Motile plant cell body: a "bug" within a "cage"

Analysis of the cytoskeleton in morphogenetically active plant cells allows us to propose a unified concept for the structural organization of eukaryotic cells. Their cytoarchitecture is determined by two principal structural complexes: nucleus-microtubule-based cell bodies ("bugs") and plasma-membrane-F-actin-based cell periphery complexes ("cages"). There are dynamic interactions between each of these entities in response to extracellular and intracellular signals. In the case of the cell body, these signals determine its polarization, rotation and migration. Interactions between cell body and cell periphery complexes determine cell growth polarity and morphogenesis throughout the eukaryotic kingdom.     

"Ant" and "grasshopper" life-history strategies in Saccharomyces cerevisiae

From the evolutionary and ecological points of view, it is essential to distinguish between the genetic and environmental components of the variability of life-history traits and of their trade-offs. Among the factors affecting this variability, the resource uptake rate deserves particular attention, because it depends on both the environment and the genetic background of the individuals. In order to unravel the bases of the life-history strategies in yeast, we grew a collection of twelve strains of Saccharomyces cerevisiae from different industrial and geographical origins in three culture media differing for their glucose content. Using a population dynamics model to fit the change of population size over time, we estimated the intrinsic growth rate (r), the carrying capacity (K), the mean cell size and the glucose consumption rate per cell. The life-history traits, as well as the glucose consumption rate, displayed large genetic and plastic variability and genetic-by-environment interactions. Within each medium, growth rate and carrying capacity were not correlated, but a marked trade-off between these traits was observed over the media, with high K and low r in the glucose rich medium and low K and high r in the other media. The cell size was tightly negatively correlated to carrying capacity in all conditions. The resource consumption rate appeared to be a clear-cut determinant of both the carrying capacity and the cell size in all media, since it accounted for 37% to 84% of the variation of those traits. In a given medium, the strains that consume glucose at high rate have large cell size and low carrying capacity, while the strains that consume glucose at low rate have small cell size but high carrying capacity. These two contrasted behaviors may be metaphorically defined as "ant" and "grasshopper" strategies of resource utilization. Interestingly, a strain may be "ant" in one medium and "grasshopper" in another. These life-history strategies are discussed with regards to yeast physiology, and in an evolutionary perspective.     

乙型肝炎病毒表面抗原a、d、r亚型单克隆抗体的制备与应用

本文成功地建立了分泌抗乙型肝炎病毒表面抗原(抗-HBsAg)a,d、r3种亚型决定簇抗体的4株杂交瘤细胞。经一系列生化、免疫学鉴定,证明4株细胞所分泌的单克隆抗体(McAb)均具有各自的亚型特异性。反复克隆培养16周,并液氮冻存8个月后复苏,抗体的效价仍稳定不变。用纯化的McAb制备RPHA诊断试剂,检测了80例有乙型肝炎自觉症     

Binding of lectins to "young" and "old" human erythrocytes

"Old" human erythrocytes showed a 21.2% decrease in cell surface area and a 2% decrease in the number of WGA receptor sites, but a 27% increase in the distribution density of the WGA (lectin) receptor site, when compared with "young" human erythrocytes. For a list of lectin abbreviations, see Materials and methods). Both "young" and "old" erythrocytes exhibited very weak binding activity for 125I-labeled PNA, but there was no difference in binding activity for PNA between "young" erythrocytes and "old" ones. Compared with "young" erythrocytes, decreases in the number and distribution density of receptor sites for five lectins including LPA, Con A, RCA-II, SBA and BPA on the cell surface were observed in aged erythrocytes. "Old" erythrocytes also showed a decrease in the number of PHA-E receptor sites, while the distribution density of the same receptor site remained unchanged. In view of these and other observations, it is thought that human erythrocyte aging is accompanied by elimination of some glycoconjugates which have affinity for six lectins, LPA, Con A, RCA-II, PHA-E, SBA and BPA, whereas no WGA receptor-containing glycoconjugates are released from erythrocyte membranes. Elimination of the glycoconjugates results in shrinkage of erythrocytes to reduce their cell surface areas.     

Actin-Based Domains of the "Cell Periphery Complex" and their Associations with Polarized "Cell Bodies" in Higher Plants

Abstract: Nascent cellulosic cell wall microfibrils and transverse (with respect of cell growth axis) arrays of cortical microtubules (MTs) beneath the plasma membrane (PM) are two well established features of the periphery of higher plant cells. Together with transmembrane synthase complexes, they represent the most characteristic form of a “cell periphery complex” of higher plant cells which determines the orientation of the diffuse (intercalary) type of their cell growth. However, there are some plant cell types having distinct cell cortex domains which are depleted of cortical MTs. These particular cell cortex domains are, instead, typically enriched with components of the actin‐based cytoskeleton. In higher plants, this feature is prominent at extending apices of two cell types displaying tip growth ‐ pollen tubes and root hairs. In the latter cell type, highly dynamic F‐actin meshworks accumulate at extending tips, and they appear to be critical for the apparently motile character of these subcellular domains. Importantly, tip growth of both root hairs and pollen tubes is immediately stopped when the most dynamic F‐actin population is depolymerized with low levels of anti‐F‐actin drugs. Intriguingly, MTs of tip‐growing plant cells are organized in the form of longitudinal arrays, throughout the cytoplasm, which interconnect the extending tips with the subapical nuclei. This suggests that actin‐rich cell cortex domains polarize plant “cell bodies” represented by nucleus‐MTs complexes. A similar polarization of “cell bodies” is typical of mitotic and cytokinetic plant cells. A further type of MT‐depleted and actomyosin‐enriched plant cell cortex domain comprises the plasmodesmata. Primary plasmodesmata are formed during cytokinesis as part of the myosin VIII‐enriched callosic cell plates, representing “juvenile” forms of the plant “cell periphery complex”. In phylogenetic terms the association between F‐actin and the PM may be considered for a more “primitive” form of cellular organization than does the association of cortical MTs with the PM. We hypothesize that the actin cytoskeleton is a natural partner of the PM in all eukaryotic cells. In most plant cells, however, it was replaced by a tubulin‐based “cell periphery apparatus” which regulates, via still unknown mechanisms, the spatial deposition of nascent cellulosic microfibrils synthesized by PM‐associated synthase complexes.     

Combination of quantification and observation methods for study of "Side Population" cells in their "in vitro" microenvironment.

BACKGROUND: Qualitative and quantitative analyses of the rare phenotypic variants in in vitro culture systems is necessary for the understanding of cell differentiation in cell culture of primary cells or cell lines. Slide-based cytometry combines image acquisition and data treatment, and associates the power of flow cytometry (FCM) and the resolution of the microscopic studies making it suitable for the analysis of cells with rare phenotype. In this paper we develop a method that applies these principles to a particularly hot problem in cell biology, the study of stem cell like cells in cultures of primary cells, cancer cells, and various cell lines. METHODS: The adherent cells were labeled by the fluorescent dye Hoechst 33342. The images of cell populations were collected by a two-photon microscope and processed by a software developed by us. The software allows the automated segmentation of the nuclei in a very dense cell environment, the measurement of the fluorescence intensity of each nucleus and the recording of their position in the plate. The cells with a given fluorescence intensity can then be located easily on the recorded image of the culture plate for further analysis. RESULTS: The potential of our method is illustrated by the identification and localization of SP cells in the cultures of the C2C12 cell line. Although these cells represent only about 1% of the total population as calculated by flow cytometry, they can be identified in the culture plate with high precision by microscopy. CONCLUSION: Cells with the rare stem-cell like phenotype can be efficiently identified in the undisturbed cultures. Since the fluorescence intensity of rare events and the position of thousands of surrounding cells are recorded at the same time, the method associates the advantage of the FCM analysis and the microscopic observation.     

Toti "potent" repressors

A fascinating property of germ cells is their ability to maintain totipotency throughout development. At fertilization, this totipotency is unleashed and the egg generates all the cell types needed to make a brand new organism. Occasionally, germ cells differentiate precociously in the embryo or in the gonads and form teratomas, tumors containing many differentiated somatic cell types. Until recently, the genetic basis for teratoma formation was not known. The unexpected discovery of a teratoma in a C. elegans double mutant points to translational control as a key mechanism to maintain totipotency in developing germ cells.     

A molecular signature for the "master" heart cell

The vertebrate heart comprises a variety of cell types, the majority of which are cardiomyocytes, smooth muscle and endothelial cells. Their origin is still an intriguing research topic and the question is whether these cells derive from a common or from multiple distinct progenitor cell(s). Three recent publications not only suggest the existence of a single progenitor cell that can give rise to cardiovascular lineages but additionally uncovered, at least in part, the molecular identity of such a multipotent precursor cell. These findings constitute major progress in the quest for stem-cell therapies for cardiac diseases.     

"Clock-scan" protocol for image analysis

Comparative analysis of extra- and intracellular distributions of protein markers in immunohistochemical and immunofluorescent studies relies on techniques of image analysis. Line or region of interest pixel intensity scans are methods routinely used. However, although having good spatial resolution, linear pixel intensity scans fail to produce integral image of the cellular distribution of the label. On the other hand, the regions of interest scans have good integrative capacity but low spatial resolution. In this work, we describe a "clock-scan" protocol that, when applied to convex objects (such as neuronal cell bodies and the majority of cells in culture), combines advantages and circumnavigates limitations of the above-mentioned techniques. The protocol 1) collects multiple radial pixel intensity profiles scanned from the cell center to the periphery, 2) scales these profiles according to the cell radius measured in the direction of the scan, and finally, 3) averages these individual profiles into one integral radial pixel intensity profile. Because of scaling, the mean pixel intensity profiles produced by the clock-scan protocol depend on neither the cell size nor, within reasonable limits, the cell shape. This allows direct comparison or, if required, averaging or subtraction of profiles of different cells. We have successfully tested the clock-scan protocol in experiments with immunostained dorsal root ganglion neurons. In addition, the protocol seems to be equally applicable for studies in a variety of other preparations. techniques; methods; cell biology; histology; immunohistochemistry     

"Big it up": endoreduplication and cell-size control in plants

Cells undergoing endoreduplication replicate chromosomal DNA without intervening mitoses. The resulting larger, higher-ploidy nucleus is often associated with an increase in cell size, but the molecular basis for this correlation remains poorly understood. Recent advances in characterising various mutants and transgenic plants are beginning to unravel how this unique type of cell cycling is regulated and how it contributes to cell-size control. Both cell growth (i.e. increase in cytoplasmic macromolecular mass) and cell expansion (i.e. increase in cell volume through vacuolation) contribute independently to increases in cell size in plants. A total organ-size checkpoint may also help to coordinate cell size and cell number within an organ, and can contribute to final cell-size determination in plants.     

"Omic" approaches for unraveling signaling networks

Signaling pathways are crucial for cell differentiation and response to cellular environments. Recently, a large number of approaches for the global analysis of genes and proteins have been described. These have provided important new insights into the components of different pathways and the molecular and cellular responses of these pathways. This review covers genomic and proteomic (collectively referred to as "omic") approaches for the global analysis of cell signaling, including gene expression profiling and analysis, protein-protein interaction methods, protein microarrays, mass spectroscopy and gene-disruption and engineering approaches.     

Lymphoma immunophenotyping: "borderline" lymphomas

Immunophenotyping of B-cell lymphoproliferative disorders is indispensable, especially in disorders with CD19(+) CD5(+) B lymphocytes, where we have to make the distinction between low grade neoplasia, such as chronic lymphocytic leukemia with CD23(+) malignant lymphocytes, and aggressive neoplasia such as mantle cell lymphoma with CD23(-) malignant lymphocytes. We found some cases of CD19(+) CD5(+) lymphoproliferative disorders that do not meet all criteria for diagnosis of chronic lymphocytic leukemia or mantle cell lymphoma. For instance, we found cases with a low or no expression of CD23, asociated with absence of expression of FMC7 and surface immunoglobulins. These cases could be classified as "borderline" CD19(+) CD5(+) B cell lymphoproliferative disorders, with an intermediate neoplasic grade.     

Bacterial cell polarity: a "swarmer-stalked" tale of actin

The actin cytoskeleton is important for cell polarity and morphogenesis in eukaryotic organisms. A recent article describes an unexpected requirement for the actin-like protein MreB in the polarization of the bacterium Caulobacter crescentus. More surprisingly, the formation of a filamentous MreB structure that traverses the length of the cell is sufficient for randomized polar localization of cell-fate proteins. In this article, we discuss the significance of these findings and the possible mechanisms by which an actin-like cytoskeleton could mediate cell polarity in bacteria.     

Sirt1, Notch and stem cell "age asymmetry"

Almost all complex multicellular organisms on earth utilize oxygen for the production of energy. This strategy carries the risk for damaging ROS to be generated and so these biochemical pathways must be highly regulated. Because of this, regulation of oxidative-phosphorylation is tightly coordinated with every aspect of cellular physiology, including stem cell regulation during embryonic development and in adult organisms. The protein-deacetylase, SIRT1, has received much attention because of its roles in oxygen metabolism, cellular stress response, aging, and has been investigated in various species and cell types including embryonic stem cells. However, there is a dearth of information on SIRT1 in adult stem cells, which have a pivotal role in adult aging processes. Here, we discuss the potential relationships between SIRT1 and the surface receptor protein, Notch, with stem cell self-renewal, asymmetric cell division, signaling, and stem cell aging.     

"TORCing" neutrophil chemotaxis

During cell migration, chemoattractant-induced signaling pathways determine the direction of movement by controlling the spatiotemporal dynamics of cytoskeletal components. In this issue of Developmental Cell, Liu et?al. report that the target of rapamycin complex 2 (TORC2) controls cell polarity and chemotaxis through regulation of both F-actin and myosin II in migrating neutrophils.     

Energy transfer in "parasitic" cancer metabolism

It is now widely recognized that the tumor microenvironment promotes cancer cell growth and metastasis via changes in cytokine secretion and extracellular matrix remodeling. However, the role of tumor stromal cells in providing energy for epithelial cancer cell growth is a newly emerging paradigm. For example, we and others have recently proposed that tumor growth and metastasis is related to an energy imbalance. Host cells produce energy-rich nutrients via catabolism (through autophagy, mitophagy, and aerobic glycolysis), which are then transferred to cancer cells to fuel anabolic tumor growth. Stromal cell-derived L-lactate is taken up by cancer cells and is used for mitochondrial oxidative phosphorylation (OXPHOS) to produce ATP efficiently. However, “parasitic” energy transfer may be a more generalized mechanism in cancer biology than previously appreciated. Two recent papers in Science and Nature Medicine now show that lipolysis in host tissues also fuels tumor growth. These studies demonstrate that free fatty acids produced by host cell lipolysis are re-used via beta-oxidation (beta-OX) in cancer cell mitochondria. Thus, stromal catabolites (such as lactate, ketones, glutamine and free fatty acids) promote tumor growth by acting as high-energy onco-metabolites. As such, host catabolism, via autophagy, mitophagy and lipolysis, may explain the pathogenesis of cancer-associated cachexia and provides exciting new druggable targets for novel therapeutic interventions. Taken together, these findings also suggest that tumor cells promote their own growth and survival by behaving as a “parasitic organism.” Hence, we propose the term “Parasitic Cancer Metabolism” to describe this type of metabolic coupling in tumors. Targeting tumor cell mitochondria (OXPHOS and beta-OX) would effectively uncouple tumor cells from their hosts, leading to their acute starvation. In this context, we discuss new evidence that high-energy onco-metabolites (produced by the stroma) can confer drug resistance. Importantly, this metabolic chemo-resistance is reversed by blocking OXPHOS in cancer cell mitochondria with drugs like Metformin, a mitochondrial “poison.” In summary, parasitic cancer metabolism is achieved architecturally by dividing tumor tissue into at least two well-defined opposing “metabolic compartments:” catabolic and anabolic.     

Ly-1 B helper cells in autoimmune "viable motheaten" mice

Previous work has demonstrated that Ly-1 B cells from normal C57BL/6J mice help the response of B cell subsets to the 4-hydroxy-3-nitrophenylacetyl hapten (NP). This regulatory cell population, called BH, preferentially helps the expression of plaque-forming B cells which express a predominant set of serologically related determinants collectively known as the NPb idiotype family. The specificity of BH cell activity in the NP system is a reflection of NPb idiotype-specific BH cell surface receptors. Thus, BH cells recognize autologous (i.e., idiotype) antigens. Given these observations and previous associations of increased Ly-1 B cell frequency in autoimmune mice, it was hypothesized that autoreactive Ly-1 BH cells may be present in high frequencies and in an activated state in autoimmune mice. To test this hypothesis the immunologic activity of BH cells in autoimmune viable motheaten (mev/mev) mice was studied. It was determined that splenic BH cells are approximately 10 times more frequent in viable motheaten than normal mice. The fact that BH cells from viable motheaten mice are activated was suggested by the presence of NPb idiotype-specific BH replacing helper activity in sera or B cell supernatants from these autoimmune mice. The soluble helper activities constitutively produced in mev/mev splenic B cell cultures and detected in mev/mev serum were resolved into two moieties, an NPb idiotype-specific immunoglobulin and a nonimmunoglobulin lymphokine(s) fraction. Purified mev/mev B cell-derived B cell maturation factor could substitute for the lymphokine moiety in the NPb idiotype helper cell assay. These results suggest that at least two signals, anti-idiotype immunoglobulin and a late-acting B cell maturation factor, are required for BH-dependent helper activity. The relationships of these results to current concepts of B cell activation mechanisms and the possible association of Ly-1 BH cells with autoimmunity are discussed.     

Transferrin receptor polarity and recycling accuracy in "tight" and "leaky" strains of Madin-Darby canine kidney cells

We have characterized the polarity of the transferrin receptor in the epithelial Madin-Darby canine kidney (MDCK) cell line. The receptor is present in approximately 165,000 copies per cell, migrates as a diffuse band upon SDS gel electrophoresis with Mr 90,000, displays a dissociation constant for diferritransferrin at neutral pH of approximately 2 nM, and is active in essentially all of the cells of the population. Transferrin-mediated 55Fe uptake was used to measure the polarity of active transferrin receptors in filter-grown MDCK cells. The ratio of basolateral to apical receptors was approximately 800:1 for the high resistance strain I MDCK cells (typically greater than 2,000 ohm X cm2) and approximately 300:1 for the lower resistance strain II cells (less than 350 ohm X cm2). In combination with morphometric data this shows that a difference in resistance between these two strains is not reflected in a significant difference in cell surface polarity. We used the recycling of transferrin receptor in filter-grown MDCK cells to evaluate the accuracy of the sorting of a basolateral protein during endocytosis. Monitoring the amount of apically released 125I-labeled transferrin after application of 55Fe- and 125I-labeled transferrin to the basolateral surface provided a sensitive assay of the accuracy of sorting during recycling of the receptor from endosomes to the plasma membrane. The accuracy of transferrin receptor sorting (greater than 99.88%) during a single cycle of transit between the endosome and the plasma membrane is sufficient to maintain the high level of polarity of the cell.     

Regulated expression of the RB "tumor suppressor gene" in normal lymphocyte mitogenesis: elevated expression in transformed leukocytes and role as a "status quo" gene.

Expression of the RB retinoblastoma tumor suppressor gene product is regulated early during the stimulation of normal human peripheral blood lymphocytes, suggesting a regulatory role for the amount of this protein in mitogenesis of normal cells. When normal human peripheral blood lymphocytes were mitogenically stimulated with pokeweed mitogen, bivariate flow cytometric measurements of cellular DNA and RB protein content showed an early decrease in the amount of RB protein per cell, anteceding onset of S phase. A subsequent increase in the amount of RB protein per cell occurred with cell proliferation. Thus the amount of RB protein relative to the total cell mass underwent a biphasic response with mitogenesis. The resulting proliferating cells had a slightly elevated level of RB protein per cell compared to the unstimulated cells. Comparison of other proliferating leukocytes to normal lymphocytes showed that both EBV virally transformed lymphocytes and human promyelocytic leukemia cells (HL-60) had elevated levels of RB protein per cell compared to normal peripheral blood lymphocytes. Mitogenic stimulation or transformation by other means thus is associated with regulation of the amount of RB protein per cell, suggesting a regulatory role for the RB protein in normal cell growth control.     

Xeno-cannibalism: a survival "escamotage"

Several lines of evidence have demonstrated that self-cannibalism (macroautophagy) is a well regulated process of cell repair as well as of molecule and organelle recycling that allows the cells to survive. However, autophagic activity also represents a cell death pathway characterized by specific features that differentiate autophagy from other cell death processes. We found that cells that are able to exert intense autophagic activity were also able to engulf and digest entire cell siblings. This phenomenon represents a sort of xeno-cannibalism. We wonder whether these two phenomena, self and xeno-cannibalism, could be related the latter being an exacerbation of the first and providing a further survival option to the cells.