THE EFFECTS OF ISOPROPYL N-PHENYL CARBAMATE ON THE GREEN ALGA OEDOGONIUM CARDIACUM : I. Cell Division

Cell division in vegetative filaments of the green alga Oedogonium cardiacum is presented as an experimental system. We report on how we have used this system to study the effects of isopropyl N-phenylcarbamate (IPC) on the mitotic apparatus and on the phycoplast, a planar array of cytokinetic microtubules. Polymerization of microtubules was prevented when filaments, synchronized by a light/dark regime and chilled (2°C) while in metaphase or just before phycoplast formation, were exposed to 5.5 x 10-4 M IPC and then returned to room temperature. Spindles reformed or phycoplasts formed when these filaments were transferred to growth medium free of IPC. However, the orientation of both microtubular systems was disturbed: the mitotic apparatus often contained three poles, frequently forming three daughter nuclei upon karyokinesis; the phycoplast was often stellate rather than planar, and it sometimes was displaced to the side of both daughter nuclei, resulting in a binucleate and an anucleate cell upon cytokinesis. Our results suggest that IPC (a) prevents the assembly of microtubules, (b) increases the number of functional polar bodies, and (c) affects the orientation of microtubules in O. cardiacum. High voltage (1,000 kV) electron microscopy of 0.5-µm thick sections allowed us to visualize the polar structures, which were not discernible in thin sections

An Osmotic Model of the Growing Pollen Tube

Pollen tube growth is central to the sexual reproduction of plants and is a longstanding model for cellular tip growth. For rapid tip growth, cell wall deposition and hardening must balance the rate of osmotic water uptake, and this involves the control of turgor pressure. Pressure contributes directly to both the driving force for water entry and tip expansion causing thinning of wall material. Understanding tip growth requires an analysis of the coordination of these processes and their regulation. Here we develop a quantitative physiological model which includes water entry by osmosis, the incorporation of cell wall material and the spreading of that material as a film at the tip. Parameters of the model have been determined from the literature and from measurements, by light, confocal and electron microscopy, together with results from experiments made on dye entry and plasmolysis in Lilium longiflorum. The model yields values of variables such as osmotic and turgor pressure, growth rates and wall thickness. The model and its predictive capacity were tested by comparing programmed simulations with experimental observations following perturbations of the growth medium. The model explains the role of turgor pressure and its observed constancy during oscillations; the stability of wall thickness under different conditions, without which the cell would burst; and some surprising properties such as the need for restricting osmotic permeability to a constant area near the tip, which was experimentally confirmed. To achieve both constancy of pressure and wall thickness under the range of conditions observed in steady-state growth the model reveals the need for a sensor that detects the driving potential for water entry and controls the deposition rate of wall material at the tip

Mitosis in a vertebrate cell

Presents a cultured newt cell in late prophase. Chromosome condensation is well advanced, but the nuclear envelope is still visible at the periphery of the chromosomes. Shortly after the movie begins, the nuclear envelope disperses, and the influence of the spindle on the chromosomes becomes evident. The apparently random distribution of the chromosomes is modified by the action of the spindle, and many of the chromosomes move towards a region that runs across the space previously occupied by the nucleus from lower left to upper right. During this time, chromosome condensation continues, so each chromosome becomes slightly shorter, fatter, and darker (as seen in the phase microscope). With phase optics, the mitotic spindle itself is barely visible in this cell type. It can be identified in these images as a region on either side of the clustering chromosomes that appears particularly featureless. This appearance derives from the spindle's ability to exclude many granules and cytoplasmic organelles, like mitochondria, which have characteristic phase densities. Careful examination of the 'featureless' regions will show, however, a subtle fibrosity with just-detectable filaments that run from the chromosomal region to a point on either side of it; these points are the poles of the mitotic spindleComponente Curricular::Educação Superior::Ciências Biológicas::Biologia Gera

Mitosis in a vertebrate cell

Presents a cultured newt cell in late prophase. Chromosome condensation is well advanced, but the nuclear envelope is still visible at the periphery of the chromosomes. Shortly after the movie begins, the nuclear envelope disperses, and the influence of the spindle on the chromosomes becomes evident. The apparently random distribution of the chromosomes is modified by the action of the spindle, and many of the chromosomes move towards a region that runs across the space previously occupied by the nucleus from lower left to upper right. During this time, chromosome condensation continues, so each chromosome becomes slightly shorter, fatter, and darker (as seen in the phase microscope). With phase optics, the mitotic spindle itself is barely visible in this cell type. It can be identified in these images as a region on either side of the clustering chromosomes that appears particularly featureless. This appearance derives from the spindle's ability to exclude many granules and cytoplasmic organelles, like mitochondria, which have characteristic phase densities. Careful examination of the 'featureless' regions will show, however, a subtle fibrosity with just-detectable filaments that run from the chromosomal region to a point on either side of it; these points are the poles of the mitotic spindleComponente Curricular::Educação Superior::Ciências Biológicas::Biologia Gera

On the mechanism of anaphase A: evidence that ATP is needed for microtubule disassembly and not generation of polewards force

Abstract. As anaphase began, mitotic PtK ~ and newt lung epithelial cells were permeabilized with digitonin in permeabilization medium (PM). Permeabilization stopped cytoplasmic activity, chromosome movement, and cytokinesis within about 3 min, presumably due to the loss of endogenous ATE ATE GTP, or ATP-~-S added in the PM 4-7 min later restarted anaphase A while kinetochore fibers shortened. AMPPNP could not restart anaphase A; ATP was ineffective if the spindle was stabilized in PM + DMSO. Cells permeabilized in PM + taxol varied in their response to ATP depending on the stage of anaphase reached: one mid-anaphase cell showed initial movement of chromosomes back to the metaphase plate upon permeabilization but later, anaphase A resumed when ATP was added. Anaphase A was also reactivated by cold PM (,x,16 ~ or PM containing calcium (1-10 mM). Staining of fixed cells with antitubulin showed that microtu-N 'o consensus currently exists regarding the role of ATP during anaphase A (chromosome-to-pole movement) and it is difficult to evaluate the diverse data on this subject. We (Pickett-Heaps and Spurck, 1982; Spurck et al., 1986a, b) have found that the concentration of metabolic inhibitors needed to cause rapid (within 1 min) and completely reversible cessation of cellular activity (presumably by depleting cellular ATP levels) in live cells is critical; 2,4 dinitrophenol (DNP) 1 is effective in both diatoms and mammalian cells only at the seemingly high concentration of 1 mM, and lowering the concentration even to 0.5 mM results in a significantly delayed and often incomplete response. In contrast, far lower concentrations of metabolic inhibitors were used in most previous work (reviewed by Spurck et al., 1986a, b; Hepler and Palevitz, 1986). Furthermore, many who work with animal cell systems use inhibitors of oxidative phosphorylation alone. We (Spurck et al., 1986a) found that glycolysis must also be blocked by usin