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Ribosomes Associated with the Mitochondrial Surface of the Yeast Rhodotorula minuta1)
Biochem. Physiol. Pflanzen 184,471-473 (1989) VEB Gustav Fischer Verlag Jena
Short Communication
Ribosomes Associated with the Mitochondrial Surface of the Yeast Rhodotorula minuta1) CH . BEGEMANN, A. SCHMIDT and U. KRISTEN Institut für Allgemeine Botanik der Universität Hamburg, Hamburg, F.R.G. Key Term Index: Mitochondria, ribosomes; Rhodotorula minuta
Summary Electron microscopical observations of the yeast Rhodotorula minuta HARRISON, aerobically and unaerobically cultured in liquid agar, revealed a unique covering of the mitochondrial surface by presumptive 80S ribosomes. Their pattern of distribution and their association to the outer mitochondrial membrane are emphasized in comparison with the ribosom al arrangement of the endoplasmic reticulum of higher plant cells. The endoplasmic reticulum and its derivatives were thought to be the only compartements to which ribosomes are commonly attached. This assumption is subject to restriction because ribosomes were observed in close association with the acrosome membranes in Guinea pig spermatids (MOLLENHAUER and MORRE 1978), with the mitochondrial surface in spores of the slime mouldDictyostelium discoideum (COTTER et a1. 1969) and in some yeast species (W ATSON 1972; KEYHANI 1973; KELLEMS and BUTOW 1972; KELLEMS eta1. 1974). Because of the importance of these observations for the understanding of protein transfer across membranes, we looked for further occurrence of such associations in yeast species, and we found mitochondria covered with ribosome-like particles in Rhodotorula minuta HARRISON. Cells ofthe yeast were removed from dried pear fruits and were aerobically or unaerobicaIly cultured in liquid Kimmig agar (KREGER-VAN Ru 1984) for 48 h. The cells were then prepared for electron microscopy as folIows: Fixation with phosphate-buffered 2.5% glutaraldehyde (pH 7.0,20 °C, 4h); after washing, postfixation with phosphate-buffered 1.0% osmium tetroxide (pH 7.0, 20°C, 4 h); blockstaining with uranylacetate (8°C, 12 h); dehydration in an ethanol series; embedding in Spurr's medium (SPURR 1969). The ultrathin sections were poststained with lead citrate and observed with a Philips electron microscope EM201. The electron micrographs, parti all y obtained from serial sections of different developmental stages of both aerobically and unaerobically cultured cells, revealed a unique covering of the surface of all mitochondria by particles wh ich correspond with 80 S ribosomes in their diameter, shape and electron density (Figs. 1, 2, 3). The distribution of these particles and their association with the outer membrane, however, differed from what is known about endoplasmic reticulurnassociated ribosomes. The latter form typical polysomes (groups with a spiral pattern, Fig. 5) J) This articIe is based on a doctoral study by CH. BEGEMANN in the Faculty of Biology, University of Hamburg
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Figs. 1-5. Ribosomes associated with the outer mitochondrial membrane of the yeast Rhodotorula minuta. Fig . 1. Portion ofthe cell containing the Ilucleus (N) mitocholldria (M) and rough endoplasmic reticulum (ER) . X 30,000. Fig. 2. Detail of the mitochondrial envelope with ribosomes associated with the outer membrane. The ribosomes are less closely attached to the membrane (arrows) than to the ER (see Fig. 4) . X 180,000. Fig . 3. The ribosomes associated with the mitochondria are evenly distributed on the membrane surface (encircled area) in contrast to the ER-attached ribosomes whichform spirals (see Fig. 5). X 100,000. Fig. 4. Rough endoplasmic reticulum of a higher plant cell. X 53,000. Fig. 5. Polyribosomes attached to the surface of the endoplasmic reticulum of a higher plant cello X 70,000.
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and are elosel y attached to the membrane (Fig. 4), whereas the particles described here appear to be evenly distributed on the membrane surface (Fig. 3) and show a small space between them and the membrane (Fig. 2). Biochemical studies of similarribosome-like partieles bound to the outer mitochondrial membrane of Saccharomyces cerevisiae provided evidence that these partieles were cytoplasmic 80 S ribosomes (KELLEMS and BUTOW 1972). They revealed in vitro protein synthesis which could be inhibited by cyeloheximide but not by chloramphenicol. Moreover, they were labelled to the same extent with 32Pi as the cytoplasmic ribosomes. In addition, binding sites for 80 S ribosomes were detected on the surface of the yeast mitochondria (KELLEMS and BUTOW 1972, 1974). From the results of both the biochemical studies (KELLEMS and BUTOW 1972, 1974; KELLEMS etal. 1974) and our electron microseopie analysis we suggest that the particles associated with the outer mitochondrial membrane are cytoplasmic 80 S ribosomes. This suggestion is supported by several observations of at least temporary continuities between the outer mitochondrial membrane and portions of the ER in animal, plant and fungal cells (FRANKE andKARTENBECK 1971; MORREet al. 1971; BRACKER andGRovE 1971). This indicates thatthe mitochondrial envelope is partially compatible with the ER membrane.
References BRACKER, C. E., and GROVE, S. N.: Continuity between cytoplasmic endomembranes and outer mitochondrial membranes in fungi. Protoplasma 73, 15-34 (1971). COTTER, D. A., MIURA-SANTO, L. Y., and HOHL, H. R.: Ultrastructural changes during germination of Dictyostelium discoideum spores. J. Bacterio!. 100, 1020-1026 (1969). FRANKE, W. W., and KARTENBECK, J.: Outer mitochondrial membrane continuous with endoplasmic reticulum. Protoplasma 73,35-41 (1971). KELLEMS, R. E., and BUTow, R. A.: Cytoplasmic-type 80S ribosomes associated with yeast mitochondria. I. Evidence for ribosome binding sites of yeast mitochondria. J. Bio!. Chem. 247, 8043-8050 (1972). KELLEMS, R. E., and BUTow, R. A.: Cytoplasmic type 80 S ribosomes associated with yeast mitochondria. III. Changes in the amount ofbound ribosomes in response to changes in metabolie state. J. Bio!. ehern. 249, 3304-3310 (1974). KELLEMS, R. E., ALLISON, Y. F., and BUTow, R. A.: Cytoplasmic type 80 S ribosomes associated with yeast mitochondria. 11. Evidence for the association of cytoplasmic ribosomes with the outer mitochondrial membrane in situ. J. Bio!. Chem. 249, 3297 - 3303 (1974). KEYHANI. E.: Ribosomal granules associated with outer mitochondrial membrane in aerobic yeast cells. J. Cell Bio!. 58,480-484 (1973). KREGER-YAN Ru, N. Y. W.: The Yeasts: a Taxonomie Study. Elsevier Science Publishers B.Y .. Oxford 1984. MOLLENHAUER, H. H., and MORRE, D. J.: Polyribosomes associated with forming acrosome membranes in guinea pig spermatides. Science 200,85-86 (1978). MORRE, D. J., MERRITT, W. D., and LEMBI, C. A.: Connections between mitochondria andendoplasmic reticulum in rat liver and on ion stern. Protoplasma 73,43-49 (1971). SPURR, A. R.: A low-viscosity epoxy resin embedding medium for electron microscopy. 1. Ultrastruct. Res. 26, 31-43 (1969). WATSON, K.: The organization of ribosomal granules within mitochondrial structures of aerobic and an aerobic cells of Saccharomyces cerevisiae. J. Cell Bio!. 55, 721-726 (1972). Received June 21, 1988: accepted October 3, 1988 Authors' address: CHRISTA BEGEMANN , Prof. Dr. ALEXANDER SCHMIDT und Prof. Dr. U DO KRISTEN (reprint requests), Institut für Allgemeine Botanik, Universität Hamburg. Ohnhorststraße 18, D-2000 Hamburg 52. 31
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Short Communication
Ribosomes Associated with the Mitochondrial Surface of the Yeast Rhodotorula minuta1) CH . BEGEMANN, A. SCHMIDT and U. KRISTEN Institut für Allgemeine Botanik der Universität Hamburg, Hamburg, F.R.G. Key Term Index: Mitochondria, ribosomes; Rhodotorula minuta
Summary Electron microscopical observations of the yeast Rhodotorula minuta HARRISON, aerobically and unaerobically cultured in liquid agar, revealed a unique covering of the mitochondrial surface by presumptive 80S ribosomes. Their pattern of distribution and their association to the outer mitochondrial membrane are emphasized in comparison with the ribosom al arrangement of the endoplasmic reticulum of higher plant cells. The endoplasmic reticulum and its derivatives were thought to be the only compartements to which ribosomes are commonly attached. This assumption is subject to restriction because ribosomes were observed in close association with the acrosome membranes in Guinea pig spermatids (MOLLENHAUER and MORRE 1978), with the mitochondrial surface in spores of the slime mouldDictyostelium discoideum (COTTER et a1. 1969) and in some yeast species (W ATSON 1972; KEYHANI 1973; KELLEMS and BUTOW 1972; KELLEMS eta1. 1974). Because of the importance of these observations for the understanding of protein transfer across membranes, we looked for further occurrence of such associations in yeast species, and we found mitochondria covered with ribosome-like particles in Rhodotorula minuta HARRISON. Cells ofthe yeast were removed from dried pear fruits and were aerobically or unaerobicaIly cultured in liquid Kimmig agar (KREGER-VAN Ru 1984) for 48 h. The cells were then prepared for electron microscopy as folIows: Fixation with phosphate-buffered 2.5% glutaraldehyde (pH 7.0,20 °C, 4h); after washing, postfixation with phosphate-buffered 1.0% osmium tetroxide (pH 7.0, 20°C, 4 h); blockstaining with uranylacetate (8°C, 12 h); dehydration in an ethanol series; embedding in Spurr's medium (SPURR 1969). The ultrathin sections were poststained with lead citrate and observed with a Philips electron microscope EM201. The electron micrographs, parti all y obtained from serial sections of different developmental stages of both aerobically and unaerobically cultured cells, revealed a unique covering of the surface of all mitochondria by particles wh ich correspond with 80 S ribosomes in their diameter, shape and electron density (Figs. 1, 2, 3). The distribution of these particles and their association with the outer membrane, however, differed from what is known about endoplasmic reticulurnassociated ribosomes. The latter form typical polysomes (groups with a spiral pattern, Fig. 5) J) This articIe is based on a doctoral study by CH. BEGEMANN in the Faculty of Biology, University of Hamburg
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Figs. 1-5. Ribosomes associated with the outer mitochondrial membrane of the yeast Rhodotorula minuta. Fig . 1. Portion ofthe cell containing the Ilucleus (N) mitocholldria (M) and rough endoplasmic reticulum (ER) . X 30,000. Fig. 2. Detail of the mitochondrial envelope with ribosomes associated with the outer membrane. The ribosomes are less closely attached to the membrane (arrows) than to the ER (see Fig. 4) . X 180,000. Fig . 3. The ribosomes associated with the mitochondria are evenly distributed on the membrane surface (encircled area) in contrast to the ER-attached ribosomes whichform spirals (see Fig. 5). X 100,000. Fig. 4. Rough endoplasmic reticulum of a higher plant cell. X 53,000. Fig. 5. Polyribosomes attached to the surface of the endoplasmic reticulum of a higher plant cello X 70,000.
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and are elosel y attached to the membrane (Fig. 4), whereas the particles described here appear to be evenly distributed on the membrane surface (Fig. 3) and show a small space between them and the membrane (Fig. 2). Biochemical studies of similarribosome-like partieles bound to the outer mitochondrial membrane of Saccharomyces cerevisiae provided evidence that these partieles were cytoplasmic 80 S ribosomes (KELLEMS and BUTOW 1972). They revealed in vitro protein synthesis which could be inhibited by cyeloheximide but not by chloramphenicol. Moreover, they were labelled to the same extent with 32Pi as the cytoplasmic ribosomes. In addition, binding sites for 80 S ribosomes were detected on the surface of the yeast mitochondria (KELLEMS and BUTOW 1972, 1974). From the results of both the biochemical studies (KELLEMS and BUTOW 1972, 1974; KELLEMS etal. 1974) and our electron microseopie analysis we suggest that the particles associated with the outer mitochondrial membrane are cytoplasmic 80 S ribosomes. This suggestion is supported by several observations of at least temporary continuities between the outer mitochondrial membrane and portions of the ER in animal, plant and fungal cells (FRANKE andKARTENBECK 1971; MORREet al. 1971; BRACKER andGRovE 1971). This indicates thatthe mitochondrial envelope is partially compatible with the ER membrane.
References BRACKER, C. E., and GROVE, S. N.: Continuity between cytoplasmic endomembranes and outer mitochondrial membranes in fungi. Protoplasma 73, 15-34 (1971). COTTER, D. A., MIURA-SANTO, L. Y., and HOHL, H. R.: Ultrastructural changes during germination of Dictyostelium discoideum spores. J. Bacterio!. 100, 1020-1026 (1969). FRANKE, W. W., and KARTENBECK, J.: Outer mitochondrial membrane continuous with endoplasmic reticulum. Protoplasma 73,35-41 (1971). KELLEMS, R. E., and BUTow, R. A.: Cytoplasmic-type 80S ribosomes associated with yeast mitochondria. I. Evidence for ribosome binding sites of yeast mitochondria. J. Bio!. Chem. 247, 8043-8050 (1972). KELLEMS, R. E., and BUTow, R. A.: Cytoplasmic type 80 S ribosomes associated with yeast mitochondria. III. Changes in the amount ofbound ribosomes in response to changes in metabolie state. J. Bio!. ehern. 249, 3304-3310 (1974). KELLEMS, R. E., ALLISON, Y. F., and BUTow, R. A.: Cytoplasmic type 80 S ribosomes associated with yeast mitochondria. 11. Evidence for the association of cytoplasmic ribosomes with the outer mitochondrial membrane in situ. J. Bio!. Chem. 249, 3297 - 3303 (1974). KEYHANI. E.: Ribosomal granules associated with outer mitochondrial membrane in aerobic yeast cells. J. Cell Bio!. 58,480-484 (1973). KREGER-YAN Ru, N. Y. W.: The Yeasts: a Taxonomie Study. Elsevier Science Publishers B.Y .. Oxford 1984. MOLLENHAUER, H. H., and MORRE, D. J.: Polyribosomes associated with forming acrosome membranes in guinea pig spermatides. Science 200,85-86 (1978). MORRE, D. J., MERRITT, W. D., and LEMBI, C. A.: Connections between mitochondria andendoplasmic reticulum in rat liver and on ion stern. Protoplasma 73,43-49 (1971). SPURR, A. R.: A low-viscosity epoxy resin embedding medium for electron microscopy. 1. Ultrastruct. Res. 26, 31-43 (1969). WATSON, K.: The organization of ribosomal granules within mitochondrial structures of aerobic and an aerobic cells of Saccharomyces cerevisiae. J. Cell Bio!. 55, 721-726 (1972). Received June 21, 1988: accepted October 3, 1988 Authors' address: CHRISTA BEGEMANN , Prof. Dr. ALEXANDER SCHMIDT und Prof. Dr. U DO KRISTEN (reprint requests), Institut für Allgemeine Botanik, Universität Hamburg. Ohnhorststraße 18, D-2000 Hamburg 52. 31
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