RESUMO
In order to test the hypothesis that mitochondrial transcription factor A (mtTFA) regulates mitochondrial transcription in vivo, mtTFA was overexpressed in HeLa cells and imported into isolated rat liver mitochondria. Five hours after transfection with an eukaryotic expression vector, mitochondrial transcripts for cytochrome-c-oxidase subunit I and 12 S rRNA were increased over controls. In the presence of rat liver mitochondria, the 29 kDa mtTFA, generated by in vitro translation, was processed to a 24 kDa protein which was protected from protease digestion. This demonstrates that mtTFA was imported into the matrix. Incorporation of 32P-UTP into mitochondrial transcripts was stimulated following import of mTFA. We conclude that the intracellular and intramitochondrial concentration of mtTFA, respectively, indeed regulates mitochondrial transcription.
Assuntos
Mitocôndrias/genética , RNA/genética , Transativadores/genética , Transcrição Gênica , Proteínas de Xenopus , Animais , DNA Mitocondrial/genética , Células HeLa , Humanos , Mitocôndrias Hepáticas/genética , RNA Mitocondrial , Ratos , TransfecçãoRESUMO
The steady state concentration of cytochrome c oxidase subunit I mRNA and 12 S rRNA, respectively, measured by a quantitative reverse transcription/polymerase chain reaction method, was 4 and 15 molecules per molecule of mt DNA in rat liver and 2 and 9 molecules in rat muscle, respectively. These results imply that in the mitochondrial compartment, the molar concentration of all thirteen mRNAs by far exceeds the concentration of ribosomes, a situation fundamentally different from the cytosolic compartment. Following thyroid hormone treatment, both mitochondrial transcripts increased, in parallel with the mRNA encoding mitochondrial transcription factor A. We conclude that this transcription factor might be the rate limiting factor for mitochondrial transcription in vivo, at least under these conditions.
