Protoplasma

Flagellar scales from the green flagellateTetraselmis striata (Prasinophyceae) were isolated, purified by isopycnic cesium chloride-gradient and zonal sucrose gradient centrifugation and their structure and biochemical composition investigated. Three types of flagellar scales were purified to more than 90% purity, a fourth type up to 75% purity. In addition to the previously known types of flagellar scales (pentagonal scales, rod-shaped scales, hair-scales), a novel scale type (i.e., the knotted scales) was discovered. New information about the asymmetric structure of the rod-shaped scales is presented and consequently they are renamed “man scales”. Flagellar scales consist mainly of carbohydrate (50–70%), significant amounts of protein (11% of dry weight) were found only in pentagonal scales. The main sugars (90%) of the pentagonal and man scales are the unusual 2-keto-sugar acids 3-deoxy-5-O-methyl-2-octulosonic acid (5 OMeKDO), 3-deoxy-2-heptulosaric acid (DHA), and 3-deoxy-2-octulosonic acid (KDO), the knotted scales contain as major sugars galactose and arabinose in addition to KDO and 5 OMeKDO but lack DHA. 13 major polypeptides were identified in flagellar scales by one-dimensional SDS-PAGE, 11 of these are of high molecular mass (>116 kDa). While the majority of polypeptides was found associated with pentagonal scales, at least 4 polypeptides were tentatively assigned to the hair-scales and knotted scales.

A method is described for non-radioactive labeling of total mRNA [poly(A)+ RNA] in plastic-embedded plant tissue sections. Oligo-deoxythymidylic acid (oligo-dT) labeled with digoxigenin-conjugated dUTP was used for in situ hybridization to poly(A)+ RNA in sections of tobacco (Nicotiana tabacum) anthers. The digoxigenin was immuno-stained using antidigoxigenin IgG and gold-labeled protein-A, followed by silver enhancement of the gold label. Reproducibly similar positive staining patterns were obtained with digoxigenin-labeled oligo-dT and polyuridylic acid [poly(U)], but not with a similarly labeled sense probe, poly(A). In the developing anthers, from the onset of meiosis to the production of pollen grains, labeling patterns were compatible with a gradual depletion of nuclear and chromosome-associated sporophytic mRNA molecules during prophase of meiosis, followed by postmeiotic production of gametophytic mRNA in microspore nuclei and the vegetative nuclei of the pollen grains.

H. chlorinus Tul., Ascomycète parasite des carpophores deBoletus sp. est déficient partiel en biotine. Les caractéristiques ultrastructurales des hyphes obtenues en présence de biotine sont données. Différentes techniques de fixation et de coloration (Thiery, Rambourg) ont été utilisées pour définir la localisation des polysaccharides et l'origine des vésicules apicales. La présence d'un appareil de Golgi chez cet Ascomycète est discutée. La carence en vitamine entraÎne,in vitro, des variations ultrastructurales telle que: l'hypertrophie de certaines cellules; l'épaississement des parois fongiques qui ont un aspect stratifié et qui présentent des irrégularités dans leur épaisseur leur donnant un aspect irrégulier; la mise en évidence moins nette du plasmalemme; la fréquence moins importante des systèmes endomembranaires; la présence d'inclusions lipidiques et de glycogène. Ces observations sont mises en relation avec les résultats de recherches biochimiques que nous effectuons parallèlement. La carence en biotine entraÎne une déviation du métabolisme cellulaire qui s'oriente vers la constitution de réserves cytoplasmiques plutÔt que vers la synthèse de nouvelles structures, ce qui aboutit à un développement réduit du mycélium.

Endogenous pH profiles were measured around single fertilized eggs of the brown algaPelvetia during the earliest stages of development. Profiles were constructed by measuring the pH near the cell surface at several positions using a pH sensitive microelectrode. Transcellular pH differences in the medium surrounding zygotes were detected soon after fertilization, as the developmental axis was being formed. The future rhizoid end of the cell was relatively alkaline and the presumptive thallus was acidic. At germination and throughout the first 5 d of embryogenesis, the apex of the elongating rhizoid was alkaline with respect to more distal regions. However, conditions that dissipated or reversed this extracellular pH gradient had little or no effect on polarization or growth, indicating that the gradient was not essential for early development. Inhibition of respiratory electron transport by cyanide and antimycin A eliminated the pH gradient, while uncouplers of oxidative phosphorylation [2,4-dinitrophenol (DNP) and carbonylcyanide m-chlorophenylhydrazone (CCCP)] stimulated acidification of the thallus regions. Proton ATPase inhibitors had no effect. Acidification, therefore, is not generated by ATP-dependent proton pumps in the plasma membrane, and instead probably reflects secretion of metabolic acids. Localized metabolism may establish an internal pH gradient that controls regional differentiation, and we are presently investigating this possibility.

The mechanism of cytokinesis was investigated during the first asymmetric division in fucoid zygotes. A plate of actin assembled midway between daughter nuclei where microtubules interdigitated and defined the cytokinetic plane. A membrane was then deposited in islands throughout the cytokinetic plane; the islands eventually fused into a continuous partition membrane and cell plate material was deposited in the intermembrane space. All of these structures matured from the center of the cell outward (centrifugal maturation). Pharmacological agents were used to investigate the roles of microtubules, actin, and secretion in cytokinesis. The findings indicate a mechanism of cytokinesis that may be unique to the brown algae.

Suspension culture cells of sycamore (Acer pseudoplatanus L.) and carrot (Daucus carota L.) were frozen to ultralow temperatures under rapid (⩾ 100 °C s−1) and slow, controlled (1 or 2 °C min−1) rates, in the presence and absence of cryoprotective compounds. After storage at −196 °C, cells were recovered by thawing either slowly, in air at room temperature (ca. 20 °C min−1) or rapidly, in a water bath at 40 °C (ca. 100 °C min−1). The ultrastructure of the thawed cells was examined by thin-sectioning and compared with unfrozen controls and cells examined in the frozen state. Cells frozen rapidly, in the presence of cryoprotectants, or frozen slowly in their absence, suffered serious ultrastructural damage and a total loss of viability. Carrot cells frozen at a rate of 2 °C min−1 in the presence of cryoprotectants and thawed at either rate, yielded up to 70% of viable cells. The recovered aggregates of carrot cells comprised some centrally located, seriously damaged cells and, at the periphery, groups of cells with a high electron opacity neighbouring well preserved cells, showing little ultrastructural modification compared with unfrozen controls. The highest rate of survival of sycamore cells (ca. 30%) was observed when they were frozen at a rate of 1 °C min−1 and thawed rapidly. In all recoverd cells of sycamore some ultrastructural modifications were evident. These included: dilation of mitochondria, plastids, golgi and ER cisternae and the nuclear envelope, decrease in polysomes, increase in nuclear and cytoplasmic microfilaments and changes in nuclear and nucleolar granularity. The probable causes and timing of the ultrastructural changes and their effects on the potential for regrowth of the recovered cells are discussed.