Protein-synthesizing Structures associated with Mitochondria

Perlman, Stanley; Penman, Sheldon

doi:10.1038/227133a0

Cited by 72 publications

(22 citation statements)

References 43 publications

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“…4. The sedimentation characteristics of the mitochondrial protein-synthesizing structures have been described (3). The cells are labeled with radioactive methionine under conditions in which cytoplasmic protein synthesis is inhibited and only mitochondrial protein synthesis occurs (26).…”

Section: Resultsmentioning

confidence: 99%

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Mitochondrial Protein Synthesis: RNA with the Properties of Eukaryotic Messenger RNA

Perlman

Abelson

Penman

1973

Proc. Natl. Acad. Sci. U.S.A.

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A heterogeneous RNA fraction with properties resembling those of messenger RNA was identified in mammalian mitochondria. Synthesis of contaminating RNA of nuclear origin was suppressed by treatment with camptothecin. Labeling of the messenger-like RNA is completely inhibited by ethidium bromide, a specific inhibitor of mitochondrial functions.Although mitochondrial protein synthesis resembles that of prokaryotes in several regards, the messenger-like RNA is covalently linked to poly(adenylic acid) [poly(A)J. Poly(A) has thus far been found only in eukaryotic cells. The poly(A) segment has a gel electrophoretic mobility of about 4 S, corresponding to a length of 50-80 nucleotides, and thus resembles in size the poly(A) found in some mammalian viral RNAs. The messenger RNA can be released from the mitochondrial protein-synthesizing structure by treatment with puromycin.Mitochondria of eukaryotic cells have a distinct system of protein synthesis that is quite dissimilar from that found in the cellular cytoplasm. The ribosome-like structure of mitochondria is relatively small (60 S) (1-4). Mitochondrial protein synthesis is sensitive to chloramphenicol, an inhibitor of prokaryotic ribosomal function, and insensitive to cycloheximide, an inhibitor of eukaryotic protein synthesis. NFormyl-methionine has been found in mitochondria and is presumed to initiate polypeptide synthesis as in bacteria (5, 6).These properties of mitochondrial protein synthesis suggested a similarity to prokaryotic rather than eukaryotic mechanisms, and there has been speculation about a possible origin of mitochondria in a primordial prokaryotic symbiont.In this report, we describe an RNA species synthesized in mitochondria with properties that suggest it serves as messenger RNA for mitochondrial protein synthesis. Surprisingly, this RNA contains polyadenylate segments, which have, so far, been found only in the messenger RNA of eukaryotic cells (7)(8)(9)(10)(11) Analysis of Total Mitochondrial RNA. Cellular extracts were prepared and analyzed as described (13). Briefly, cells were washed and then suspended in 1 ml of RSB buffer [10 mM NaCl-1.5 mM MgCl2-10 mM Tris* HCO (pH 7.4)] before they were broken in a Dounce homogenizer. Nuclei were removed by centrifugation at 800 X g for 2 min. The supernatant was again centrifuged at 7700 X g for 8 min. This pellet (mitochondrial fraction) was resuspended in sodium dodecyl sulfate (SDS) buffer and extracted by the hot phenol-SDS method (14). The RNA was analyzed by electrophoresis on 2.7% acrylamide gels for 3.5 hr at 7.5 V/cm. Gels were fractionated as described (15).Analysis of Poly(A)-Containing RNA. Mitochondrial RNA was labeled in the presence of camptothecin, and a crude mitochondrial fraction was prepared. Mitochondria were suspended in MSB buffer [100 mM NaCl-10 mM MgCl2-10 mM Tris*HCl (pH 7.4)] and lysed with 0.5% sodium deoxycholate-0.5% Brij 58. (Brij 58 was obtained from the Hercules Powder, Co.) The lysed mitochondria were layered on a 15-30% MBS-sucrose gradients and centrifuged ...

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Section: Resultsmentioning

confidence: 99%

“…The ribosome-like structure of mitochondria is relatively small (60 S) (1)(2)(3)(4). Mitochondrial protein synthesis is sensitive to chloramphenicol, an inhibitor of prokaryotic ribosomal function, and insensitive to cycloheximide, an inhibitor of eukaryotic protein synthesis.…”

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confidence: 96%

Mitochondrial Protein Synthesis: RNA with the Properties of Eukaryotic Messenger RNA

Perlman

Abelson

Penman

1973

Proc. Natl. Acad. Sci. U.S.A.

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“…This similarity of dose responses suggests a common molecular basis, which is at present unknown . Ethidium bromide also inhibits mitochondrial protein synthesis (4,5) . Experiments on cellular DNA polymerases in vitro (9) 1 -mitochondrial enzymes, to an alteration of mitochondrial structure, or to some other effect .…”

Section: Discussionmentioning

confidence: 99%

“…1 16 in the cell at the time the drug was administered (3) . In animal cells, EBr has been shown to inhibit mitochondrial protein synthesis (4,5) and mitochondrial RNA synthesis (6,7) .…”

Section: Introductionmentioning

confidence: 99%

THE EFFECT OF ETHIDIUM BROMIDE ON MITOCHONDRIAL DNA SYNTHESIS AND MITOCHONDRIAL DNA STRUCTURE IN HELA CELLS

Leibowitz

1971

The Journal of Cell Biology

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The synthesis of mitochondrial DNA (mDNA) in HeLa cells is selectively inhibited by relatively low concentrations of ethidium bromide . After exposure of cells to strongly inhibitory concentrations of the drug, the apparent superhelix density of mDNA is rapidly increased, as judged by its buoyant density in CsC1 in the presence of ethidium bromide . Mitochondrial DNA synthesized in the presence of partially inhibitory concentrations of ethidium bromide is also altered in its buoyant density in the presence of the dye, but is more heterogeneous in this respect . However, the change in buoyant density of newly synthesized mDNA may be explained by changes in structure other than a change in superhelix density, as indicated by its increased resistance to digestion by pancreatic DNase .

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“…Mild treatment with pancreatic ribonuclease converted the 95 S structure into 55 S particles, whereas treatment with EDTA caused it to sediment at 35 S, without releasing the nascent protein, however. The authors [68] point out that the 95 S structure may represent a 55 S polysome and we may add that the tenacious binding of nascent protein even in the presence of EDTA and the low buoyant density of the 95 S structure could be due to the association of the 55 S ribosome with membrane fragments. Therefore, these results are not necessarily in contradiction with those presented in table 4.…”

Section: Mitochondrial Ribosomes Of Animal Tissuesmentioning

confidence: 99%