Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To determine the role of poly(A) polymerase in 3'-end processing of mRNA, the effect of purified poly(A) polymerase antibodies on endonucleolytic cleavage and polyadenylation was studied in HeLa nuclear extracts, using adenovirus L3 pre-mRNA as the substrate. Both Mg2+- and Mn2+-dependent reactions catalyzing addition of 200 to 250 and 400 to 800 adenylic acid residues, respectively, were inhibited by the antibodies, which suggested that the two reactions were catalyzed by the same enzyme. Anti-poly(A) polymerase antibodies also inhibited the cleavage reaction when the reaction was coupled or chemically uncoupled with polyadenylation. These antibodies also prevented formation of specific complexes between the RNA substrate and components of nuclear extracts during cleavage or polyadenylation, with the concurrent appearance of another, antibody-specific complex. These studies demonstrate that (i) previously characterized poly(A) polymerase is the enzyme responsible for addition of the poly(A) tract at the correct cleavage site and probably for the elongation of poly(A) chains and (ii) the coupling of these two 3'-end processing reactions appears to result from the potential requirement of poly(A) polymerase for the cleavage reaction. The results suggest that the specific endonuclease is associated with poly(A) polymerase in a functional complex.
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PMID:Role of poly(A) polymerase in the cleavage and polyadenylation of mRNA precursor. 256 10

Thermal stress induces expression of a family of heat shock proteins which may regulate the synthesis of various cellular genes. We investigated the effect of heat shock on polyadenylation in Epstein-Barr Virus (EBV) negative and EBV transformed human Burkitt's lymphoma (BL) B-cell lines. Incubation of the BL B-cell line P3HR-1, carrying the defective EBV genome [EBV nuclear antigen-2 gene deletion] at 46 degrees C for 15 min increased nuclear poly(A) polymerase (PAP) activity. Thereafter, enzymatic activity declined and at 60 min it was reduced to about 50% of that observed in cells incubated at 37 degrees C. In contrast, no significant increase in PAP activity was observed at 15 min or thereafter in an EBV- BL cell line, ST-486, in response to elevated temperature. Furthermore, no heat shock mediated change in nuclear poly(A)-specific endonuclease activity was observed in either P3HR-1 or ST-486 cells suggesting a specific effect on PAP activity. However, thermal stress dependent increase in c-myc expression was detected only in P3HR-I cells. These results suggest an association between EBV transformation and enhanced expression of c-myc and PAP activity. To further determine the role of EBV, and EBV- BL cell line, BL-30, and BL-30 cells infected in vitro with a wild type strain of EBV, BL-30/B95-8, were investigated. BL-30/B-95-8, unlike the parental BL-30 cells, exhibited c-myc and PAP gene upregulation at 15 min but were downregulated at 60 min following exposure of cells to elevated temperatures. These results suggest that infection of human B-cells with EBV is associated with their ability to respond to thermal stress by increased PAP activity which may stabilize mRNA through enhanced polyadenylation.
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PMID:Differential effect of heat shock on RNA metabolism in human Burkitt's lymphoma B-cell lines. 855

Ribosomal RNAs are generally synthesized as long, primary transcripts that must be extensively processed to generate the mature, functional species. In Escherichia coli, it is known that the initial 30S precursor is cleaved during its synthesis by the endonuclease RNase III to generate precursors to the 16S, 23S, and 5S rRNAs. However, despite extensive study, the processes by which these intermediate products are converted to their mature forms are poorly understood. In this article, we describe the maturation of 23S rRNA. Based on Northern analysis of RNA isolated from a variety of mutant strains lacking one or multiple ribonucleases, we show that maturation of the 3' terminus requires the action of RNase T, an enzyme previously implicated in the end turnover of tRNA and in the maturation of small, stable RNAs. Although other exoribonucleases can participate in shortening the 3' end of the initial RNase III cleavage product, RNase T is required for removal of the last few residues. In the absence of RNase T, 23S rRNA products with extra 3' residues accumulate and are incorporated into ribosomes, with only small effects on cell growth. Purified RNase T accurately and efficiently converts these immature ribosomes to their mature forms in vitro, whereas free RNA is processed relatively poorly. In vivo, the processing defect at the 3' end has no effect on 5' maturation, indicating that the latter process proceeds independently. We also find that a portion of the 23S rRNA that accumulates in many RNase T- cells becomes polyadenylated because of the action of poly(A) polymerase I. The requirement for RNase T in 23S rRNA maturation is discussed in relation to a model in which only this enzyme, among the eight exoribonucleases present in E. coli, is able to efficiently remove nucleotides close to the double-stranded stem generated by the pairing of the 5' and 3' termini of most stable RNAs.
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PMID:Maturation of 23S ribosomal RNA requires the exoribonuclease RNase T. 991 73

RNAI is a short RNA, 108 nt in length, which regulates the replication of the plasmid ColE1. RNAI turns over rapidly, enabling plasmid replication rate to respond quickly to changes in plasmid copy number. Because RNAI is produced in abundance, is easily extracted and turns over quickly, it has been used as a model for mRNA in studying RNA decay pathways. The enzymes polynucleotide phosphorylase, poly(A) polymerase and RNase E have been demonstrated to have roles in both messenger and RNAI decay; it is reported here that these enzymes can work independently of one another to facilitate RNAI decay. The roles in RNAI decay of two further enzymes which facilitate mRNA decay, the exonuclease RNase II and the endonuclease RNase III, are also examined. RNase II does not appear to accelerate RNAI decay but it is found that, in the absence of RNase III, polyadenylated RNAI, unprocessed by RNase E, accumulates. It is also shown that RNase III can cut RNAI near nt 82 or 98 in vitro. An RNAI fragment corresponding to the longer of these can be found in extracts of an mc+ pcnB strain (which produces RNase III) but not of an rnc pcnB strain, suggesting that RNAI may be a substrate for RNase III in vivo. A possible pathway for the early steps in RNAI decay which incorporates this information is suggested.
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PMID:Absence of RNASE III alters the pathway by which RNAI, the antisense inhibitor of ColE1 replication, decays. 1058 16

The stability of mRNA in prokaryotes depends on multiple factors and it has not yet been possible to describe the process of mRNA degradation in terms of a unique pathway. However, important advances have been made in the past 10 years with the characterization of the cis-acting RNA elements and the trans-acting cellular proteins that control mRNA decay. The trans-acting proteins are mainly four nucleases, two endo- (RNase E and RNase III) and two exonucleases (PNPase and RNase II), and poly(A) polymerase. RNase E and PNPase are found in a multienzyme complex called the degradosome. In addition to the host nucleases, phage T4 encodes a specific endonuclease called RegB. The cis-acting elements that protect mRNA from degradation are stable stem-loops at the 5' end of the transcript and terminators or REP sequences at their 3' end. The rate-limiting step in mRNA decay is usually an initial endonucleolytic cleavage that often occurs at the 5' extremity. This initial step is followed by directional 3' to 5' degradation by the two exonucleases. Several examples, reviewed here, indicate that mRNA degradation is an important step at which gene expression can be controlled. This regulation can be either global, as in the case of growth rate-dependent control, or specific, in response to changes in the environmental conditions.
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PMID:Messenger RNA stability and its role in control of gene expression in bacteria and phages. 1069 Apr 8

Sequences at the immediate 3' terminus of several eukaryotic primary transcripts, synthesised just before the termination of transcription, are often lost during RNA processing. The rna82.1 mutation in Saccharomyces cerevisiae appears to result in a deficiency of the endonuclease that removes such sequences from certain yeast transcripts. Some small RNAs of rna82.1 cells are a few nucleotides longer than their counterparts in wild-type S. cerevisiae. The 5S rRNAs made during very short pulse-labellings of the mutant have, relative to the mature 121 nucleotide 5S RNA of wild-type cells, an additional 7, 11 or 13 nucleotides at their 3' terminus. These 5S forms reveal sites upon 5S genes where transcription probably terminates in vivo. The extra nucleotides upon 5S RNAs in rna82.1 cells are lost very slowly by sequential removal from the 3' terminus. Through this 3'-5' exonuclease action the total 5S RNA of the mutant possesses several 3'-terminal sequences yet is mostly only 0-3 nucleotides longer than in wild-type S. cerevisiae. Just one or two of these 3'-terminal sequences serve as a substrate in vivo for a poly(A) polymerase since a small proportion of rna82.1 5S RNAs terminate in the sequence: CAAUCUUU(A)n.
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PMID:Altered maturation of sequences at the 3' terminus of 5S gene transcripts in a Saccharomyces cerevisiae mutant that lacks a RNA processing endonuclease. 1189 49

The exosome is a multi-subunit 3'-5' exonucleolytic complex that is conserved in structure and function in all eukaryotes studied to date. The complex is present in both the nucleus and cytoplasm, where it continuously works to ensure adequate quantities and quality of RNAs by facilitating normal RNA processing and turnover, as well as by participating in more complex RNA quality-control mechanisms. Recent progress in the field has convincingly shown that the nucleolytic activity of the exosome is maintained by only two exonuclease co-factors, one of which is also an endonuclease. The additional association of the exosome with RNA-helicase and poly(A) polymerase activities results in a flexible molecular machine that is capable of dealing with the multitude of cellular RNA substrates that are found in eukaryotic cells. Interestingly, the same basic set of enzymatic activities is found in prokaryotic cells, which might therefore illustrate the evolutionary origin of the eukaryotic system. In this Commentary, we compare the structural and functional characteristics of the eukaryotic and prokaryotic RNA-degradation systems, with an emphasis on some of the functional networks in which the RNA exosome participates in eukaryotes.
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PMID:Origins and activities of the eukaryotic exosome. 1942 Feb 35