Gottfried Schatz

DEOXYRIBONUCLEIC ACID ASSOCIATED WITH YEAST MITOCHONDRIA.

Abstract DNA is generally considered to be confined to the nucleus. Small amounts of DNA sometimes found in other cell fractions are usually attributed to contamination by nuclear material ( Allfrey 1959 ). The presence of significant amounts of extranuclear DNA occurring in the cytoplasm of the amphibian oocyte ( Stich 1962 ), the kinetosomes ( Clark and Wallace 1960 ), and in the chloroplasts of plant cells ( Chun et al. 1963 ) is well documented but is regarded rather as an exception than as a rule, because in these cases DNA is associated with cellular components absent from higher cells. This paper presents evidence that preparations of mitochondria from bakers yeast, purified by flotation in density gradients, contain a significant quantity of DNA. The amount found is far in excess of that accounted for by the polydeoxyribonucleotide component ( Appleby and Morton, 1960 ) of mitochondrial cytochrome b2. Since the mitochondrion represents an organelle present in most cells, including those of mammals, the occurrence of DNA in yeast mitochondria suggests the possibility, that extranuclear DNA is much more common that hitherto suspected Mitochondrial function in the yeast cell is in part controlled by extrachromosomal genetic factors ( Ephrussi and Hottinguer 1951 ), whose exact chemical nature is unknown. The presence of DNA in yeast mitochondria is thus of special interest.

TRIPHOSPHOPYRIDINE NUCLEOTIDE: CYTOCHROME C REDUCTASE OF SACCHAROMYCES CEREVISIAE: A "MICROSOMAL" ENZYME.

Abstract 1. 1. The intracellular location and function of TPNH: cytochrome c reductase (EC 1.6.2.3) and of DPNH; cytochrome c reductase system in the yeast cell have been investigated. 2. 2. Both enzyme activities are associated with particles. 3. 3. Homogenates of anaerobically grown yeast and of the “petite” mutant, both of which are unable to respire, exhibit practically the same specific TPNH: cytochrome c reductase activity as homogenates of respiring bakers yeast. TPNH: cytochrome c reductase activity in antimycin-insensitive, does not interact with endogenous cytochrome c of yeast mitochondria and is predominantly associated with microsomal particles which have not yet been described for yeast. Electron micrographs of these particles are presented. 4. 4. In contrast, DPNH: cytochrome c reductase activity, which is greatly lowered in homogenates of anaerobic or “peptide” mutant yeast, is 95% inhibited by antimycin and interacts rapidly with endogenous cytochrome c of yeast mitochondria. More than 95% of this reductase activity is mitochondrial, the remainder being located in microsomes. Microsomal DPNH: cytochrome c reductase activity is not inhibited by antimycin. 5. 5. It is concluded, that TPNH: cytochrome c reductase of Saccharomyces cerevisine is not a respiratory-chain enzyme. Some implications of these findings are discussed.

The isolation of possible mitochondrial precursor structures from aerobically grown baker'-s yeast

Abstract At present, the steps involved in the formation of mitochondria are unknown, primarily because of the lack of a suitable experimental system, since most cells containing mitochondria will not tolerate any extensive modification of their respiratory apparatus. However, such limitations do not apply to the facultative anaerobe, Saccharomyces cerevisiae , whose respiratory system can be drastically altered either by mutation or by environmental conditions, even though the electron transfer steps of aerobic yeast as well as its mitochondria are quite similar to those of mammalian cells (Slonimski 1953; Chance 1959; vitols et al. 1961). Results from earlier work failed to provide evidence in favor of a genetic continuity of yeast mitochondria (Schatz et al. 1963). Thus, de novo synthesis of yeast mitochondria appeared possible. In a preliminary communication, Linnane et al. reported the absence of mitochondria in anaerobically grown cells of Torulopsis utilis . An intracellular membrane system was identified in these cells, tentatively designated by the authors as a mitochondrial precursor system (Linnane et al. 1962). The present report deals with the identification and isolation of as yet unknown cytoplasmic particles from aerobic cells of S. cerevisiae . The properties of these particles are consistent with those expected for mitochondrial precursors. The experimental approach was based on previous studies by Ephrussi et al. who had shown that glucose specifically interferes with mitochondrial development in these cells (Ephrussi et al. 1956). Aerobic growth in various concentrations of glucose thus enabled the large scale isolation of cells with a graded development of their mitochondrial apparatus.

Intramitochondriale Deoxyribonukleinsäure in Säugetierzellen

Highly purified mitochondria from liver and kidney of the rat as well as from bovine heart have been found to contain from 0.17 to 0.57 μg deoxyribonucleic acid per mg mitochondrial protein. This amount of DNA is attributable neither to contamination of the mitochondrial fractions by nuclei nor to adsorption artefacts. Purification of the mitochondria was achieved by differential centrifugation and subsequent flotation in a density gradient of “Urografin”, an X-ray contrasting agent. It is suggested that deoxyribonucleic acid is a universal component of genetical importance in all mitochondria.

The Oxygen-Induced Synthesis of Respiratory Enzymes

Many organisms require oxygen in order to live. Others, however, are poisened by this element and fail to grow if its concentration in the environment reaches a critical threshold level. Still another group of organisms can grow both in the presence and absence of oxygen. This last group is collectively termed “facultative anaerobes” and forms the subject of the present discussion.

Extranukleare Desoxyribonukleinsäure in aerob und anaerob gezüchteter normaler Bäckerhefe sowie in der atmungsdefizienten „petite”-Mutante

Pankreas-DNAase** greift bei niedriger Temperatur die in Hefemitochondrien vorhandeneDNA nicht merkbar an, wahrend verunreinigende nicht-mitochondrialeDNA zu saureloslichen Bruchstucken abgebaut wird. Eine darauf beruhende Methode eignet sich zur selektiven Bestimmung mitochondrialerDNA in Mitochondrienfraktionen, die durch differentielles Zentrifugieren von Hefehomogenaten gewonnen worden und noch mit Zellkernmaterial verunreinigt sind.

Intrazelluläre Lokalisierung und Funktion von NADPH-Cytochromc-Reduktase in Saccharomyces cerevisiae

Das Flavoprotein NADPH-Cytochromc-Reduktase aus Hefe ist im Gegensatz zu bisherigen Anschauungen kein Enzym der mitochondralen Atmungskette, sondern an bisher unbekannte subzellulare Partikel gebunden, die nach Homogenisieren der Hefezellen von den Mitochondrien abgetrennt werden konnen. Die Beteiligung von Cytochromb2 oder einer Transhydrogenase-reaktion an der Elektronenubertragung von NADPH auf Cytochromc in einem Hefehomogenat konnte ausgeschlossen werden. Die Natur der neuen Hefepartikel sowie die physiologische Funktion des Enzyms NADPH-Cytochromc-Reduktase werden diskutiert.