Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Exoribonucleases”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 145 records · Page 8Linked to original sources
PubMed1994

Oat phytochrome A mRNA degradation appears to occur via two distinct pathways.

We have identified possible mechanisms for the degradation of oat phytochrome A (PHYA) mRNA. The majority of PHYA mRNA molecules appeared to be degraded prior to removal of the poly(A) tail, a pathway that differs from that reported for the degradation of other eukaryotic mRNAs. Polyadenylated PHYA mRNA contained a pattern of putative degradation products that is consistent with a 5'-->3' exoribonuclease, although the participation of a stochastic endoribonuclease cannot be excluded. The poly(A) tail of PHYA mRNA was heterogeneous in size and ranged from approximately 14 to 220 nucleotides. Early PHYA mRNA degradation events did not appear to involve site-specific endoribonucleases. Approximately 25% of the apparently full-length PHYA mRNA was poly(A) deficient. Oat H4 histone, beta-tubulin, and actin mRNA populations had lower amounts of apparently full-length mRNAs that were poly(A) deficient. Degradation of the poly(A)-deficient PHYA mRNA, a second pathway, appeared to be initiated by a 3'-->5' exoribonucleolytic removal of the poly(A) tail followed by both 5'-->3' and 3'-->5' exoribonuclease activities. Polysome-associated RNA contained putative PHYA mRNA degradation products and was a mixture of polyadenylated and deadenylated PHYA messages, suggesting that the two distinct degradation pathways are polysome associated.

Base Sequence↗
PubMed2025

Rho-dependent termination and RNase E-mediated cleavage: dual pathways for RNA 3' end processing in polycistronic mRNA.

"Pre-full-length" transcripts are produced at the end of the polycistronic galactose (gal) operon, 5' galE-galT-galK-galM 3', via Rho-dependent transcription termination (RDT) and -independent transcription termination. The 3' end of the full-length galETKM mRNA is acquired by exonucleolytic processing of the 3'-OH ends of the pre-full-length transcripts. However, the gal operon produces an mRNA termed galE whose 3' end forms approximately 120 nucleotides downstream of the galE stop codon, within the subsequent gene, galT, thereby establishing polarity in gene expression. In this study, we investigated the molecular processes that generate the 3' end of galE mRNA. We discovered that the 3' ends of pre-galE mRNA are produced in the middle of galT as a result of the combination of two separate molecular processes-one previously reported as RDT and the other as unreported RNase E-mediated transcript cleavage. The 3' ends of pre-galE mRNA undergo exonucleolytic processing to the 3' end of galE mRNA observed in vivo. A hairpin structure containing an 8 bp stem and a 4-nucleotide loop, located 5-10 nucleotides upstream of the 3' ends of galE mRNA, blocks exoribonuclease digestion and renders transcript stability. These findings demonstrate that RNase E-contrary to its general role in mRNA degradation-produces RNA 3' ends that regulate polarity in gene expression.IMPORTANCEThis study reports the findings of two molecular mechanisms that generate the 3' ends of pre-galE mRNA in the gal operon, viz., Rho-dependent transcription termination and RNase E-mediated cleavage. These 3' ends are subsequently processed to produce stable galE mRNA with a hairpin structure that prevents exoribonuclease degradation. This mechanism establishes gene expression polarity by generating the 3' end of galE mRNA within galT in contrast to the usual mRNA degradation role of RNase E. The study reveals a unique role of RNase E in mRNA processing and stability.

RNA, Messenger↗
PubMed1998

Telomere length regulation and telomeric chromatin require the nonsense-mediated mRNA decay pathway.

Rap1p localization factor 4 (RLF4) is a Saccharomyces cerevisiae gene that was identified in a screen for mutants that affect telomere function and alter the localization of the telomere binding protein Rap1p. In rlf4 mutants, telomeric silencing is reduced and telomere DNA tracts are shorter, indicating that RLF4 is required for both the establishment and/or maintenance of telomeric chromatin and for the control of telomere length. In this paper, we demonstrate that RLF4 is allelic to NMD2/UPF2, a gene required for the nonsense-mediated mRNA decay (NMD) pathway (Y. Cui, K. W. Hagan, S. Zhang, and S. W. Peltz, Mol. Cell. Biol. 9:423-436, 1995, and F. He and A. Jacobson, Genes Dev. 9:437-454, 1995). The NMD pathway, which requires Nmd2p/Rlf4p together with two other proteins, (Upf1p and Upf3p), targets nonsense messages for degradation in the cytoplasm by the exoribonuclease Xrn1p. Deletion of UPF1 and UPF3 caused telomere-associated defects like those caused by rlf4 mutations, implying that the NMD pathway, rather than an NMD-independent function of Nmd2p/Rlf4p, is required for telomere functions. In addition, telomere length regulation required Xrn1p but not Rat1p, a nuclear exoribonuclease with functional similarity to Xrn1p (A. W. Johnson, Mol. Cell. Biol. 17:6122-6130, 1997). In contrast, telomere-associated defects were not observed in pan2, pan3, or pan2 pan3 strains, which are defective in the intrinsic deadenylation-dependent decay of normal (as opposed to nonsense) mRNAs. Thus, loss of the NMD pathway specifically causes defects at telomeres, demonstrating a physiological requirement for the NMD pathway in normal cell functions. We propose a model in which the NMD pathway regulates the levels of specific mRNAs that are important for telomere functions.

Adaptor Proteins, Signal Transducing↗
PubMed2004

5-fluorouracil enhances exosome-dependent accumulation of polyadenylated rRNAs.

The antimetabolite 5-fluorouracil (5FU) is a widely used chemotherapeutic for the treatment of solid tumors. Although 5FU slows DNA synthesis by inhibiting the ability of thymidylate synthetase to produce dTMP, the drug also has significant effects on RNA metabolism. Recent genome-wide assays for 5FU-induced haploinsufficiency in Saccharomyces cerevisiae identified genes encoding components of the RNA processing exosome as potential targets of the drug. In this report, we used DNA microarrays to analyze the effect of 5FU on the yeast transcriptome and found that the drug causes the accumulation of polyadenylated fragments of the 27S rRNA precursor and that defects in the nuclear exoribonuclease Rrp6p enhance this effect. The size distribution of these RNAs and their sensitivity to Rrp6p suggest that they are normally degraded by the nuclear exosome and a 5'-3' exoribonuclease. Consistent with this hypothesis, 5FU inhibits the growth of RRP6 mutants with defects in the degradation function of the enzyme and it interferes with the degradation of an rRNA precursor. The detection of poly(A)(+) pre-RNAs in strains defective in various steps in ribosome biogenesis suggests that the production of poly(A)(+) pre-rRNAs may be a general result of defects in rRNA processing. These findings suggest that 5FU inhibits an exosome-dependent surveillance pathway that degrades polyadenylated precursor rRNAs.

Antimetabolites↗
PubMed2005

Rat1p and Rai1p function with the nuclear exosome in the processing and degradation of rRNA precursors.

Exoribonucleases function in the processing and degradation of a variety of RNAs in all organisms. These enzymes play a particularly important role in the maturation of rRNAs and in a quality-control pathway that degrades rRNA precursors upon inhibition of ribosome biogenesis. Strains with defects in 3'-5' exoribonucleolytic components of the RNA processing exosome accumulate polyadenylated precursor rRNAs that also arise in strains with ribosome biogenesis defects. These findings suggested that polyadenylation might target pre-rRNAs for degradation by the exosome. Here we report experiments that indicate a role for the 5'-3' exoribonuclease Rat1p and its associated protein Rai1p in the degradation of poly(A)(+) pre-rRNAs. Depletion of Rat1p enhances the amount of poly(A)(+) pre-rRNA that accumulates in strains deleted for the exosome subunit Rrp6p and decreases their 5' heterogeneity. Deletion of RAI1 results in the accumulation of poly(A)(+) pre-rRNAs, and inhibits Rat1p-dependent 5'-end processing and Rrp6p-dependent 3'-end processing of 5.8S rRNA. RAT1 and RAI1 mutations cause synergistic growth defects in the presence of rrp6-Delta, consistent with the interdependence of 5'-end and 3'-end processing pathways. These findings suggest that Rai1p may coordinate the 5'-end and 3'-end processing and degradation activities of Rat1p and the nuclear exosome.

Antimetabolites↗
PubMed2000

Degradation of mRNA in bacteria: emergence of ubiquitous features.

The amount of a messenger RNA available for protein synthesis depends on the efficiency of its transcription and stability. The mechanisms of degradation that determine the stability of mRNAs in bacteria have been investigated extensively during the last decade and have begun to be better understood. Several endo- and exoribonucleases involved in the mRNA metabolism have been characterized as well as structural features of mRNA which account for its stability have been determined. The most important recent developments have been the discovery that the degradosome-a multiprotein complex containing an endoribonuclease (RNase E), an exoribonuclease (polynucleotide phosphorylase), and a DEAD box helicase (RhlB)-has a central role in mRNA degradation and that oligo(A) tails synthesized by poly(A) polymerase facilitate the degradation of mRNAs and RNA fragments. Moreover, the phosphorylation status and the base pairing of 5' extremities, together with 3' secondary structures of transcriptional terminators, contribute to the stability of primary transcripts. Degradation of mRNAs can follow several independent pathways. Interestingly, poly(A) tails and multienzyme complexes also control the stability and the degradation of eukaryotic mRNAs. These discoveries have led to the development of refined models of mRNA degradation.

Bacteria↗
PubMed1994

Control of mRNA processing and decay in prokaryotes.

Post-transcriptional mechanisms operate in regulation of gene expression in bacteria, the amount of a given gene product being also dependent on the inactivation rate of its own message. Moreover, segmental differences in mRNA stability of polycistronic transcripts may be responsible for differential expression of genes clustered in operons. Given the absence of 5' to 3' exoribonucleolytic activities in prokaryotes, both endoribonucleases and 3' to 5' exoribonucleases are involved in chemical decay of mRNA. As the 3' to 5' exoribonucleolytic activities are readily blocked by stem-loop structures which are usual at the 3' ends of bacterial messages, the rate of decay is primarily determined by the rate of the first endonucleolytic cleavage within the transcripts, after which the resulting mRNA intermediates are degraded by the 3' to 5' exoribonucleases. Consequently, the stability of a given transcript is determined by the accessibility of suitable target sites to endonucleolytic activities. A considerable number of bacterial messages decay with a net 5' to 3' directionality. Two different alternative models have been proposed to explain such a finding, the first invoking the presence of functional coupling between degradation and the movement of the ribosomes along the transcripts, the second one implying the existence of a 5' to 3' processive '5' binding nuclease'. The different systems by which these two current models of mRNA decay have been tested will be presented with particular emphasis on polycistronic transcripts.

Bacteria↗
PubMed2003

A gene encoding an RNase D exonuclease-like protein is required for post-transcriptional silencing in Arabidopsis.

Post-transcriptional gene silencing (PTGS) and the closely related phenomenon RNA interference (RNAi) result from the initial endonucleolytic cleavage of target mRNAs, which are then presumed to be completely hydrolyzed by exoribonucleases. To date, no plant genes required for PTGS are known to encode exoribonucleases. The Arabidopsis Werner Syndrome-like exonuclease (WEX) gene encodes an RNase D domain most similar to that in human Werner Syndrome protein (WRN), but lacks the RecQ helicase domain. It is also related to Caenorhabditis elegans mut-7, which is essential for RNAi, PTGS, and transposon activity. We isolated a loss-of-function mutant, wex-1, that showed greatly reduced expression of WEX mRNA and early flowering. Although wex-1 did not affect expression of a robust marker for transcriptional gene silencing (TGS), PTGS of a green-fluorescent-protein (GFP) reporter gene was blocked in wex-1 and restored by ectopic expression of WEX, indicating that WEX is required for PTGS but not TGS. Thus, members of the RNase D protein family are required for PTGS in both plants and animals. Interestingly, WEX has been shown to interact with an Arabidopsis RecQ helicase, suggesting that these proteins might comprise a functional equivalent of WRN.

Arabidopsis↗
PubMed1974

Polyriboadenylate sequences at the 3'-termini of ribonucleic acid obtained from mammalian leukemia and sarcoma viruses.

The location of poly(A) sequences in the RNA of mammalian RNA-tumor viruses was determined by enzymatic analyses. The 56-64S viral genomic RNAs, the 20-40S viral subunit RNAs, and the 4-5S poly(A) sequences excised from these viral RNAs were subjected to either hydrolysis with a 3'-OH specific exoribonuclease from Ehrlich ascites tumor cells or phosphorolysis from the 3'-termini with polynucleotide phosphorylase from Micrococcus luteus. Purified adenosine-labeled poly(A) fragments, excised from genomic viral RNAs by RNase A and T(1) digestion, were hydrolyzed with the 3'-OH specific exoribonuclease for various periods of time. Poly(U) filter binding studies of the residual poly(A) indicated that 97% of the poly(A) fragments were hydrolyzed. Adenosine-labeled genomic and subunit viral RNAs and excised poly(A) fragments were phosphorolyzed from their 3'-termini for various periods of time with polynucleotide phosphorylase. The degree of phosphorolysis was monitored by poly(U) filter binding studies, and CCl(3)COOH insolubility and solubility determinations. There was an initial preferential rate of phosphorolysis of the poly(A) sequences of genomic and subunit viral RNAs as compared to the total adenosine-labeled viral RNAs. The data from these two different enzymatic mechanisms of action indicated conclusively that the poly(A) sequences were located at the 3'-termini of genomic and subunit viral RNAs.

Adenine Nucleotides↗
PubMed1993

Substitution of the 3' terminal adenosine residue of transfer RNA in vivo.

We have altered by site-directed mutagenesis the 3' terminal adenosine residue of a tRNA(Tyrsu3+) gene encoded on a single-copy plasmid and examined the consequences of these substitutions on suppressor activity in vivo. Our data show that mutant su3 genes containing 3'-CCC, -CCG, or -CCU termini instead of -CCA can be efficiently transcribed and processed in Escherichia coli to generate functional suppressor tRNAs. However, in contrast to normal tRNA genes, both tRNA nucleotidyltransferase and exoribonuclease activities are required to obtain suppression by the mutant tRNAs, indicating that removal of the incorrect 3' terminal residue and resynthesis of the normal -CCA terminus are occurring in this situation. In addition, a low level of suppressor activity and tRNA repair was found in cells devoid of tRNA nucleotidyltransferase, suggesting that an additional activity able to partially repair the 3' end of tRNA is present in E. coli. The use of mutant strains lacking one or several exoribonucleases revealed that the various RNAses have very different specificities for removal of incorrect 3' residues and that these differ greatly from their action on CCA-ending tRNA. These data show that the 3' terminal adenosine residue is necessary for tRNA function in vivo and that cells can compensate for its alteration by changes in the normal pathway of tRNA metabolism.

Adenosine↗
PubMed1995

Polyadenylylation destabilizes the rpsO mRNA of Escherichia coli.

The rpsO mRNA, encoding ribosomal protein S15, is only partly stabilized when the three ribonucleases implicated in its degradation--RNase E, polynucleotide phosphorylase, and RNase II--are inactivated. In the strain deficient for RNase E and 3'-to-5' exoribonucleases, degradation of this mRNA is correlated with the appearance of posttranscriptionally elongated molecules. We report that these elongated mRNAs harbor poly(A) tails, most of which are fused downstream of the 3'-terminal hairpin at the site where transcription terminates. Poly(A) tails are shorter in strains containing 3'-to-5' exoribonucleases. Inactivation of poly(A) polymerase I (pcnB) prevents polyadenylylation and stabilizes the rpsO mRNA if RNase E is inactive. In contrast polyadenylylation does not significantly modify the stability of rpsO mRNA undergoing RNase E-mediated degradation.

Base Sequence↗
PubMed1998

Polyadenylation of stable RNA precursors in vivo.

Polyadenylation at the 3' terminus has long been considered a specific feature of mRNA and a few other unstable RNA species. Here we show that stable RNAs in Escherichia coli can be polyadenylated as well. RNA molecules with poly(A) tails are the major products that accumulate for essentially all stable RNA precursors when RNA maturation is slowed because of the absence of processing exoribonucleases; poly(A) tails vary from one to seven residues in length. The polyadenylation process depends on the presence of poly(A) polymerase I. A stochastic competition between the exoribonucleases and poly(A) polymerase is proposed to explain the accumulation of polyadenylated RNAs. These data indicate that polyadenylation is not unique to mRNA, and its widespread occurrence suggests that it serves a more general function in RNA metabolism.

Base Sequence↗
PubMed2002

The yeast mitochondrial degradosome. Its composition, interplay between RNA helicase and RNase activities and the role in mitochondrial RNA metabolism.

The yeast mitochondrial degradosome (mtEXO) is an NTP-dependent exoribonuclease involved in mitochondrial RNA metabolism. Previous purifications suggested that it was composed of three subunits. Our results suggest that the degradosome is composed of only two large subunits: an RNase and a RNA helicase encoded by nuclear genes DSS1 and SUV3, respectively, and that it co-purifies with mitochondrial ribosomes. We have found that the purified degradosome has RNA helicase activity that precedes and is essential for exoribonuclease activity of this complex. The degradosome RNase activity is necessary for mitochondrial biogenesis but in vitro the degradosome without RNase activity is still able to unwind RNA. In yeast strains lacking degradosome components there is a strong accumulation of mitochondrial mRNA and rRNA precursors not processed at 3'- and 5'-ends. The observed accumulation of precursors is probably the result of lack of degradation rather than direct inhibition of processing. We suggest that the degradosome is a central part of a mitochondrial RNA surveillance system responsible for degradation of aberrant and unprocessed RNAs.

Amino Acid Sequence↗
PubMed1977

Alterations of activities of ribonucleases and polyadenylate polymerase in synchronized mouse L cells.

The activities of the three known catabolic and the one anabolic polyadenylate enzymes have been determined in synchronized L5178y cells: endoribonuclease, exoribonuclease, 5'-nucleotidase and poly(A) polymerase (Mg2+-dependent). These four enzymes were found primarily in the nuclear fraction. The activity of poly(A) polymerase remains essentially constant during the transition from G1 to S phase. However, the poly(A) catabolic enzyme activities increase parallel with DNA synthesis; the endoribonuclease activity increases 4-fold during G1 to S phase, the exoribonuclease and the nucleotidase activities increasing 30-fold and 16-fold. During the S phase the poly(A)-degrading enzymes are far more active than the poly(A)-synthesizing activity of poly(A) polymerase. We conclude that in L5178y cells the poly(A)-degrading enzymes probably function in regulation of the post-transcriptional net-polyadenylation of heterogeneous nuclear RNA during the phase of DNA synthesis.

Cell Division↗
PubMed1979

Age-dependent enzymatic poly (A) metabolism in quail oviduct.

The poly(A) metabolism in oviducts from adult and senescent quails has been studied. The incorporation studies by double-labelling of mRNA with [3H]adenosine and [3H]uridine revealed, that after inhibition of transcription by actinomycin D, the incorporation ratio adenosine/uridine increases drastically in adult animals compared with the ratio determined for mRNA from senescent animals. This finding is a hint that in senescent animals the poly(A) stretch of mRNA is shorter than in adult animals. This assumption is supported by the finding that the activity of the extractable poly(A) exoribonuclease is higher in senescent than in adult animals. The activity of poly(A) exoribonuclease is even enhanced after stimulation of senescent animals with progesterone. The activity of the other catabolic enzyme studied, the poly (A) endoribonuclease, as well as the activity of anabolic poly(A) enzyme, the poly(A) polymerase, are almost identical in oviducts from adult and senescent animals. The properties of the catabolic poly(A) enzymes and of the anabolic poly(A) polymerase seem to be identical.

Adenosine↗
PubMed2005

Co-evolution of tRNA 3' trailer sequences with 3' processing enzymes in bacteria.

Maturation of the tRNA 3' terminus is a complicated process in bacteria. Usually, it is initiated by an endonucleolytic cleavage carried out by RNase E and Z in different bacteria. In Escherichia coli, RNase E cleaves AU-rich sequences downstream of tRNA, producing processing intermediates with a few extra residues at the 3' end; these are then removed by exoribonuclease trimming to generate the mature 3' end. Here we show that essentially all E. coli tRNA precursors contain a potential RNase E cleavage site, the AU-rich sequence element (AUE), in the 3' trailer. This suggests that RNase E cleavage and exonucleolytic trimming is a general pathway for tRNA maturation in this organism. Remarkably, the AUE immediately downstream of each tRNA is selectively conserved in bacteria having RNase E and tRNA-specific exoribonucleases, suggesting that this pathway for tRNA processing is also commonly used in these bacteria. Two types of RNase E-like proteins are identified in actinobacteria and the alpha-subdivision of proteobacteria. The tRNA 3' proximal AUE is conserved in bacteria with only one type of E-like protein. Selective conservation of the AUE is usually not observed in bacteria without RNase E. These results demonstrate a novel example of co-evolution of RNA sequences with processing activities.

Bacteria↗
PubMed1988

Alteration of nuclear (2'-5')oligoriboadenylate synthetase and nuclease activities preceding replication of human immunodeficiency virus in H9 cells.

After infection of the respective target cells with the human immunodeficiency virus (HIV-1) viral progeny is produced only after a short temporary delay of some days, depending on cell type. After this period of time a sudden onset of HIV-1 protein synthesis with a dramatic increase in virus release occurs. (2'-5')Oligoriboadenylates [(2'-5')A], capable to activate a latent ribonuclease (RNase L) degrading both mRNA and rRNA, are known mediators involved in the early response of cells to virus infection. Here we show that the (2'-5')A-synthesizing (2'-5')A synthetase, which is inducible by interferon and activated by double-stranded RNA, as well as a (2'-5')A nuclease (2',3'-exoribonuclease) are associated with the nuclear matrix of uninfected and infected H9 cells, also in the absence of interferon. Infection of H9 cells with HIV-1 was found to cause a strong (7.7-fold) enhancement of (2'-5')A synthetase activity and a smaller (2-fold) increase of 2',3'-exoribonuclease activity. Simultaneously the concentration of synthesized (2'-5')A increased 5 to 10 times in isolated nuclei. After incubation for 2 to 3 days both enzyme activities reached a maximum and then dropped below their initial values. Concomitantly a drastic increase in virus production occurred, as judged by reverse transcriptase activity in the culture fluid. These results suggest that the (nuclear matrix-associated) (2'-5')A system might be important during the initial stage of HIV infection, also by destructing matrix-bound viral messengers.

2',5'-Oligoadenylate Synthetase↗
PubMed1998

Yeast nuclear PET127 gene can suppress deletions of the SUV3 or DSS1 genes: an indication of a functional interaction between 3' and 5' ends of mitochondrial mRNAs.

Saccharomyces cerevisiae nuclear genes SUV3 and DSS1 encode putative RNA helicase and RNase II, respectively, which are subunits of the mitochondrial degradosome (mtEXO): a three-protein complex which has a 3' to 5' exoribonuclease activity and plays a major role in regulating stability of mitochondrial RNA. Lack of either of the two gene products results in a respiratory negative phenotype, while on the molecular level it causes a total block of mitochondrial translation, loss of the in vitro exoribonuclease activity and changes in stability and processing of many mtRNAs. We have found that the yeast nuclear gene PET127 present on a low or high copy number vector can effectively suppress the effects of the SUV3 or DSS1 gene disruptions. Since the product of the PET127 gene is involved in processing of the 5' ends of mitochondrial mRNAs, we suggest that there is a functional coupling between the 5' and 3' ends of mitochondrial mRNAs.

Blotting, Northern↗