xylem in Eucommia ulmoides Shoots

The result from in situ end-labelling of fragmented DNA indicated that the vessel element differentiation of the secondary xylem in Eucommia ulmoides Oliv. was a typical programmed cell death (PCD) which involved a series of events, viz. synthesis of components essential for the secondary wall formation and a well organized succession of protoplast degeneration and autolysis in the tracheary cells. The nuclei gradually became irregular with highly condensed chromatin. In some nuclei, the cistema of the nuclear envelope became unevenly dilated within which some inner membrane protrusion enclosed with nuclear materials were present. The nuclear envelope underwent disruption and the nucleus eventually degenerated. However, as the nucleus was one of the most stable components in the cell, it was among the last organelles disappeared during the autolytic process. In the process, there were two forms of degeneration in the mitochondria (Mit). In one form the Mit shrank and became disorganized; in the other, part of the matrix in the Mit became electron-lucent with breakage of the membrane nearby. The cytoplasmic component residues were phagocitized and sequestered by the dilated rough endoplasmic reticulum (RER) cisternae. The RER and vacuoles did play a vital role in the further degeneration of other organelles just similar to the lysosomes acting in the animal cells. The autolyzed debri might be utilized in situ by taking part in the formation of secondary wall or be transported to the adjacent cells through the pits.     

Distribution of microtubules in prolactin cells of lactating rats

The intracellular distribution of microtubules was studied using serial sections of prolactin cells in anterior pituitary glands from lactating rats. Numerous microtubules were present in these cells following fixation with glutaraldehyde and osmium tetroxide. The greatest number of microtubules were present in the Golgi complex, situated around the perimeter and in association with the cisternae, vesicles and developing secretory granules. Microtubules were found in channels between groups of parallel cisternae of rough surfaced endoplasmic reticulum and in close proximity to small vesicles. They were also located adjacent to mitochondria, the plasmalemma, the nuclear envelope, and among mature secretory granules. Due to their orientation within the cell, it is suggested that the microtubules may act to direct the movement of organelles from one region of the cell to another and to give internal support to the cell.     

Detection of intracellular phosphatidylserine in living cells

To demonstrate the intracellular phosphatidylserine (PS) distribution in neuronal cells, neuroblastoma cells and hippocampal neurons expressing green fluorescence protein (GFP)-AnnexinV were stimulated with a calcium ionophore and localization of GFP-AnnexinV was monitored by fluorescence microscopy. Initially, GFP-AnnexinV distributed evenly in the cytosol and nucleus. Raising the intracellular calcium level with ionomycin-induced translocation of cytoplasmic GFP-AnnexinV to the plasma membrane but not to the nuclear membrane, indicating that PS distributes in the cytoplasmic side of the plasma membrane. Nuclear GFP-AnnexinV subsequently translocated to the nuclear membrane, indicating PS localization in the nuclear envelope. GFP-AnnexinV also localized in a juxtanuclear organelle that was identified as the recycling endosome. However, minimal fluorescence was detected in any other subcellular organelles including mitochondria, endoplasmic reticulum, Golgi complex, and lysosomes, strongly suggesting that PS distribution in the cytoplasmic face in these organelles is negligible. Similarly, in hippocampal primary neurons PS distributed in the inner leaflet of plasma membranes of cell body and dendrites, and in the nuclear envelope. To our knowledge, this is the first demonstration of intracellular PS localization in living cells, providing an insight for specific sites of PS interaction with soluble proteins involved in signaling processes.     

A new type of microtubular cytoskeleton in microsporogenesis of Lavatera thuringiaca L.

Summary. In Lavatera thuringiaca, kariokinesis and simultaneous cytokinesis during the meiotic division of microsporogenesis follow a procedure similar to that which takes place in the majority of members of the class Angiospermae. However, chondriokinesis occurs in a unique way found only in species from the family Malvaceae. Chondriokinesis in such species is well documented, but the relationship between the tubulin cytoskeleton and rearrangement of cell organelles during meiosis in L. thuringiaca has not been precisely defined so far. In this study, the microtubular cytoskeleton was investigated in dividing microsporocytes of L. thuringiaca by immunofluorescence. The meiotic stages and positions of cell organelles were identified by staining with 4′,6-diamidino-2-phenylindole. We observed that, during prophase I and II, changes in microtubular cytoskeleton configurations have unique features, which have not been described for other plant species. At the end of prophase I, organelles (mostly plastids and mitochondria) form a compact envelope around the nucleus, and the subsequent phases of kariokinesis take place within this arrangement. At this point of cell division, microtubules surround the organelle envelope and separate it from the peripheral cytoplasm, which is devoid of plastids and mitochondria. In telophase I, two newly formed nuclei are tightly surrounded by the cell organelle envelopes, and these are separated by the phragmoplast. Later, when the phragmoplast disappears, cell organelles still surround the nuclei but also move a little, starting to occupy the place of the disappearing phragmoplast. After the breakup of tetrads, the radial microtubule system is well developed, and cell organelles can still be observed as a dense envelope around the nuclei. At a very late stage of sporoderm development, the radial microtubule system disappears, and cell organelles become gradually scattered in the cytoplasm of the microspores. Using colchicines, specific inhibitors of microtubule formation, we investigated the relationship between the tubulin cytoskeleton and the distribution of cell organelles. Our analysis demonstrates that impairment of microtubule organization, which constitutes only a single component of the cytoskeleton, is enough to disturb typical chondriokinesis in L. thuringiaca. This indicates that microtubules (independent of microfilaments) are responsible for the reorganization of cell organelles during meiotic division. Correspondence: D. Tchórzewska, Department of Plant Anatomy and Cytology, Maria Curie-Skłodowska University, Akademicka 19, 20-033 Lublin, Poland.     

FURTHER OBSERVATIONS ON THE PHENOMENON OF SECONDARY VACUOLATION IN LIVING CELLS

Invagination of the plasma membrane in plant cells forms peripheral or endocytic structures which often contain a complement of membrane-bound vesicles. These structures, or secondary vacuoles, move with the streaming cytoplasm although their velocities are somewhat slower than that for the various organelles within the cytoplasm. They glide over the nucleus or flow from the peripheral cytoplasm onto a transvacuolar strand and continue unabated along the length of a strand. These structures may detach from the plasma membrane as sacs to become positioned in the cytoplasm directly under the tonoplast and project into the primary vacuole. Some endocytic vacuoles may separate from the peripheral cytoplasm and remain free within the primary vacuole; subsequently they can re-associate with the cytoplasm. While the content and function of these vacuoles are yet to be determined, indirect evidence indicates that they are pinocytic in character since the content of an invagination is confined to the sac upon its detachment from the plasma membrane and is subsequently transported throughout the cell by cyclosis.     

杜仲次生木质部导管分子分化中的程序性死亡

运用原位末端标记法,证明了杜仲（ＥｕｃｏｍｍｉａｕｌｍｏｉｄｅｓＯｌｉｖ．）次生木质部的导管分子分化过程是程序性死亡（ｐｒｏｇｒａｍｍｅｄｃｅｌｄｅａｔｈ,ＰＣＤ）过程。它包括一系列的物质合成和细胞结构的解体和自溶：细胞核逐步变得极不规则,染色质高度凝集,有些细胞核的核周腔膨大,核周腔内存在着包裹有核物质的内膜凸起,最后,核膜消失,核解体。细胞核是ＰＣＤ中最后消失的细胞器之一。线粒体有两种解体方式,一种为线粒体内部结构紊乱,嵴极不清晰,线粒体皱缩变形;另一种为线粒体出现一些电子密度低的透明区,进而裂开。内质网囊泡化,相邻的内质网槽库之间互相连接,形成分隔。内质网和液泡共同行使溶酶体功能,吞噬各种细胞器残体。解体后的物质可能有两种去向,一是通过纹孔运输到邻近的细胞中,二是转化成构建自身次生壁的物质     

Ultrastructure of vacuolar inclusions in root tips

Summary Vacuoles in plant cells often contain inclusions which at early stages of development are bounded by a single membrane. The inclusion bodies (IBs) comprise a diversity of forms and various stages of differentiation are recognizable. IBs are divided into two categories: those which have a matrix without internal membranes, and those which contain cytoplasmic organelles and other membranous material. The internal membranes may be tightly coiled or in the form of vesicles. IBs develop from invaginations of the tonoplast which become detached into the vacuole. They are initiated mainly during active cell growth but may remain within the vacuole in differentiated cells. Various components contribute to the contents of IBs: endoplasmic reticulum, nuclear envelope, Golgi vesicles, extruded portions of mitochondria and plastids, ribosomes and groundplasm. In most IBs the limiting membrane and contents eventually disappear within the vacuole. Some IBs prior to their breakdown within the vacuole also function as sites for the formation of material not found elsewhere in the cell. The disappearance of IBs from vacuoles suggests that such vacuoles behave as lysosomes.     

PARTICIPATION OF THE CYTOPLASMIC MEMBRANE IN THE GROWTH AND SPORE FORMATION OF BACILLI

A polyester embedding technique was used to study the early stages of spore formation in members of the genus Bacillus in order to investigate further the origin and nature of the initial spore septum and the resulting forespore envelope. Whereas previously, with a methacrylate procedure, this layer had appeared to be continuous with the cell wall, this study reveals it as a double layer of cytoplasmic membrane. Perisporal, membranous organelles connected both to the developing forspore envelope and to the cytoplasmic membrane were encountered in the four species studied. Similar organelles were prominent during growth at the sites of transverse septa formation. These were connected to, or continuous with, the cytoplasmic membrane and often adherent to the chromatin bodies of the dividing bacilli.     

FINE STRUCTURAL STUDIES ON THE INTERPHASE AND DIVIDING APICAL CELLS OF SPHACELARIA TRIBULOIDES (PHAEOPHYTA)1

The apical cells of Sphacelaria tribuloides Menegh. are larger than other thallus cells, contain more organelles and appear polarized. Their tip portion, where they grow, contains a well developed Golgi apparatus, abundant endoplasmic reticulum (ER) membranes, mitochondria, chloroplasts and a large number of small vacuoles. It seems likely that a continuous flow of membranous material from the ER membranes to the dictyosomes and from the latter to the plasmalemma of the extending tip portion takes place. In contrast, the basal pole possesses fewer organelles and is occupied mainly by large-sized, sometimes central vacuoles. The apical cells undergo two distinct types of highly asymmetrical differential divisions giving rise to cells of the thallus and hair initials. During the early stages of mitosis the nuclear envelope remains intact, except for fenestrated poles. Microtubules pass through the fenestrae into the nucleoplasm. During meta-phase, a typical chromosome plate is organized. The sites of attachment of spindle microtubules to the chromosomes are structurally different from the rest of the chromosomes. At late anaphase, the nuclear envelope breaks down completely. During telophase, a new membrane encloses the chromosomes which are decondensed and the nucleoli are reorganized. Cytokinesis proceeds long after mitosis at a stage in which the nuclei have increased in size and have moved farther apart. A membranous furrow develops centripetally, without the participation of microtubules. However, microtubules traverse the thin cytoplasmic strands which, in both interphase and cytokinetic cells, meander among the vacuoles of the basal pole of the cell and the internuclear space. Dictyosomes appear to be involved in the subsequent wall deposition.