Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.8 (polynucleotide phosphorylase)
723 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Polyadenylation plays an important role in RNA degradation in bacterial cells. In Escherichia coli, exoribonucleases, mostly RNase II and polynucleotide phosphorylase, antagonize the synthesis of poly(A) tails by poly(A) polymerase I (PAP I). In accordance with earlier observations showing that only a small fraction of bacterial RNA is polyadenylated, we demonstrate here that approximately 10% of rpsO mRNA harbors short oligo(A) tails ranging from one to five A residues in wild-type cells. We also compared the length, frequency and distribution of poly(A) tails at the 3'-end of rpsO transcripts in vivo in the presence and absence of Hfq, a host factor that in vitro stimulates the activity of PAP I, and found that Hfq affects all three parameters. In the hfq(+) strain the average length of oligo(A) tails and frequency of polyadenylated transcripts was higher than in the hfq(-) strain and a smaller proportion of tails was found at the 3' end of transcripts terminated at the Rho- independent terminator. Our data led us to the conclusion that Hfq is involved in the recognition of 3' RNA extremities by PAP I.
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PMID:Hfq affects the length and the frequency of short oligo(A) tails at the 3' end of Escherichia coli rpsO mRNAs. 1285 18

In Escherichia coli, REP-stabilizers are structural elements in polycistronic messages that protect 5'-proximal cistrons from 3'-->5' exonucleolytic degradation. The stabilization of a protected cistron can be an important determinant in the level of gene expression. Our results suggest that RNase E, an endoribonuclease, initiates the degradation of REP-stabilized mRNA. However, subsequent degradation of mRNA fragments containing a REP-stabilizer poses a special challenge to the mRNA degradation machinery. Two enzymes, the DEAD-box RNA helicase, RhlB and poly(A) polymerase (PAP) are required to facilitate the degradation of REP-stabilizers by polynucleotide phosphorylase (PNPase). This is the first in vivo evidence that these enzymes are required for the degradation of REP-stabilizers. Furthermore, our results show that REP degradation by RhlB and PNPase requires their association with RNase E as components of the RNA degradosome, thus providing the first in vivo evidence that this ribonucleolytic multienzyme complex is involved in the degradation of structured mRNA fragments.
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PMID:The RNA degradosome and poly(A) polymerase of Escherichia coli are required in vivo for the degradation of small mRNA decay intermediates containing REP-stabilizers. 1473 Dec 78

In Escherichia coli, the post-transcriptional addition of poly(A) tails by poly(A) polymerase I (PAP I, pcnB) plays a significant role in cellular RNA metabolism. However, many important features of this system, including its regulation and the selection of polyadenylation sites, are still poorly understood. Here we show that the inactivation of Hfq (hfq), an abundant RNA-binding protein, leads to the reduction in the ability of PAP I to add poly(A) tails at the 3' termini of mRNAs containing Rho-independent transcription terminators even though PAP I protein levels remain unchanged. Those poly(A) tails that are synthesized in the absence of Hfq are shorter in length, even in the absence of polynucleotide phosphorylase (PNPase), RNase II and RNase E. In fact, the biosynthetic activity of PNPase in the hfq single mutant is enhanced and it becomes the primary polynucleotide polymerase, adding heteropolymeric tails almost exclusively to 3' truncated mRNAs. Surprisingly, both PNPase and Hfq co-purified with His-tagged PAP I under native conditions indicating a potential complex among these proteins. Immunoprecipitation experiments using PNPase- and Hfq-specific antibodies confirmed the protein-protein interactions among PAP I, PNPase and Hfq. Analysis of mRNA half-lives in hfq, deltapcnB and hfq deltapcnB mutants suggests that Hfq and PAP I function in the same mRNA decay pathway.
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PMID:The Sm-like protein Hfq regulates polyadenylation dependent mRNA decay in Escherichia coli. 1552 76

Mammalian mitochondrial (mt) mRNAs have short poly(A) tails at their 3' termini that are post-transcriptionally synthesized by mt poly(A) polymerase (PAP). The polyadenylation of mt mRNAs is known to be a key process needed to create UAA stop codons that are not encoded in mtDNA. In some cases, polyadenylation is required for the tRNA maturation by editing of its 3' terminus. However, little is known about the functional roles the poly(A) tail of mt mRNAs plays in mt translation and RNA turnover. Here we show human mt PAP (hmtPAP) and human polynucleotide phosphorylase (hPNPase) control poly(A) synthesis in human mitochondria. Partial inactivation of hmtPAP by RNA interference using small interfering RNA in HeLa cells resulted in shortened poly(A) tails and decreased steady state levels of some mt mRNAs as well as their translational products. Moreover, knocking down hmtPAP generated markedly defective mt membrane potentials and reduced oxygen consumption. In contrast, knocking down hPNPase showed significantly extended poly(A) tails of mt mRNAs. These results demonstrate that the poly(A) length of human mt mRNAs is controlled by polyadenylation by hmtPAP and deadenylation by hPNPase, and polyadenylation is required for the stability of mt mRNAs.
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PMID:Human mitochondrial mRNAs are stabilized with polyadenylation regulated by mitochondria-specific poly(A) polymerase and polynucleotide phosphorylase. 1576 37

Polyadenylation plays a role in decay of some bacterial mRNAs, as well as in the quality control of stable RNA. In Escherichia coli, poly(A) polymerase I (PAP I) is the main polyadenylating enzyme, but the addition of 3' tails also occurs in the absence of PAP I via the synthetic activity of polynucleotide phosphorylase (PNPase). The nature of 3'-tail addition in Bacillus subtilis, which lacks an identifiable PAP I homologue, was studied. Sizing of poly(A) sequences revealed a similar pattern in wild-type and PNPase-deficient strains. Sequencing of 152 cloned cDNAs, representing 3'-end sequences of nontranslated and translated RNAs, revealed modified ends mostly on incomplete transcripts, which are likely to be decay intermediates. The 3'-end additions consisted of either short poly(A) sequences or longer heteropolymeric ends with a mean size of about 40 nucleotides. Interestingly, multiple independent clones exhibited complex heteropolymeric ends of very similar but not identical nucleotide sequences. Similar polyadenylated and heteropolymeric ends were observed at 3' ends of RNA isolated from wild-type and pnpA mutant strains. These data demonstrated that, unlike the case of some other bacterial species and chloroplasts, PNPase of Bacillus subtilis is not the major enzyme responsible for the addition of nucleotides to RNA 3' ends.
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PMID:Addition of poly(A) and heteropolymeric 3' ends in Bacillus subtilis wild-type and polynucleotide phosphorylase-deficient strains. 1599 84

Polyadenylation of RNAs by poly(A) polymerase I (PAP I) in Escherichia coli plays a significant role in mRNA decay and general RNA quality control. However, many important features of this system, including the prevalence of polyadenylated mRNAs in the bacterium, are still poorly understood. By comparing the transcriptomes of wild-type and pcnB deletion strains using macroarray analysis, we demonstrate that >90% of E.coli open reading frames (ORFs) transcribed during exponential growth undergo some degree of polyadenylation by PAP I, either as full-length transcripts or decay intermediates. Detailed analysis of over 240 transcripts suggests that Rho-independent transcription terminators serve as polyadenylation signals. Conversely, mRNAs terminated in a Rho-dependent fashion are probably not substrates for PAP I, but can be modified by the addition of long polynucleotide tails through the biosynthetic activity of polynucleotide phosphorylase (PNPase). Furthermore, real-time PCR analysis indicates that the extent of polyadenylation of individual full-length transcripts such as lpp and ompA varies significantly in wild-type cells. The data presented here demonstrates that polyadenylation in E.coli occurs much more frequently than previously envisioned.
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PMID:The majority of Escherichia coli mRNAs undergo post-transcriptional modification in exponentially growing cells. 1704 Aug 98

Polyadenylation in animal mitochondria is very unique. Unlike other systems, polyadenylation is needed to generate UAA stop codons that are not encoded in mitochondrial (mt) DNA. In some cases, polyadenylation is required for the mt tRNA maturation by editing of its 3' termini. Furthermore, recent studies on human mt poly(A) polymerase (PAP) and PNPase provide new insights and questions for the regulatory mechanism and functional role of polyadenylation in human mitochondria.
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PMID:Polyadenylation in mammalian mitochondria: insights from recent studies. 1831 63

Replication of the ColE2 plasmid requires a plasmid-coded initiator protein (Rep). Rep expression is controlled by antisense RNA (RNAI), which prevents the Rep mRNA translation. In this paper, we examined the effects of RNA degradation enzymes on the degradation pathways of RNAI of the ColE2 plasmid. In the DeltapcnB strain lacking the poly(A) polymerase I (PAP I) the RNAI degradation intermediate (RNAI(*)) accumulates much more than that in the wt strain. RNAI(*) is produced by the RNase E cleavage. RNase II and PNPase are involved in further degradation of RNAI(*) and PAP I is necessary for efficient degradation. The degradation process of ColE2 RNAI is similar to those of R1 CopA RNA and ColE1 RNAI, although the nucleotide sequences and fine secondary structures of these three RNAs are different. ColE2 RNAI is cleaved at multiple positions in the 5' end region by RNase E. The degradation pathway of ColE2 RNAI shown here is quite different from that of the ColE2 Rep mRNA which we have previously reported. In the DeltapcnB strain used for RNA analysis the copy number of the ColE2 plasmid decreases to about a half as compared with that in the isogenic wt strain.
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PMID:The effects of RNA degradation enzymes on antisense RNAI controlling ColE2 plasmid copy number. 1868 57

The polyadenylation-stimulated RNA degradation pathway takes place in plant and algal organelles, yet the identities of the enzymes that catalyze the addition of the tails remain to be clarified. In a search for the enzymes responsible for adding poly(A) tails in Chlamydomonas and Arabidopsis organelles, reverse genetic and biochemical approaches were employed. The involvement of candidate enzymes including members of the nucleotidyltransferase (Ntr) family and polynucleotide phosphorylase (PNPase) was examined. For several of the analyzed nuclear-encoded proteins, mitochondrial localization was established and possible dual targeting to mitochondria and chloroplasts could be predicted. We found that certain members of the Ntr family, when expressed in bacteria, displayed poly(A) polymerase (PAP) activity and partially complemented an Escherichia coli strain lacking the endogenous PAP1 enzyme. Other Ntr proteins appeared to be specific for tRNA maturation. When the expression of PNPase was down-regulated by RNAi in Chlamydomonas, very few poly(A) tails were detected in chloroplasts for the atpB transcript, suggesting that this enzyme may be solely responsible for chloroplast polyadenylation activity in this species. Depletion of PNPase did not affect the number or sequence of mitochondrial mRNA poly(A) tails, where unexpectedly we found, in addition to polyadenylation, poly(U)-rich tails. Together, our results identify several Ntr-PAPs and PNPase in organelle polyadenylation, and reveal novel poly(U)-rich sequences in Chlamydomonas mitochondria.
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PMID:Polyadenylation in Arabidopsis and Chlamydomonas organelles: the input of nucleotidyltransferases, poly(A) polymerases and polynucleotide phosphorylase. 1930 54

In this work, we describe RapA-dependent polyadenylation of model RNA substrates and endogenous, RNA polymerase-associated nucleic acid fragments. We demonstrate that the Escherichia coli RNA polymerase obtained through the classic purification procedure carries endogenous RNA oligonucleotides, which, in the presence of ATP, are polyriboadenylated in a RapA-dependent manner by an accessory poly(rA) polymerase. RNA polymerase isolated from poly(A) polymerase- (PAP-) and polynucleotide phosphorylase- (PNP-) deficient E. coli strain lacks accessory (rA)(n)-synthetic activity. Experiments with reconstituted RNA polymerase-PAP and RNA polymerase-PNP mixtures suggest that RapA enables the polyadenylation by PAP of RNA polymerase-associated RNA. Mutations disrupting RapA's ATP-hydrolytic function disrupt RapA-dependent polyadenylation, and the rapA(-)E. coli strain displays a measurable reduction in RNA polyadenylation. RapA-dependent polyadenylation can also be modulated by mutations in the section of RapA's SWI/SNF domain linked to interaction with single-stranded nucleic acid. We have developed enzymatic assays in which model, synthetic RNAs are polyriboadenylated in a RapA-dependent manner. Taken together, our results are consistent with RapA acting as an RNA polymerase-associated, ATP-dependent RNA translocase. Our work further links RapA to RNA remodeling and provides new mechanistic insights into the functional interaction between RNA polymerase and RapA.
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PMID:RapA, Escherichia coli RNA polymerase SWI/SNF subunit-dependent polyadenylation of RNA. 2129 17


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