Condensation-decondensation of the gamma-tubulin containing material in the absence of a structurally visible organelle during the cell cycle of Physarum plasmodia.

Genetic evidence has shown the presence of a common spindle pole organiser in Physarum amoebae and plasmodia. But the typical centrosome and mitosis observed in amoebae are replaced in plasmodia by an intranuclear mitosis devoid of any structurally defined organelle. The fate of gamma-tubulin and of another component (TPH17) of the centrosome of Physarum amoebae was investigated in the nuclei of synchronous plasmodia. These two amoebal centrosomal elements were present in the nuclear compartment during the entire cell cycle and exhibited similar relocalisation from metaphase to telophase. Three preparation methods showed that gamma-tubulin containing material was dispersed in the nucleoplasm during interphase. It constituted an intranuclear thread-like structure. In contrast, the TPH17 epitope exhibited a localisation close to the nucleolus. In late G2-phase, the gamma-tubulin containing elements condensed in a single organelle which further divided. Intranuclear microtubules appeared before the condensation of the gamma-tubulin material and treatment with microtubule poisons suggested that microtubules were required in this process. The TPH17 epitope relocalised in the intranuclear spindle later than the gamma-tubulin containing material suggesting a maturation process of the mitotic poles. The decondensation of the gamma-tubulin material and of the material containing the TPH17 epitope occurred immediately after telophase. Hence in the absence of a structurally defined centrosome homologue, the microtubule nucleating material undergoes a cycle of condensation and decondensation during the cell cycle.

A single gamma-tubulin gene and mRNA, but two gamma-tubulin polypeptides differing by their binding to the spindle pole organizing centres.

Cells of eukaryotic organisms exhibit microtubules with various functions during the different developmental stages. The identification of multiple forms of alpha- and beta-tubulins had raised the question of their possible physiological roles. In the myxomycete Physarum polycephalum a complex polymorphism for alpha- and beta-tubulins has been correlated with a specific developmental expression pattern. Here, we have investigated the potential heterogeneity of gamma-tubulin in this organism. A single gene, with 3 introns and 4 exons, and a single mRNA coding for gamma-tubulin were detected. They coded for a polypeptide of 454 amino acids, with a predicted molecular mass of 50,674, which presented 64-76% identity with other gamma-tubulins. However, immunological studies identified two gamma-tubulin polypeptides, both present in the two developmental stages of the organism, uninucleate amoebae and multinucleate plasmodia. The two gamma-tubulins, called gamma s- and gamma f-tubulin for slow and fast electrophoretic mobility, exhibited apparent molecular masses of 52,000 and 50,000, respectively. They were recognized by two antibodies (R70 and JH46) raised against two distinct conserved sequences of gamma-tubulins. They were present both in the preparations of amoebal centrosomes possessing two centrioles and in the preparations of plasmodial nuclear metaphases devoid of structurally distinct polar structures. These two gamma-tubulins exhibited different sedimentation properties as shown by ultracentrifugation and sedimentation in sucrose gradients. Moreover, gamma s-tubulin was tightly bound to microtubule organizing centers (MTOCs) while gamma f-tubulin was loosely associated with these structures. This first demonstration of the presence of two gamma-tubulins with distinct properties in the same MTOC suggests a more complex physiological role than previously assumed.

Changes in phospholipids during the cell cycle of myxomycete Physarum polycephalum.

The rate of 32Pi incorporation into the main membrane phospholipid fractions, i.e. phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylinositol (PI), as well as their contents in the cells during synchronous growth of the myxomycete, Physarum polycephalum, have been studied. It has been found that both the phospholipid levels and the rates of 32Pi incorporation increase during the S phase till the early G2 phase, remain nearly constant during the G2 phase and fall to the initial level at the end of the G2 phase and in mitosis. It has been revealed that the rate of 32Pi incorporation into PC is very low compared to PE and PI, in spite of the fact that the PC level is the highest. The possible reasons of this phenomenon are discussed.

[Differences in the DNA nucleotide sequences of the nuclear matrix of the myxomycete Physarum polycephalum in the G2- and S-phases of the cell cycle]. / Razlichiia nukleotidnykh posledovatel'nostei DNK iadernogo matriksa miksomitseta Physarum polycephalum v G2- i S-fazakh kletochnogo tsikla.

The extent of homology of the nuclear matrix DNA isolated from the macroplasmodium of Physarum polycephalum at G2- and S-phases of the cell cycle was studied using the molecular reassociation method. The results obtained point to the incomplete homology of DNA samples prepared at G2- phases as compared to those obtained at S-phase. Bearing this in mind it seems reasonable to suppose that the DNA template may transfer through the nuclear matrix-attached replicative complex under the replication.

[Study of the mitochondrial Ca2+-transport system on lipid bilayer membranes (LBM)]. / Izuchenie Ca2+-transportiruiushchei sistemy mitokhondrii na bisloinykh lipidnykh membranakh (BLM)

Electrical properties of BLM formed from lipid components of rat liver mitochondria were studied. The lipids were obtained after preliminary 40 second incubation of mitochondria in the medium containing succinic acid. BLM electroconductivity in the medium with calcium chloride (8--20 mM) is 10--100 times higher than the initial electroconductivity in tris-HC1 at pH 7.5, and in the presence of potassium chloride and magnesium chloride it differes little from the initial value. The transmembrane potential difference per 10-fold concentration gradient of Ca2+, Mg2+, and K+ is approximately 27 mV, approximately 15 mV and approximately 22 mV correspondingly. By means of biionic potential method the following selectivety line for cations is formed: Ca2+ greater than Sr2 greater than or equal to Mg2+ greater than K+. La3+ ions inhibit the transport of Ca2+ through the BLM studied.