A NUCLEOSIDE TRIPHOSPHATE-REGULATED, 3' EXONUCLEOLYTIC MECHANISM IS INVOLVED IN TURNOVER OF YEAST MITOCHONDRIAL RNAS

We have employed cell-free transcription reactions with mitochondria i solated from Saccharomyces cerevisiae to study the mechanism of RNA tu rnover. The specificity of RNA turnover was preserved in these prepara tions, as were other RNA-processing reactions, including splicing, 3' end formation of mRNAs, and maturation of rRNAs. Turnover of nascent R NAs was found to occur exonucleolytically; endonucleolytic cleavage pr oducts were not detected during turnover of the omega intron RNA, whic h was studied in detail. However, these experiments still leave open t he possibility that endonucleolytic cleavage products with very short half-lives are kinetic intermediates in the decay of omega RNA. Exonuc leolytic turnover was regulated by nucleotide triphosphates and requir ed their hydrolysis. A unique signature of this regulation was that an y one of the eight standard ribo- or deoxyribonucleotide triphosphates supported RNA turnover. A novel hybrid selection protocol was used to determine the turnover rates of the 5', middle, and 3' portions of on e mitochondrial transcript, the omega intron RNA. The results suggeste d that degradation along that transcript occurred with a 3'-->5' polar ity. The similarity between features of mitochondrial RNA turnover and the properties of a nucleotide triphosphate-dependent 3' exoribonucle ase that has been purified from yeast mitochondria suggests that this single enzyme is a key activity whose regulation is involved in the sp ecificity of mitochondrial RNA turnover.