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M Osumi

Publications and source records attributed to M Osumi.

At least 19 recordsLinked to original sources
PubMed1992

Beta-oxidation of butyrate, the short-chain-length fatty acid, occurs in peroxisomes in the yeast Candida tropicalis.

When an n-alkane-utilizable yeast, Candida tropicalis pK233, was cultivated on butyrate, the fatty acid of shortest chain-length for beta-oxidation, as the sole source of carbon and energy, catalase and the enzymes of the fatty acid beta-oxidation system were inducibly synthesized at high levels. As in the alkane-grown cells, the proliferation of peroxisomes was harmonized with the induction of peroxisomal enzymes. The results of subcellular fractionation and immunoelectronmicroscopy indicated the localization of these enzymes in peroxisomes, not in mitochondria. It was suggested that only peroxisomes have a role in fatty acid beta-oxidation in the yeast cells, unlike in mammalian cells, in which cooperation between peroxisomes and mitochondria is essential.

Acetyl-CoA C-Acyltransferase↗
PubMed1992

Physiological roles of acetoacetyl-CoA thiolase in n-alkane-utilizable yeast, Candida tropicalis: possible contribution to alkane degradation and sterol biosynthesis.

The presence of two types of thiolases, acetoacetyl-CoA thiolase and 3-ketoacyl-CoA thiolase, was demonstrated in peroxisomes of n-alkane-grown Candida tropicalis [Kurihara, T., Ueda, M., & Tanaka, A. (1989) J. Biochem. 106, 474-478], while acetoacetyl-CoA thiolase was also shown to be present in cytosol. The activity of the enzyme in cytosol was constant irrespective of culture conditions, while the peroxisomal enzyme was inducibly synthesized in the alkane-grown yeast cells. These results indicate that peroxisomal acetoacetyl-CoA thiolase participates in alkane degradation, while the cytosolic enzyme is associated with other fundamental metabolic processes, probably sterol biosynthesis, because this enzyme can catalyze the first step of the sterol biosynthesis. 3-Hydroxy-3-methylglutaryl (HMG)-CoA reductase, a key regulatory enzyme of sterol biosynthesis, was found to be localized exclusively in microsomes of the alkane-grown yeast cells. These results suggest that yeast peroxisomes do not contribute to sterol biosynthesis, unlike the case of mammalian cells.

Acetyl-CoA C-Acetyltransferase↗
PubMed1992

Morphological changes associated with growth inhibition of Trichophyton mentagrophytes by amorolfine.

Amorolfine inhibited the in-vitro growth of Trichophyton mentagrophytes to some extent at a low drug concentration of 0.8 ng/ml. Corresponding to the growth inhibition, SEM studies revealed a slight modification of hyphal morphology, i.e. a waving of the hyphal surface. These morphological alterations were more extensive with increases in drug concentration and treatment period: collapsed and distorted hyphae and exfoliation of the surface of T. mentagrophytes occurred at 8 ng/ml and marked deformation and disruption of the hyphal structure at 80 ng/ml of amorolfine. TEM revealed thickening of the cell walls and the accumulation of electron-dense granular structures in both the wall and cytoplasm in thin-sectioned cells pretreated with 8 ng/ml or more of amorolfine, although the nuclear and mitochondrial architecture was not noticeably influenced. Cytoplasmic membranes and other membranous structures of organelles such as nuclei and mitochondria were disrupted or fused, thereby losing their essential physiological activity in hyphal cells pretreated with 80 ng/ml of amorolfine. The ultrastructural study thus supports the observation that morphological changes of T. mentagrophytes caused by amorolfine were associated with its growth-inhibitory and killing activity, which depended on the drug concentration and treatment time.

Antifungal Agents↗
PubMed1992

Immunoelectron microscopic localization of thiolases, beta-oxidation enzymes of an n-alkane-utilizable yeast, Candida tropicalis.

The location of acetoacetyl-CoA thiolase (T-I) and 3-ketoacyl-CoA thiolase (T-III), enzymes of the fatty acid beta-oxidation system, was studied in n-alkane-grown Candida tropicalis cells by immunoelectron microscopy using a post-embedding method with colloidal gold conjugated IgG. The deposition of gold particles for T-I was detected in the microbodies and cytoplasm and that of gold particles for T-III specifically in the microbodies. The double labeling technique confirmed that T-I and T-III occurred concurrently in a microbody and T-I also in cytoplasm. These results were consistent with the biochemical data based on subcellular fractionation and indicated that the yeast beta-oxidation system operates efficiently only in the microbodies.

Acetyl-CoA C-Acetyltransferase↗
PubMed1991

Electron microscopic observation of recombinant Escherichia coli cells overproducing human tumor necrosis factor-alpha mutant as inclusion bodies.

Escherichia coli C600 r-m- carrying plasmid pTNF483 (E. coli [pTNF483]) produces a tumor necrosis factor-alpha (TNF-alpha) mutant protein in an insoluble form. A swollen region was observed in the SEM images to encircle the outside of most of the E. coli [pTNF483] cells just like a bandage. On the other hand, inclusion bodies of the TNF-alpha mutant as large as the short axis of the cell were observed in TEM images. This position was regarded as coinciding with the swollen region of SEM images. The inclusion bodies revealed in the swollen region of the cell envelopes were clearly observed in SEM images of isolated insoluble structures obtained by centrifugal sedimentation of a sonicated cell suspension.

Escherichia coli↗
PubMed1989

Serial section reconstruction using a computer graphics system: applications to intracellular structures in yeast cells and to the periodontal structure of dogs' teeth.

A computer graphics system for reconstruction from serial section micrographs was applied to intracellular details of a yeast target cell (Saccharomyces cerevisiae cell) induced by the alpha factor mating pheromone and was also applied to a periodontal structure of a dog tooth moved orthodontically. In the former, intracellular organelles and a distribution of vesicles could be clearly observed through the cell membrane using the transparent display method in which the smoothing of the reconstructed outer cell membrane surface by computer processing was applied to the transparent display. In the latter case, by cutting through a reconstructed dog tooth and its periodontal tissues, labiolingual and mesiodistal cut surfaces of the tooth and of adjacent alveolar bone could be observed with fine details (232 sections were used).

Animals↗
PubMed1989

Three-dimensional analysis of morphogenesis induced by mating pheromone alpha factor in Saccharomyces cerevisiae.

Ultrastructural analyses of cytoplasmic changes in Saccharomyces cerevisiae X2180-1A (MATa) that had been treated with alpha factor were performed by using the freeze-substitution fixation method. After alpha factor treatment, cells exhibited a pointed projection, which is a unique pattern of oriented cell surface growth. The relationship between projection formation and intracellular organelles was examined using serial thin sections and computer-aided three-dimensional reconstructions. Using these analyses membrane vesicles and other organelles were detected, and studies on their dynamic structural reorganization became feasible. Production of membrane vesicles (average 65 nm in diameter) was induced upon exposure of the cells to alpha factor before projection emergence. The total number of membrane vesicles increased at the early stage and decreased at the late stage of projection formation. Three-dimensional analysis indicated that the vesicles were at first dispersed throughout the cell, then accumulated at the site where the projection formed. Morphological changes and multiplication of the Golgi body were seen during the process of projection formation. Other intracellular organelles (nucleus, vacuole, rough endoplasmic reticulum and mitochondria) were also rearranged, showing a polar organization of the cytoplasm during projection formation.

Mating Factor↗
PubMed1989

Actin is associated with the formation of the cell wall in reverting protoplasts of the fission yeast Schizosaccharomyces pombe.

To clarify the involvement of actin in the formation of the yeast cell wall, reverting protoplasts of Schizosaccharomyces pombe were used as a simple model system. Actin of reverting protoplasts was labeled with rhodamine-conjugated phalloidin and observed by conventional fluorescence microscopy and laser scanning confocal microscopy. A close spatial as well as temporal relationship between actin and cell wall formation was observed in protoplast reversion. That is, the site of actin 'dots' in the reverting protoplasts coincided with the site of new wall formation and the timing of rearrangement of actin coincided with the initiation of cell wall formation and with the timing of cell wall expansion. Treatment of reverting protoplasts with cytochalasin D (CD) further clarified the close relationship between actin and cell wall organization. The effect of CD was dose dependent. A high dose of CD caused the absence of actin as well as the complete inhibition of cell wall formation. A low dose of CD caused weakly stained unlocalized actin, which induced grossly aberrant cell wall deposition as well as substantial changes in the morphology of the reverting protoplasts. These results demonstrated that actin is associated with initiation of cell wall formation, the proper deposition of cell wall materials, and maintaining the normal morphology of reverting protoplasts. Scanning electron microscopy revealed the presence of a fibrillar net structure on the surface of non-treated control reverting protoplasts. However, the absence of a fibrillar network on the surface of reverting protoplasts was observed in the presence of a high concentration of CD. Lack of localization of microfibrils as well as poor development of the fibrillar network were also observed in the presence of a low concentration of CD. Recovery experiments confirmed the close relationship between actin and cell wall formation.

Actins↗
PubMed1989

Cell wall formation in regenerating protoplasts of Schizosaccharomyces pombe: study by high resolution, low voltage scanning electron microscopy.

The ultrastructure of regenerating cell wall in Schizosaccharomyces pombe protoplasts was studied with a high resolution, low voltage scanning electron microscope (LVSEM). In contrast to the transmission electron microscopy, the LVSEM images give three-dimensional information on the cell wall regeneration in yeast protoplasts. We found that, after only a few minutes of incubation, the protoplasts began to show protuberances in a unipolar manner, and a fibrilar network was formed asymmetrically which covered the whole surface of the protoplasts after 5 hr. The network consisted of microfibrils about 8 to 10 nm wide, forming flat and wavy bundles of various widths and lengths, up to about 200 nm wide and 1 micron long, mainly made of yeast glucan. Free ends of microfibrils were seldom found. Interfibrillar spaces were progressively filled with granular particles and finally the complete cell wall was formed after 12 hr. The fibrillar network was destroyed by the digestion with beta (1----3)-glucanase. When protoplasts were regenerating in the presence of aculeacin A, the fibrillar networks were not formed, resulting in incomplete cell wall formation. These observations suggest that beta-glucan is the main component of the microfibrils and that it plays an important role in the formation of the cell wall in S. pombe.

Antifungal Agents↗
PubMed1987

Effect of aculeacin A on reverting protoplasts of Candida albicans.

Protoplasts of Candida albicans were prepared by digestion with Zymolyase and the effect of aculeacin A, a wall-active antibiotic, on the synthesis of microfibrillar structures by the protoplasts incubated for 5 hr in an osmotically stabilized medium was studied using several electron microscopical techniques. Chemical analyses of the protoplasts before and after reversion with or without the antibiotic were also performed. Aculeacin A not only inhibited synthesis of microfibrils mainly composed of alkali-insoluble beta-glucan, but also their assembly to form thicker bundles. The antibiotic appeared to have no effect on other wall components constituting the surface structure of reverting protoplasts. These data confirmed our previous postulation that aculeacin A is a specific and potent inhibitor of beta-glucan synthesis as well as biogenesis of cell walls including beta-glucans in susceptible yeasts.

Alkalies↗
PubMed1986

Properties of catalase purified from a methanol-grown yeast, Kloeckera sp. 2201.

Catalase, a marker enzyme of peroxisomes, was purified to homogeneity from whole cells of Kloeckera sp. 2201 (a strain of Candida boidinii) grown on methanol by means of ammonium sulfate fractionation followed by hydroxyapatite, Sephacryl S-300 and DEAE-Sepharose column chromatographies. Crystallized catalase was brown-coloured and needle-like. The molecular mass of the enzyme was about 240 000 daltons consisting of four identical subunits of 62 000 daltons. The minimum size of catalase molecule was estimated to be about 6 X 10 nm from an electron micrograph. Judging from the absorption spectrum, the enzyme seemed to belong to a group of T-type catalase. The Km value of the enzyme for hydrogen peroxide (catalatic activity) was 25 mM, while that for methanol (peroxidatic activity) was 83 mM. Catalase from Kloeckera sp. cells showed a certain degree of similarity to the enzyme purified from alkane-grown Candida tropicalis [T. Yamada et al. (1982) Eur. J. Biochem. 125, 517-521 and 129, 251-255] in its immunochemical properties.

Animals↗
PubMed1985

Effect of aculeacin A, a wall-active antibiotic, on synthesis of the yeast cell wall.

A wall-active, amphophilic antibiotic aculeacin A significantly but incompletely inhibited in vitro the activity of beta-(1,3)glucan synthase prepared from highly susceptible yeasts Saccharomyces cerevisiae and Candida albicans. In contrast, comparable cell-free preparations from S. cerevisiae active in chitin synthase or mannan synthase were insensitive to the antibiotic, suggesting selectivity of its action in synthesis of the yeast cell wall. An electron microscopic study of the effects of aculeacin A at 0.31 micrograms/ml, the optimally active concentration, on osmotically stabilized C. albicans cells revealed morphological alterations in both cell walls and cell membranes. Deformation in contour and derangement of the layered structure of the cell wall were prominent. In addition, massive fibrous material of beta-glucan-like microfibrils was occasionally extruded from the cell surface. Accompanying this effect on the cytology of the cell wall, ultrastructural and functional impairment of the cell membrane was demonstrated by transmission and freeze-fracture electron microscopic techniques. These data suggest that aculeacin A affects synthesis of the yeast cell wall through not only selective blockage of beta-(1,3)glucan synthase, as a result of a primary interaction with the cell membrane, but also inhibition of the fabrication of beta-glucan or other wall components into well-organized cell walls.

Antifungal Agents↗