Browsing by Subject "POLYNUCLEOTIDE PHOSPHORYLASE"

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  • Maes, Alexandre; Gracia, Celine; Innocenti, Nicolas; Zhang, Kaiyang; Aurell, Erik; Hajnsdorf, Eliane (2017)
    Polyadenylation is thought to be involved in the degradation and quality control of bacterial RNAs but relatively few examples have been investigated. We used a combination of 5 '-tagRACE and RNA-seq to analyze the total RNA content from a wild-type strain and from a poly(A) polymerase deleted Mutant. A total of 178 transcripts were either up- or down-regulated in the mutant when compared to the wild-type strain. Poly(A) polymerase up-regulates the expression of all genes related to the FliA regulon and several previously unknown transcripts, including numerous transporters. Notable down-regulation of genes in the expression of antigen 43 and components of the type 1 fimbriae was detected. The major consequence of the absence of poly(A) polymerase was the accumulation of numerous sRNAs, antisense transcripts, REP sequences and RNA fragments resulting from the processing of entire transcripts. A new algorithm to analyze the position and composition of post-transcriptional modifications based on the sequence of unencoded 3 '-ends, was developed to identify polyadenylated molecules. Overall our results shed new light on the broad spectrum of action of polyadenylation on gene expression and demonstrate the importance of poly(A) dependent degradation to remove structured RNA fragments.
  • Toompuu, Marina; Tuomela, Tea; Laine, Pia; Paulin, Lars; Dufour, Eric; Jacobs, Howard T. (2018)
    RNA 3' polyadenylation is known to serve diverse purposes in biology, in particular, regulating mRNA stability and translation. Here we determined that, upon exposure to high levels of the intercalating agent ethidium bromide (EtBr), greater than those required to suppress mitochondrial transcription, mitochondrial tRNAs in human cells became polyadenylated. Relaxation of the inducing stress led to rapid turnover of the polyadenylated tRNAs. The extent, kinetics and duration of tRNA polyadenylation were EtBr dose-dependent, with mitochondrial tRNAs differentially sensitive to the stress. RNA interference and inhibitor studies indicated that ongoing mitochondrial ATP synthesis, plus the mitochondrial poly(A) polymerase and SUV3 helicase were required for tRNA polyadenylation, while polynucleotide phosphorylase counteracted the process and was needed, along with SUV3, for degradation of the polyadenylated tRNAs. Doxycycline treatment inhibited both tRNA polyadenylation and turnover, suggesting a possible involvement of the mitoribosome, although other translational inhibitors had only minor effects. The dysfunctional tRNA(Leu(UUR)) bearing the pathological A3243G mutation was constitutively polyadenylated at a low level, but this was markedly enhanced after doxycycline treatment. We propose that polyadenylation of structurally and functionally abnormal mitochondrial tRNAs entrains their PNPase/SUV3-mediated destruction, and that this pathway could play an important role in mitochondrial diseases associated with tRNA mutations.