On the formation of ρ−petites in yeast

Bechmann, Helga; Krüger, Melanie; Böker, E.; Bandlow, Wolfhard; Schweyen, Rudolf J.; Kaudewitz, F.

doi:10.1007/bf00268559

Cited by 16 publications

(7 citation statements)

References 28 publications

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“…rho--clones generated by mutant tsm8 during growth in yeast/peptone/glucose medium at 35 "C are found to retain, with high frequency, deleted mtDNA [7]. These findings point to a mode of petite formation according to the second alternative above.…”

Section: Growth Ratesmentioning

confidence: 79%

“…It was shown previously [7,3 81 that growth rates of wild-type and mutant tsm8 are similar at 23 "C, regardless of the carbon source used. At 35 "C the growth rates on fermentable carbon sources are also comparable.…”

Section: Growth Ratesmentioning

confidence: 79%

“…The iso-nuclear diploid strains SM 551 tsm8 and SM 552 tsm+, SM 201 tsm8 and SM 202 tsm+, the haploid mutant strains D 12161-1 and 1211-1B and the parent strain M 12 a, i h s , trp2, ural have been used [7,8,11]. Media (yeast extract/ peptone/glycerol and yeast extract/peptone/glucose) and growth conditions (permissive temperature 23 "C ; non-permissive temperature 35 "C) were as published previously [7,8,11]. Mitochondria were prepared after breaking cells by shaking them by hand with glass beads [12], or after lysis of spheroplasts [13] as will be indicated in Results.…”

Section: Strains Cell Culture Harvesting Procedures and Preparationmentioning

confidence: 99%

“…No further budding is observed. Concomitantly, whole cell respiration decreases gradually after a short initial increase [7,18]. Fig.…”

Section: Cytochromes Respiratory Function and Energy Conservation Inmentioning

confidence: 99%

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Macromolecular Synthesis and Energy Level in a Mitochondrial Conditional Yeast Mutant, tsm-8

et al. 1977

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Mitochondrial DNA, protein and ATP syntheses persist at non-permissive temperature (35 "C) in the mitochondrial, conditionally rho-petites forming yeast mutant, tsm8. Protein and ATP syntheses, however, are diminished during prolonged incubation at 35 "C in non-fermentable substrate.Mitochondrial RNA synthesis decreases rapidly to a residual constant level of about 10 % of the initial value after the shift to 35 "C. The decrease is reversed by returning to permissive conditions. Evidence is presented that this temperature-induced decrease in mitochondrial transcription rate is effected by a mutationally altered regulatory process rather than by temperature sensitivity of mitochondrial RNA polymerase. It is concluded that rho-petite formation in mutant tsm8 is not effected by complete inhibition of macromolecular and ATP syntheses but is correlated with a reduction in mitochondrial transcription.Considerable progress towards the identification of mitochondrial genes and gene products has been made by the genetic and biochemical analysis of recently described mit-mutants of Saccharomyces cerevisiae. It has been shown that mit-mutations mapping in different regions of mitochondrial DNA (mtDNA) cause deficiencies in respiratory complexes I11 or IV or in the ATPase complex. In some mutants an alteration in, or a lack of, a (mitochondrially synthesized) protein component of one or the other of these complexes could be related to the mit-mutation Different from these mit-mutants which are deficient in one or the other functional complex, is a conditional mit-mutant previously described by this laboratoy [ 5 ] . Pleiotropic deficiencies in mitochondrial respiratory functions and a lack in several mitochondrially synthesized components of the inner membrane were observed when cells of this mutant, tsm8, were grown at 35 "C [6]. At this temperature growth on non-fermentable substrate of mutant cells was arrested, whereas on fermentable substrates growth is not limited but results in a rapid conversion of cells into rho-Detailed genetic analysis has revealed that mutation tsm8 is a mutation in a mitochondrial gene locus different from previously known mit-loci and also [l-41. Abbreviation. FCCP, carbonylcyanide p-trifluoromethoxyphenylhydrazone.different from the loci determining drug resistance/ sensitivity or coding for mitochondrial rRNA [8,9]. The analysis of mitochondrial proteins and functions affected by mutation tsm8 thus promises to lead to the identification of a new mitochondrial gene product. Furthermore, the knowledge of gene product and functions altered by mutation tsm8 may help to understand the exceptionally high rho factor instability observed in this mutant.This paper intends to present a study of mitochondrial macromolecular syntheses in mutant tsm8. The results indicate a conditional reduction in expression of part of or all mitochondrial genes in this mutant, which may cause the pleiotropic effects observed in mutant cells at 35 "C, including the exceptionally high rate of the formation of rho-peti...

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Section: Growth Ratesmentioning

confidence: 79%

Section: Growth Ratesmentioning

confidence: 79%

Section: Strains Cell Culture Harvesting Procedures and Preparationmentioning

confidence: 99%

“…No further budding is observed. Concomitantly, whole cell respiration decreases gradually after a short initial increase [7,18]. Fig.…”

Section: Cytochromes Respiratory Function and Energy Conservation Inmentioning

confidence: 99%

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Macromolecular Synthesis and Energy Level in a Mitochondrial Conditional Yeast Mutant, tsm-8

et al. 1977

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“…The Sacchnromyces cerevisiae strain SM202 [25] was grown in YPG medium (10 g/1 yeast extract, 10 g/l bacto-peptone, 10 g/l galactose, 1 g/1 glucose) in a Labroferm FS-300 New Brunswick fermentor at 28 -30°C with constant agitation and aeration until Asoo = 12 (late exponential phase). The wet cells (700 g from 36 1 medium) were harvested by a 5 -10-min centrifugation at 3000 x g , washed twice with cold demineralized water and incubated during 30 rnin at room temperature in 3.8 1 buffer A (50 mM EDTA pH 7.5, 0.75 M 2-mercaptoethanol) to disturb the cell wall.…”

Section: Mitochondria1 Trnamentioning

confidence: 99%

Purification of the yeast mitochondrial methionyl‐tRNA synthetase

Schwob

Sanni

Fasiolo

et al. 1988

European Journal of Biochemistry

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Yeast-mitochondria1 methionyl-tRNA synthetase was purified 1060-fold from mitochondrial matrix proteins of Saccharomyces cerevisiae using a four-step procedure based on affinity chromatography (heparin-Ultrogel, tRNAMe'-Sepharose, Agarose-hexyl-AMP) to yield a single polypeptide of high specific activity (1800 U/mg). Like the cytoplasmic methionyl-tRNA synthetase ( M , 85 000), the mitochondrial isoenzyme is a monomer, but of significantly smaller polypeptide size ( M , 65 000). In contrast, the corresponding enzyme of Escherichia coli is a dimer (Mr 152000) made up of identical subunits. The measured affinity constants of the purified mitochondrial enzyme for methionine and tRNA'" are similar to those of the cytoplasmic isoenzyme. However, the two yeast enzymes exhibit clearly different patterns of aminoacylation of heterologous yeast and E. coli tRNAMe'. Furthermore, polyclonal antibodies raised against the two proteins did not show any cross-reactivity by inhibition of enzymatic activity and by the highly sensitive immunoblotting technique, indicating that the two enzymes share little, if any, common antigenic determinants. Taken together, our results further support the belief that the yeast mitochondrial and cytoplasmic methionyl-tRNA synthetases are different proteins coded for by two distinct nuclear genes.Like the yeast cytoplasmic aminoacyl-tRNA synthetases, the mitochondrial enzymes displayed affinity for immobilized heparin. This distinguishes them from the corresponding enzymes of E. coli. Such an unexpected property of the mitochondrial enzymes suggests that they have acquired during evolution a domain for binding to negatively charged cellular components.The components of the yeast mitochondrial translation apparatus are of dual origin. While the mitochondrial (mt) DNA codes for a complete set of transfer RNAs and the 15s and 21s ribosomal RNAs, the proteins required for mitochondrial protein synthesis such as the ribosomal proteins, the aminoacyl-tRNA synthetases, and the translation factors are coded for by nuclear genes. The sole exception is the ribosomal protein varl which is encoded by the mt DNA (for reviews, see [l, 21).Aminoacyl-tRNA synthetases constitute a divergent family of enzymes differing in size and subunit structure but catalyzing the same reaction, the selective attachment of amino acids to their cognate tRNAs. The molecular and catalytic properties of these enzymes have been extensively studied over the past 20 years [3, 41. However, information about mitochondrial aminoacyl-t RNA synthetases remained scarce until recently. Previous studies on phenylalanyl-, leucyl-, and methionyl-tRNA synthetases from yeast mitochondria and cytoplasm indicated that the two isoenzymes exhibit distinctive properties [5 -71. By contrast, no significant differences could be observed between the mitochondrial and cytoplasmic valyl-tRNA synthetases IS]. Recently, cloning and molecular analysis of genes coding for mitochondrial aminoacyl-tRNA synthetases revealed the existence of two classes of nuclea...

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The Petite Mutation in Yeast

Whittaker¹

1979

Subcellular Biochemistry

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On the formation of ρ−petites in yeast

Cited by 16 publications

References 28 publications

Macromolecular Synthesis and Energy Level in a Mitochondrial Conditional Yeast Mutant, tsm-8

Macromolecular Synthesis and Energy Level in a Mitochondrial Conditional Yeast Mutant, tsm-8

Purification of the yeast mitochondrial methionyl‐tRNA synthetase

The Petite Mutation in Yeast

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