DNA SYNTHESIS AND MITOSIS IN MERISTEMS: REQUIREMENTS FOR RNA AND PROTEIN SYNTHESIS

Following provision of sucrose to starved, stationary phase pea root meristems, G₁ and G₂ cells enter DNA synthesis and mitosis, respectively. Puromycin (450 μg/ml) and cycloheximide (5 μg/ml) completely prevent this initiation of progression through the cell cycle. Actinomycin D (10 μg/ml) has no effect on the initial entry of G₁ and G₂ cells into S and mitosis, although later entry is prevented. The resistance of the cells to actinomycin D is lost slowly with time in medium without sucrose, suggesting that an RNA required for the resumption of proliferative activity is being gradually lost. The effects of the inhibitors on transitional and proliferative phase meristem cells indicate that such dividing cells do indeed have sufficient of the requisite RNA for 8-12 hr progression through the cycle, but that protein synthesis is required continuously. It is suggested that this RNA is the one lost slowly during starvation, allowing starved cells to reinitiate progression through the cycle in the presence of actinomycin D.     

THE PERMEABILITY OF CELLS FOR OXYGEN AND ITS SIGNIFICANCE FOR THE THEORY OF STIMULATION

It can be demonstrated by an indicator method that living cells are as freely permeable to oxygen as dead cells, and that sudden admission of oxygen to the cell cannot account for increased oxidation as a result of stimulation. Oxygen penetrates as readily as carbon dioxide among the acids and ammonia among the alkalies. Exposure of living plant cells to high oxygen pressures does not initiate certain oxidations (except after some hours), which proceed readily in dead plant cells in the air. In the light of the preceding statement, about the permeability of cells for oxygen, this is interpreted to mean that more oxygen enters the cell at high pressure, but that the reacting substances (chromogen and oxidase) are kept apart by some phase boundary as long as the cell is alive. Increased oxygen concentration eventually produces injury to the cell.     

A TECHNIQUE FOR ULTRACRYOTOMY OF CELL SUSPENSIONS AND TISSUES

Ultracryotomy of fixed tissue has been investigated for a number of years but, so far, success has been limited for several reasons. The simple technique herein reported allows the ultracryotomy not only of a variety of tissues but also of single cells in suspension, with a preservation and visualization of ultrastructural detail at least equivalent to that obtained with conventional embedding procedures. In this technique, sucrose is infused into glutaraldehyde-fixed tissue pieces before freezing for the purpose of controlling the sectioning consistency. By choosing the proper combinations of sucrose concentration and sectioning temperature, a wide variety of tissues can be smoothly sectioned. Isolated cells, suspended in a sucrose solution, are sectioned by sectioning the frozen droplet of the suspension. A small liquid droplet of a saturated or near-saturated sucrose solution, suspended on the tip of an eyelash probe, is used to transfer frozen sections from the knife edge onto a grid substrate or a water surface. Upon melting of the sections on the surface of the sucrose droplet, they are spread flat and smooth due to surface tension. When the section of a suspension of single cells melts, individual sections of cells remain confined to the small area of the droplet surface. These devices make it possible to cut wide dry sections, and to avoid flotation on dimethyl sulfoxide solutions. With appropriate staining procedures, well-preserved ultrastructural detail can be observed. The technique is illustrated with a number of tissue preparations and with suspensions of erythrocytes and bacterial cells.     

NEWER THERAPY FOR LEUKEMIA,POLYCYTHEMIA, AND LYMPHOMA

X-radiation remains the treatment of choice in most cases of leukemia and lymphoma, but new agents are playing an increasing role in therapy. Radioactive phosphorus does not produce radiation sickness and results with it are comparable to those of x-ray therapy in chronic leukemia. Urethane and nitrogen mustard may produce remissions in patients with chronic leukemia who have become resistant to radiation. Triethylene melamine may be administered orally with nitrogen mustard-like effects and is undergoing further trial. Aminopterin, ACTH and cortisone often cause short remissions in acute leukemia. Urethane is the best treatment available for multiple myeloma. Polycythemia vera is well controlled by radioactive phosphorus combined with venesection. Nitrogen mustard is often effective and triethylene melamine shows promise in Hodgkin''s disease. Antianemic substances such as iron and liver extract are of no value in the treatment of anemia caused by leukemia, lymphoma and myeloma.     

THE MOLLUSCAN RHOGOCYTE (PORE-CELL, BLASENZELLE, CELLULE NUCALE), AND ITS SIGNIFICANCE FOR IDEAS ON NEPHRIDIAL EVOLUTION

The rhogocyte (Leydig's cell, cellule nucale, Blasen-zelle,pore cell) is a specific molluscan cell type that occurs throughoutthe animal's primary body cavity, i.e free in the haemocoelor embedded in connective tissue. Rhogocytes closely resemblecyrtocytes and podocytes jn having slit-Uke diaphragms withan encoating extracellular matrix at their surface which probablyacts as a molecular sieve However, rhogocytes are solitary cells,whereas cyrtocytes and podocytes form epitheha Occurrence, variability,naming, and possible functions of the rhogocyte among the MoUuscaare reviewed and discussed. In general, rhogocytes play an importantrole in metal ion metabolism of molluscs, they are possiblyinvolved in the recycling of respiratory pigments and may alsoact in detoxification. Up to now direct homologues of the molluscan rhogocyte havenot been described in any other bi-laterian phylum, so-called‘pore cells’ of other phyla show entirely differentfine-structures However, at least partial (serial) homologycan be accepted between rhogocytes (free cells), arthropod nephro-cytes(cell aggregates or solitary cells) and podocytes (true epithelialcells), and there is also a cytological and functional continuumof both rhogocytes and podocytes to protonephndial cyrto- respectivelysolenocytes. Implications of this postulated ‘cell-family’with slit-diaphragms and their respective ultrafiltration systemsfor theories on the evolution of filtration nephndia are discussed. (Received 10 February 1995; accepted 10 October 1995)     

EVIDENCE FOR SEXUAL REPRODUCTION IN THE MARINE DINOFLAGELLATES, PYROCYSTIS NOCTILUCA AND PYROCYSTIS LUNULA (DINOPHYTA)

Evidence for sexual reproduction was observed in two oceanic dinoflagellate species, Pyrocystis noctiluca Murray ex Haeckel and Pyrocystis lunula (Schütt) Schütt. Observations suggest that cells underwent fertilization as opposed to cell division because of the following: first, fusing cells had a conspicuous pore (fusion pore) connecting the two gametes; dividing cells lacked this feature. In culture, about 0.1% of P. noctiluca cells had a fusion pore, which serves as a possible site for gamete recognition on the cell wall. Second, we document a temporal progression of plasmogamy and karyogamy. Fusion events in both species were observed at the beginning of the day, whereas division stages were most apparent at the end of the day.     

Fe-S Cluster Biogenesis in Isolated Mammalian Mitochondria: COORDINATED USE OF PERSULFIDE SULFUR AND IRON AND REQUIREMENTS FOR GTP,NADH, AND ATP*

Iron-sulfur (Fe-S) clusters are essential cofactors, and mitochondria contain several Fe-S proteins, including the [4Fe-4S] protein aconitase and the [2Fe-2S] protein ferredoxin. Fe-S cluster assembly of these proteins occurs within mitochondria. Although considerable data exist for yeast mitochondria, this biosynthetic process has never been directly demonstrated in mammalian mitochondria. Using [³⁵S]cysteine as the source of sulfur, here we show that mitochondria isolated from Cath.A-derived cells, a murine neuronal cell line, can synthesize and insert new Fe-³⁵S clusters into aconitase and ferredoxins. The process requires GTP, NADH, ATP, and iron, and hydrolysis of both GTP and ATP is necessary. Importantly, we have identified the ³⁵S-labeled persulfide on the NFS1 cysteine desulfurase as a genuine intermediate en route to Fe-S cluster synthesis. In physiological settings, the persulfide sulfur is released from NFS1 and transferred to a scaffold protein, where it combines with iron to form an Fe-S cluster intermediate. We found that the release of persulfide sulfur from NFS1 requires iron, showing that the use of iron and sulfur for the synthesis of Fe-S cluster intermediates is a highly coordinated process. The release of persulfide sulfur also requires GTP and NADH, probably mediated by a GTPase and a reductase, respectively. ATP, a cofactor for a multifunctional Hsp70 chaperone, is not required at this step. The experimental system described here may help to define the biochemical basis of diseases that are associated with impaired Fe-S cluster biogenesis in mitochondria, such as Friedreich ataxia.     

实验性脾虚证大鼠结肠和盲肠组织化学、免疫组织化学研究

成年雄性Wistar大鼠60只,分正常对照组;脾虚组;自然恢复组和四君子汤治疗组。四组动物取结肠和盲肠分别进行H-E;酶组织化学和5-羟色胺细胞PAP免疫组织化学观察。结果表明,脾虚组结肠和盲肠粘膜上皮的杯状细胞较对照组明显增多,淋巴细胞及淋巴集结也增多。固有层毛细血管充血并有炎症细胞的增多。上皮细胞和腺细胞琥珀酸脱氢酶(SDH)减弱,乳酸脱氢酶(LDH)增强;结肠上皮细胞和腺细胞碱性磷酸酶(AIP)和三磷酸腺苷酶(ATPase)增强,而盲肠上皮细胞和腺细胞AIP和ATPase减弱。结肠和盲肠5-HT细胞免疫组化反应减弱。这些结果表明肠上皮细胞酶的变化和5-HT细胞分泌活性的变化与脾虚证的发生密切相关,可能是导致脾虚证原因之一。经四君子汤治疗后以上各项指标均比自然恢复组更接近于正常对照组,说明此药对消化道粘膜上皮酶活性和5-HT细胞分泌活性恢复正常有明显效果,从而起治疗作用。     

A MODEL FOR DEVELOPMENT AND EVOLUTION OF COMPLEX MORPHOLOGICAL STRUCTURES

How 'complex' or composite morphological structures like the mammalian craniomandibular region arise during development and how they are altered during evolution are two major unresolved questions in biology. Herein, we have described a model for the development and evolution of complex morphological structures. The model assumes that natural selection acts upon an array of phenotypes generated by variation in a variety of underlying genetic and epigenetic controlling factors. Selection refines the integration of the various morphogenetic components during ontogeny in order to produce a functioning structure and to adapt the organisms to differing patterns of environmental heterogeneity. The model was applied to the development and evolution of the mammalian mandible (which is used as a paradigm of complex morphological structures). The embryology of the mandible was examined in detail in order to identify the fundamental developmental units which are necessary to assemble the final morphological structure. The model is quite general since equivalent units exist for the development of many other biological structures. This model could be applied to many other developing morphological structures as well as other groups of organisms. For example, it can be applied to cell parameters during Drosophila development (Atchley, 1987). The model as discussed in this paper assumes that morphological changes in the mandible result from evolutionary changes in its underlying developmental units. The developmental units relate to characteristics of cellular condensations which are produced from the differentiation of embryonic neural crest cells. The developmental units include: the number of stem cells in preskeletal condensations (n), the time of initiation of condensation formation (t), the fraction of cells that is mitotically active within a condensation (f), the rate of division of these cells (r), and their rate of cell death (d). These units and their derivative structures are discussed in terms of types of tissue differentiation (chondrogenesis, osteogenesis, primary/secondary osteogenesis, intramembranous/endochondral ossification) and growth properties of major morphological regions of the mandible. Variation in these five units provides the developmental basis for ontogenetic and phylogenetic modification of mandibular morphology. We have discussed how these developmental units are influenced by (a) the cell lineage from which they arise, (b) epithelial-mesenchymal (inductive tissue) interactions, (c) regulation of cell differentiation, and (d) extrinsic factors such as muscles, teeth and hormones. Evidence was provided that variation in mandibular morphology is heritable, subject to modification by natural selection, and that divergence among different genetic stocks has apparently occurred through changes in these developmental units and their derivative structures.(ABSTRACT TRUNCATED AT 400 WORDS)     

CHOREONEMA (CORALLINALES, RHODOPHYTA): 18S rDNA PHYLOGENY AND RESURRECTION OF THE HAPALIDIACEAE FOR THE SUBFAMILIES CHOREONEMATOIDEAE, AUSTROLITHOIDEAE, AND MELOBESIOIDEAE

Phylogenetic analyses of 18S rDNA gene data for Choreonema thuretii (Corallinales, Rhodophyta) and available data for other coralline red algae indicated that Choreonema belongs to the same lineage as other taxa of Corallinales possessing tetra/bisporangial conceptacles with multiporate plates. These results, when integrated with extant morphological/anatomical data, ultrastructural data, and taxonomic data led to the conclusion that all taxa of Corallinales possessing multiporate conceptacles belong to a distinct family, the Hapalidiaceae. Recognition of the Hapalidiaceae as a distinct family was supported both phylogenetically and phenetically. The Hapalidiaceae includes those taxa of Corallinales whose tetrasporangia produce zonately arranged spores and whose tetra/bisporangia are borne in conceptacles, produce apical plugs, and develop beneath multiporate plates. The Hapalidiaceae includes the subfamilies Choreonematoideae, Melobesioideae, and Austrolithoideae, formerly placed in the Corallinaceae sensu lato . The Choreonematoideae lack cell connections between adjacent vegetative filaments and have a multiporate plate that is acellular at maturity, consisting only of a calcium carbonate matrix. The Austrolithoideae and Melobesioideae both have cellular pore plates; taxa of Melobesioideae have cell fusions between cells of adjacent vegetative filaments, whereas taxa of Austrolithoideae lack cellular connections between adjacent vegetative filaments. Inclusion of the Austrolithoideae in the Hapalidiaceae was based entirely on morphological/anatomical evidence; molecular evidence currently is lacking. Relevant historical and nomenclatural data are included.     

NEURONAL AND GLIAL SYSTEMS FOR γ-AMINOBUTYRIC ACID METABOLISM

—Bulk prepared neuronal perikarya, nerve endings and glial cells have been used to study amino acid concentrations and GABA metabolism in vitro. All amino acids were more concentrated in synaptosomes and glial cells than in neuronal perikarya. Cell specificity was found with respect to the relative distribution of some amino acids. Glutamate decarboxylase activity was considerably higher in synaptosomes than in glial cells. The inhibitory effect of amino-oxyacetic acid on glutamate decarboxylase activity differed between synaptosomes and glial cells. γ-Aminobutyric acid-α-ketoglutarate transaminase had the highest activity in the glial cell fraction; the inhibition of amino-oxyacetic acid differed between glial and neuronal material. The metabolism of exogenous GABA just accumulated by a cell showed similar time characteristics in neuronal and glial material.     

NUCLEOCYTOPLASMIC RATIO REQUIREMENTS FOR THE INITIATION OF DNA REPLICATION AND FISSION IN TETRAHYMENA

Hydroxyurea (10 mM) arrests the exponential growth of Tetrahymena by blocking DNA replication during S-phase. After removal of the hydroxyurea (HU), they have a long recovery period during which they are active in DNA synthesis. ³H-TdR uptake showed that on completion of the recovery period, the cells divide (recovery division) and enter a cell cycle which lacks G₁. The frequency, size and DNA content of the extranuclear chromatin bodies (ECB) formed at this division are all markedly increased (2–4) over the corresponding values obtained from exponential growth phase controls. Microspectrophotometric analysis of macronuclear DNA content (N) coupled with the cytoplasmic dry mass (C) values suggest that specific N to C ratios (N/C) are required for the initiation of DNA replication and fission: during a normal (exponential growth) cell cycle, both N and C double, but asynchronously, so that the N/C of both post-fission-daughter cells and pre-fission cells is identical (standardized to N/C = 1) but late G₁ cells have a low N/C. During a 10 hr exposure to HU, the N remains essentially the same whereas the C increases. When the HU is removed, the N increases by 4× and the C continues to increase until just prior to recovery division when it also reaches a value 4× that of the original daughter cells. Thus, the N/C = 1 is re-established. The enlarged ECB formed during recovery division may function to lower the N/C in the daughter cells, which in turn may in some way stimulate immediate DNA replication, thus eliminating G₁. The elimination of G₁ (and shortening in a few subsequent cell cycles) allows less time for cytoplasmic growth and results in the return of the cells to the generation time and the N and C values observed prior to the HU treatment.     

QUANTITATIVE GENETIC MODELS FOR DEVELOPMENT,EPIGENETIC SELECTION,AND PHENOTYPIC EVOLUTION

Herein we describe a general multivariate quantitative genetic model that incorporates two potentially important developmental phenomena, maternal effects and epigenetic effects. Maternal and epigenetic effects are defined as partial regression coefficients and phenotypic variances are derived in terms of age-specific genetic and environmental variances. As a starting point, the traditional quantitative genetic model of additive gene effects and random environmental effects is cast in a developmental time framework. From this framework, we first extend a maternal effects model to include multiple developmental ages for the occurrence of maternal effects. An example of maternal effects occurring at multiple developmental ages is prenatal and postnatal maternal effects in mammals. Subsequently, a model of intrinsic and epigenetic effects in the absence of maternal effects is described. It is shown that genetic correlations can arise through epigenetic effects, and in the absence of other developmental effects, epigenetic effects are in general confounded with age-specific intrinsic genetic effects. Finally, the two effects are incorporated into the basic quantitative genetic model. For this more biologically realistic model combining maternal and epigenetic effects, it is shown that the phenotypic regressions of offspring on mother and offspring on father can be used in some cases to estimate simultaneously maternal effects and epigenetic effects.     

HIGH AND LOW DENSITY WATER AND RESTING,ACTIVE AND TRANSFORMED CELLS

Resting and active states of cells are described in terms of the expectation, derived from experiments with aqueous polymers, that they contain two modified forms of water: high density, reactive, fluid water and low density, inert, viscous water. Low density water predominates in a resting cell and is converted to high density water in an active cell. It is proposed that switching from one state to another is an integral part of cellular function. When this ability is lost cells are transformed either to a state of rigor or to a hyperactive state in which they no longer depend upon external signals.     

DEVELOPMENTAL ANATOMY OF THE STEM OF WELFIA GEORGII,IRIARTEA GIGANTEA,AND OTHER ARBORESCENT PALMS: IMPLICATIONS FOR MECHANICAL SUPPORT

Changes in stem anatomy with radial position and height were studied for the arborescent palms Welfia georgii, Iriartea gigantea, Socratea durissima, Euterpe macrospadix, Prestoea decurrens, and Cryosophila albida. Vascular bundles are concentrated toward the stem periphery and peripheral bundles contain more fibers than central bundles. Expansion and cell wall thickening of fibers within vascular bundles continues throughout the life of a palm, even in the oldest tissue. Within individual vascular bundles, the fibers nearest the phloem expand first and fiber cell walls become heavily thickened. A front of expanding fibers moves outward from the phloem until all fibers within a vascular bundle are fully expanded and have thick cell walls. Peripheral vascular bundles differentiate first and inner bundles later. In the stem beneath the crown, vascular bundles and ground tissue cells show little or no size increase, but marked cell wall thickening during development for Welfia georgii. Beneath the crown, diameters of peripheral vascular bundles increase more than twofold for Iriartea gigantea, while diameters of central bundles do not increase. In Iriartea stems, ground tissue cells at the periphery elongate to accommodate expanding vascular bundles and cell walls become thickened to a lesser degree than in fibers; central ground tissue cells elongate markedly, but cell walls do not become thickened; and large lacunae form between central parenchyma cells. For Iriartea, Socratea, and Euterpe, sustained cell expansion results in limited, but significant increases in stem diameter. For all species, sustained cell wall thickening results in dramatic increases in stem stiffness and strength.     

不同固定液和染色方法显示山羊子宫内肥大细胞效果的比较

目的探讨用不同固定液和染色方法对显示处于间情期山羊子宫肥大细胞的影响。方法用四种不同的固定方法,应用改良甲苯胺蓝（MTB）染色法和阿尔辛蓝-番红花红（AB-S）染色法显示处于间情期山羊子宫肥大细胞。结果山羊子宫组织采用Carnoy氏液固定,MTB和AB-S染色对所有的肥大细胞均可获得良好的染色反应,但10％中性福尔马林,4％多聚甲醛,Bouin氏液固定的组织仅有少量肥大细胞着染。结论MTB和AB-S染色法均是山羊子宫肥大细胞良好的染色方法。     

EVIDENCE FOR WIDESPREAD COURTSHIP DURING COPULATION IN 131 SPECIES OF INSECTS AND SPIDERS,AND IMPLICATIONS FOR CRYPTIC FEMALE CHOICE

Male courtship behavior is generally thought to function prior to copulation, as an inducement to the female to allow the male to copulate with her; this study indicates however, that male courtship during and following copulation (“copulatory courtship”) is common in insects and spiders (81% of 131 species in 102 genera and 49 families, mostly Coleoptera, Hemiptera, Diptera, and Araneioidea). Copulatory courtship is apparently evolutionarily labile, as expected if it is under sexual selection; intrageneric variation occurred in all 17 genera in which more than one species was observed. In 81% of 94 species with copulatory courtship, the male abandoned the female soon after copulation ended; thus, copulatory courtship appears not to function generally to induce acceptance of further copulatory attempts. The most likely explanation for copulatory courtship is that it represents attempts by males to influence cryptic female choice. This suggests that an aspect of sexual selection by female choice not considered by Darwin may be more important than previously appreciated and that the common practice in evolutionary studies of measuring male reproductive success by counting numbers of copulations may sometimes be misleading because of cryptic female choice during and after copulation.     

A MODIFIED PROCEDURE FOR ISOLATION OF ASTROCYTE- AND NEURON-ENRICHED FRACTIONS FROM RAT BRAIN

Our previously published method for isolation of neurons with extensive processes (Farooq et al., 1977) has been modified to permit the isolation of both astrocyte- and neuron-enriched fractions. Rat cerebral tissue is incubated with acetylated trypsin and disrupted. The cell suspension is separated first by differential centrifugation and then by gradient centrifugation on discontinuous Ficoll gradients. The method is reproducible and is applicable equally well to immature and adult animals. The yield of astrocytes of 57% particle purity, and higher weight purity, is 4–7 × 10⁶ cells/brain, amounting to 1.5–2.0 mg of protein. The astrocytes appear to be a mixture of fibrous and protoplasmic types. The yield of neurons of 90% particle purity is 10–14 × 10⁶ cells/brain, amounting to 2.4–3.0 mg of protein. A total yield of neurons of 28–37 × 10⁶ cells/brain can be obtained at 70% purity. These preparations have been characterized by light microscopy and protein, RNA and DNA content.     

AN IMPROVED METHOD OF CELL ENUMERATION FOR FILAMENTOUS ALGAE AND BACTERIA1

A new simple method for estimating the number of individual cells per milliliter in suspensions of filamentous microorganisms is described. This 2-part procedure utilizes a standard microscopic counting chamber and is independent of filament length or individual cell size. A statistical analysis of the method is also presented.     

SOME MODELS FOR DEVELOPMENT,GROWTH, AND MORPHOMETRIC CORRELATION

Genetic and phenotypic correlations between morphometric traits can be a direct consequence of shared developmental history and common systems of growth regulation. Correlation between traits, therefore, need not imply direct functional or adaptive constraints on those traits. Useful models of the developmental origins of correlations will consider mechanisms that can reduce initially high correlation of traits that arise from a single developmental precursor. Several models presented here predict such correlations for different modes of fission of a precursor. Timing of developmental events may also affect correlations and respond to selection on adult traits. The models may apply to development of the tetrapod limb bud, including variance and covariance induced by known developmental mutants.