1887

Microbiology Society logo

Volume 144, Issue 7

Degradation pathway of CopA, the antisense RNA that controls replication of plasmid R1 Free

Abstract

RNA decay in bacteria is carried out by a number of enzymes that participate in the coordinated degradation of their substrates. Endo- and exonucleolytic cleavages as well as polyadenylation are generally involved in determining the half-life of RNAs. Small, untranslated antisense RNAs are suitable model systems to study decay. A study of the pathway of degradation of CopA, the copy number regulator RNA of plasmid R1, is reported here. Strains carrying mutations in the genes encoding RNase E, polynucleotide phosphorylase (PNPase), RNase II and poly(A) polymerase I (PcnB/PAP I) -- alone or in combination -- were used to investigate degradation patterns and relative half-lives of CopA. The results obtained suggest that RNase E initiates CopA decay. Both PNPase and RNase II can degrade the major 3-cleavage product generated by RNase E. This exonucleolytic degradation is aided by PcnB, which may imply a requirement for A-tailing. RNase II can partially protect CopA's 3′-end from PNPase-dependent degradation. Other RNases are probably involved in decay, since in rnblpnp double mutants, decay still occurs, albeit at a reduced rate. Experiments using purified RNase E identified cleavage sites in CopA in the vicinity of, but not identical to, those mapped in vivo, suggesting that the cleavage site specificity of this RNase is modulated by additional proteins in the cell. A model of CopA decay is presented and discussed.

  • Received:
  • Accepted:
  • Revised:
  • Published Online:
Keyword(s): antisense RNA , exoribonucleases , poly(A) polymerase , RNA decay and RNase E
© Society for General Microbiology 1998
Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-144-7-1907
1998-07-01
2024-04-25
Loading full text...

Full text loading...

/deliver/fulltext/micro/144/7/mic-144-7-1907.html?itemId=/content/journal/micro/10.1099/00221287-144-7-1907&mimeType=html&fmt=ahah

References

  1. Arraiano C.M., Yancey S.D., Kushner S.R. 1988; Stabilization of discrete mRNA breakdown products in ams pnp rnb multiple mutants of Escherichia coli K-12. . J Bacteriol 170:4625–4633
     [Google Scholar]
  2. Bertani G. 1951; Studies on lysogenesis. I. The mode of phage liberation by lysogenic Escherichia coli. . J Bacteriol 62:293–300
     [Google Scholar]
  3. Blomberg P., Wagner E.G.H., Nordström K. 1990; Control of replication of plasmid R1: the duplex between the antisense RNA, CopA, and its target, CopT, is processed specifically in vivo and in vitro by RNase III. . EMBO J 9:2331–2340
     [Google Scholar]
  4. Brantl S., Wagner E.G.H. 1996; An unusually long-lived antisense RNA in plasmid copy number control: in vivo RNAs encoded by the streptococcal plasmid pIP501. . J Mol Biol 255:275–288
     [Google Scholar]
  5. Braun F., Hajnsdorf E., Regniér P. 1996; Polynucleotide phosphorylase is required for the rapid degradation of the RNase E-processed rpsO mRNA of Escherichia coli devoid of its 3ʹ- hairpin. . Mol Microbiol 19:997–1005
     [Google Scholar]
  6. Cao G.J., Sarkar N. 1992; Identification of the gene for an Escherichia coli poly (A) polymerase. . Proc Natl Acad Sci USA 8910380–10384
     [Google Scholar]
  7. Carpousis A.J., Houve G.V., Ehretsmann C., Krisch H.M. 1994; Copurification of E. coli RNAse E and PNPase: evidence for a specific association between two enzymes important in RNA processing and degradation. . Cell 76:889–900
     [Google Scholar]
  8. Coburn G.A., Mackie G.A. 1996a; Differential sensitivities of portions of the mRNA for ribosomal protein S20 to 3ʹ- exonucleases dependent on oligoadenylation and RNA secondary structure. . J Biol Chem 271:15776–15781
     [Google Scholar]
  9. Coburn G.A., Mackie G.A. 1996b; Overexpression, purification, and properties of Escherichia coli ribonuclease II. . J Biol Chem 271:1048–1053
     [Google Scholar]
  10. Cohen S.N., McDowall K.J. 1997; RNase E: still a wonderfully mysterious enzyme. . Mol Microbiol 23:1099–1106
     [Google Scholar]
  11. Deutscher M.P. 1993; Ribonuclease multiplicity, diversity and complexity. . J Biol Chem 268:13011–13014
     [Google Scholar]
  12. Donovan W.P., Kushner S.R. 1986; Polynucleotide phosphorylase and ribonuclease II are required for cell viability and mRNA turnover in Escherichia coli K-12. . Proc Natl Acad Sci USA 83120–124
     [Google Scholar]
  13. Eguchi Y., Itoh T., Tomizawa J. 1991; Antisense RNA. . Annu Rev Biochem 60:631–652
     [Google Scholar]
  14. Georgellis D., Sohlberg B., Hartl F.-U., von Gabain A. 1995; Identification of GroEL as a constituent of an mRNA protection complex in Escherichia coli. . Mol Microbiol 16:1259–1268
     [Google Scholar]
  15. Guarneros G., Portier C. 1991; Different specificities of ribonuclease I1 and polynucleotide phosphorylase in 3σ mRNA decay.. Biochimie 73543–549
     [Google Scholar]
  16. Hajnsdorf E., Braun F., Haugel-Nielsen J., Régnier P. 1994; Polyadenylation destabilizes the rpsO mRNA of Escherichia coli. . Proc Natl Acad Sci USA 923973–3977
     [Google Scholar]
  17. He L, Söderborn F., Wagner E.G.H., Binnie U., Binns N., Masters M. 1993; PcnB is required for the rapid degradation of RNA I, the antisense RNA that controls the copy number of ColEl-related plasmids. . Mol Microbiol 9:1131–1142
     [Google Scholar]
  18. Hjalt T.Å.H., Wagner E.G.H. 1995; Bulged-out nucleotides protect an antisense RNA from RNase III cleavage. . Nucleic Acids Res 23:571–579
     [Google Scholar]
  19. Ingle C.A., Kushner S.R. 1996; Development of an in vitro mRNA decay system for Escherichia coli : poly(A)polymerase I is necessary to trigger degradation. . Proc Natl Acad Sci USA 9312926–12931
     [Google Scholar]
  20. Jensen R.B., Grohmann E., Schwab H., Diaz-Orejas R., Gerdes K. 1995; Comparison of ccd of F, parDE of RP4, and parD of R1 using a novel conditional replication control system of plasmid Rl. . Mol Microbiol 17:211–220
     [Google Scholar]
  21. Kaberdin V.R., Chao Y.H., Lin-Chao S. 1996; RNase E cleaves at multiple sites in bubble regions of RNA I stem loops yielding products that dissociate differentially from the enzyme. . J Biol Chem 271:13103–13109
     [Google Scholar]
  22. Lin-Chao S., Cohen S.N. 1991; The rate of processing and degradation of antisense RNA I regulates the replication of ColEl-type plasmids in vivo. . Cell 65:1233–1242
     [Google Scholar]
  23. Lopilato J., Bortner S., Beckwith J. 1986; Mutations in a new chromosomal gene of Escherichia coli K-12, pcnB, reduce plasmid copy number of pBR322 and its derivatives. . Mol Gen Genet 205:285–290
     [Google Scholar]
  24. McDowall K.J., Lin-Chao S., Cohen S.N. 1994; A + U content rather than a particular nucleotide order determines the specificity of RNase E cleavage. . J Biol Chem 269:10790–10796
     [Google Scholar]
  25. McLaren R.S., Newbury S.F., Dance G.S., Causton H.C., Higgins C.F. 1991; mRNA degradation by processive 3ʹ-5ʹ exoribonucleases in vitro and the implications for prokaryotic mRNA decay in vivo. . J Mol Biol 221:81–95
     [Google Scholar]
  26. Masters M., Colioms M.D., Oliver I.R., He L., Macnaughton E.J., Charters Y. 1993; The pcnB gene of Escherichia coli, which is required for ColEl copy number maintenance, is dispensable. . J Bacteriol 175:4405–4413
     [Google Scholar]
  27. Miczak A., Kaberdin V.R., Wei C.L., Lin-Chao S. 1996; Proteins associated with RNase E in a multicomponent ribo- nucleolytic complex. . Proc Natl Acad Sci USA 933865–3869
     [Google Scholar]
  28. Mikkelsen N.D., Gerdes K. 1997; Sok antisense RNA from plasmid Rl is functionally inactivated by RNase E and poly- adenylated by poly(A) polymerase I. . Mol Microbiol 26:311–320
     [Google Scholar]
  29. Naureckiene S., Uhlin B.E. 1996; In vitro analysis of mRNA processing by RNase E in the pap operon of Escherichia coli. . Mol Microbiol 21:55–68
     [Google Scholar]
  30. Nierlich D.P., Murakawa G.J. 1996; The decay of bacterial messenger RNA. . Prog Nucleic Acid Res Mol Biol 52:153–216
     [Google Scholar]
  31. Nordström K., Wagner E.G.H. 1994; Kinetic aspects of control of plasmid replication by antisense RNA. . Trends Biochem Sei 19:294–300
     [Google Scholar]
  32. Pepe C.M., Maslesa-Galic S., Simons R.W. 1994; Decay of the 7S10 antisense RNA by 3ʹ exoribonucleases: evidence that RNase II stabilizes RNA-OUT against PNPase attack. . Mol Microbiol 13:1133–1142
     [Google Scholar]
  33. Py B., Causton H., Mudd E.A., Higgins C.F. 1994; A protein complex mediating mRNA degradation in Escherichia coli. . Mol Microbiol 14:717–729
     [Google Scholar]
  34. Py B., Higgins C.F., Krisch H.M., Carpousis A.J. 1996; A DEAD-box RNA helicase in the Escherichia coli RNA degrado-some. . Nature 381:169–172
     [Google Scholar]
  35. Söderbom F., Binnie U., Masters M., Wagner E.G.H. 1997; Regulation of plasmid R1 replication: PcnB and RNase E expedite the decay of the antisense RNA, CopA. . Mol Microbiol 26:493–504
     [Google Scholar]
  36. Sohlberg B., Lundberg U., Hartl F.-U., von Gabain A. 1993; Functional interaction of heatshock protein GroEL-with an RNase E-like activity in Escherichia coli. . Proc Natl Acad Sei USA 90277–281
     [Google Scholar]
  37. Wagner E.G.H., Nordström K. 1986; Structural analysis of an RNA molecule involved in replication of plasmid Rl. . Nucleic Acids Res 14:2523–2538
     [Google Scholar]
  38. Wagner E.G.H., Simons R.W. 1994; Antisense RNA control in bacteria, phage and plasmids. . Annu Rev Microbiol 48:713–742
     [Google Scholar]
  39. Xu F., Cohen S. N. 1995; RNA degradation in Escherichia coli regulated by 3σ adenylation and 5σ phosphorylation.. Nature 374180–183
     [Google Scholar]
  40. Xu F., Lin-Chao S., Cohen S.N. 1993; The Escherichia coli pcnB gene promotes adnylation of antisense RNA I of ColEl- type plasmids in vivo and degradation of RNA I decay intermediates. . Proc Natl Acad Sei USA 906756–6760
     [Google Scholar]
  41. Zilhão R., Cairrao F., Régnier P., Arraiano C.M. 1996; PNPase modulates RNase II expression in Escherichia coli: implications for mRNA decay and cell metabolism. . Mol Microbiol 20:1033–1042
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-144-7-1907
Loading
Degradation pathway of CopA, the antisense RNA that controls replication of plasmid R1
Microbiology 144, 1907 (1998); https://doi.org/10.1099/00221287-144-7-1907
/content/journal/micro/10.1099/00221287-144-7-1907
/content/journal/micro/10.1099/00221287-144-7-1907
Loading

Data & Media loading...

Most cited Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error