Naegleria’s mitotic spindles are built from unique tubulins and highlight core spindle features

ABSTRACTNaegleria gruberi is a unicellular eukaryote whose evolutionary distance from animals and fungi has made it useful for developing hypotheses about the last common eukaryotic ancestor. Naegleria amoebae lack a cytoplasmic microtubule cytoskeleton and assemble microtubules only during mitosis, and thus provides a unique system to study the evolution and functional specificity of mitotic tubulins and the resulting spindle. Previous studies showed that Naegleria amoebae express a divergent α-tubulin during mitosis and we now show that Naegleria amoebae express a second mitotic α- and two mitotic β-tubulins. The mitotic tubulins are evolutionarily divergent relative to typical α- and β- tubulins, contain residues that suggest distinct microtubule properties, and may represent drug targets for the “brain-eating amoeba” Naegleria fowleri. Using quantitative light microscopy, we find that Naegleria’s mitotic spindle is a distinctive barrel-like structure built from a ring of microtubule bundles. Similar to those of other species, Naegleria’s spindle is twisted and its length increases during mitosis suggesting that these aspects of mitosis are ancestral features. Because bundle numbers change during metaphase, we hypothesize that the initial bundles represent kinetochore fibers, and secondary bundles function as bridging fibers.

Kinetics of Chalcopyrite Leaching by Hydrogen Peroxide in Sulfuric Acid

In ores, chalcopyrite is usually associated with other sulfide minerals, such as sphalerite, galena, and pyrite, in a dispersed form, with complex mineralogical structures. Concentrates obtained by flotation of such ores are unsuitable for pyrometallurgical processing owing to their poor quality and low metal recovery. This paper presents the leaching of chalcopyrite concentrate from the location “Rudnik, Serbia”. The samples from the flotation plant were treated with hydrogen peroxide in sulfuric acid. The influences of temperature, particle size, stirring speed, as well as the concentrations of hydrogen peroxide and sulfuric acid were followed and discussed. Hence, the main objective was to optimize the relevant conditions and to determine the reaction kinetics. It was remarked that the increase in temperature, hydrogen peroxide content, and sulfuric acid concentration, as well as the decrease in particle size and stirring speed, contribute to the dissolution of chalcopyrite. The dissolution kinetics follow a model controlled by diffusion, and the lixiviant diffusion controls the rate of reaction through the sulfur layer. Finally, the main characterization methods used to corroborate the obtained results were X-ray diffraction (XRD) as well as qualitative and quantitative light microscopy of the chalcopyrite concentrate samples and the leach residue.

Applications of Modern Stereology to Quantitative Light Microscopy

Extended abstract of a paper presented at Microscopy and Microanalysis 2009 in Richmond, Virginia, USA, July 26 – July 30, 2009

Ultrastructural identification of uncoated caveolin-independent early endocytic vehicles

Using quantitative light microscopy and a modified immunoelectron microscopic technique, we have characterized the entry pathway of the cholera toxin binding subunit (CTB) in primary embryonic fibroblasts. CTB trafficking to the Golgi complex was identical in caveolin-1null (Cav1−/−) mouse embryonic fibroblasts (MEFs) and wild-type (WT) MEFs. CTB entry in the Cav1−/− MEFs was predominantly clathrin and dynamin independent but relatively cholesterol dependent. Immunoelectron microscopy was used to quantify budded and surface-connected caveolae and to identify noncaveolar endocytic vehicles. In WT MEFs, a small fraction of the total Cav1-positive structures were shown to bud from the plasma membrane (2% per minute), and budding increased upon okadaic acid or lactosyl ceramide treatment. However, the major carriers involved in initial entry of CTB were identified as uncoated tubular or ring-shaped structures. These carriers contained GPI-anchored proteins and fluid phase markers and represented the major vehicles mediating CTB uptake in both WT and caveolae-null cells.