Polyadenylation Is Dispensable for Encapsidation and Reverse Transcription of Hepatitis B viral Pregenomic RNA

  • Lee, Hye-Jin (Department of Biochemistry, Yonsei University) ;
  • Lee, Jehan (Department of Biochemistry, Yonsei University) ;
  • Shin, Myeong-Kyun (Department of Biochemistry, Yonsei University) ;
  • Ryu, Wang-Shick (Department of Biochemistry, Yonsei University)
  • Received : 2007.10.30
  • Accepted : 2007.12.11
  • Published : 2008.06.30

Abstract

A hepadnaviruses replicates its DNA genome via reverse transcription of an RNA template (pregenomic RNA or pgRNA), which has a cap structure at the 5' end and a poly(A) tail at the 3' end. We have previously shown that the 5' cap is indispensable for encapsidation of the pgRNA. A speculative extension of the above finding is that the cap contributes to encapsidation via its interaction with the poly(A) tail, possibly involving eIF4E-eIF4G-PABP interaction. To test this hypothesis, poly(A)-less pgRNAs were generated via cleavage by a cis-acting hepatitis delta virus ribozyme sequence. We found that accumulation of the poly(A)-less pgRNA was markedly diminished, mostly likely due to its reduced stability. Importantly, however, the remaining poly(A)-less pgRNAs were nonetheless encapsidated and reverse transcribed normally when the reduced stability was taken account. Our finding clearly contradicts the notion that the poly(A) tail has any function in encapsidation and viral reverse transcription.

Keywords

Acknowledgement

Supported by : Korean Research Foundation

References

  1. Abraham, T.M., and Loeb, D.D. (2006). Base pairing between the 5′ half of epsilon and a cis-acting sequence, phi, makes a contribution to the synthesis of minus-strand DNA for human hepatitis B virus. J. Virol. 80, 4380-4387 https://doi.org/10.1128/JVI.80.9.4380-4387.2006
  2. Allen, M.I., Deslauriers, M., Andrews, C.W., Tipples, G.A., Walters, K.A., Tyrrell, D.L., Brown, N., and Condreay, L.D. (1998). Identification and characterization of mutations in hepatitis B virus resistant to lamivudine. Lamivudine Clinical Investigation Group. Hepatology 27, 1670-1677 https://doi.org/10.1002/hep.510270628
  3. Bartenschlager, R., and Schaller, H. (1992). Hepadnaviral assembly is initiated by polymerase binding to the encapsidation signal in the viral RNA genome. EMBO. J. 11, 3413-3420
  4. Galibert, F., Mandart, E., Fitoussi, F., Tiollais, P., and Charnay, P. (1979). Nucleotide sequence of the hepatitis B virus genome (subtype ayw) cloned in E. coli. Nature 281, 646-650 https://doi.org/10.1038/281646a0
  5. Ganem, D., and Schneider, R. (2001). Hepadnaviridae: the viruses and their replication. In Fields Virology, P.M.H. D. M. Knipe, ed. (Philadelphia, USA: Lippincott-Raven Publishers), pp. 2923-2970
  6. Gebauer, F., and Hentze, M.W. (2004). Molecular mechanisms of translational control. Nat. Rev. Mol. Cell Biol. 5, 827-835 https://doi.org/10.1038/nrm1488
  7. Havert, M.B., and Loeb, D.D. (1997). cis-Acting sequences in addition to donor and acceptor sites are required for template switching during synthesis of plus-strand DNA for duck hepatitis B virus. J. Virol. 71, 5336-5344
  8. Jeong, J.K., Yoon, G.S., and Ryu, W.S. (2000). Evidence that the 5′-end cap structure is essential for encapsidation of hepatitis B virus pregenomic RNA. J. Virol. 74, 5502-5508 https://doi.org/10.1128/JVI.74.12.5502-5508.2000
  9. Junker-Niepmann, M., Bartenschlager, R., and Schaller, H. (1990). A short cis-acting sequence is required for hepatitis B virus pregenome encapsidation and sufficient for packaging of foreign RNA. EMBO J. 9, 3389-3396
  10. Lee, J., Lee, H.-J., Shin, M.-K., and Ryu, W.-S. (2004). Versatile PCR-mediated insertion or deletion mutagenesis. BioTechniques 36, 398-400
  11. Loeb, D.D., and Tian, R. (1995). Transfer of the minus strand of DNA during hepadnavirus replication is not invariable but prefers a specific location. J. Virol. 69, 6886-6891
  12. Munroe, D., and Jacobson, A. (1990). mRNA poly(A) tail, a 3′ enhancer of translational initiation. Mol. Cell. Biol. 10, 3441-3455 https://doi.org/10.1128/MCB.10.7.3441
  13. Nassal, M., and Rieger, A. (1996). A bulged region of the hepatitis B virus RNA encapsidation signal contains the replication origin for discontinuous first-strand DNA synthesis. J. Virol. 70, 2764-2773
  14. Oropeza, C.E., and McLachlan, A. (2007). Complementarity between epsilon and phi sequences in pregenomic RNA influences hepatitis B virus replication efficiency. Virology 359, 371-381 https://doi.org/10.1016/j.virol.2006.08.036
  15. Ostrow, K.M., and Loeb, D.D. (2004). Underrepresentation of the 3′ region of the capsid pregenomic RNA of duck hepatitis B virus. J. Virol. 78, 2179-2186 https://doi.org/10.1128/JVI.78.5.2179-2186.2004
  16. Pattnaik, A.K., Ball, L.A., LeGrone, A.W., and Wertz, G.W. (1992). Infectious defective interfering particles of VSV from transcripts of a cDNA clone. Cell 69, 1011-1020 https://doi.org/10.1016/0092-8674(92)90619-N
  17. Perrotta, A.T., and Been, M.D. (1996). Core sequences and a cleavage site wobble pair required for HDV antigenomic ribozyme self-cleavage. Nucleic Acids Res. 24, 1314-1321 https://doi.org/10.1093/nar/24.7.1314
  18. Pollack, J.R., and Ganem, D. (1993). An RNA stem-loop structure directs hepatitis B virus genomic RNA encapsidation. J. Virol. 67, 3254-3263
  19. Pollack, J.R., and Ganem, D. (1994). Site-specific RNA binding by a hepatitis B virus reverse transcriptase initiates two distinct reactions: RNA packaging and DNA synthesis. J. Virol. 68, 5579-5587
  20. Rieger, A., and Nassal, M. (1996). Specific hepatitis B virus minus-strand DNA synthesis requires only the 5′ encapsidation signal and the 3′-proximal direct repeat DR1. J. Virol. 70, 585-589
  21. Sharmeen, L., Kuo, M.Y., Dinter-Gottlieb, G., and Taylor, J. (1988). Antigenomic RNA of human hepatitis delta virus can undergo self-cleavage. J. Virol. 62, 2674-2679
  22. Shin, M.-K., Lee, J., and Ryu, W.-S. (2004). A novel cis-acting element facilitates minus-strand DNA synthesis during reverse transcription of the hepatitis B viruses genome. J. Virol. 78, 6252-6262 https://doi.org/10.1128/JVI.78.12.6252-6262.2004
  23. Shin, M.K., Kim, J.H., Ryu, D.K., and Ryu, W.S. (2008). Circularization of an RNA template via long-range base pairing is critical for hepadnaviral reverse transcription. Virology 371, 362-373 https://doi.org/10.1016/j.virol.2007.09.042
  24. Tang, H., and McLachlan, A. (2002). A pregenomic RNA sequence adjacent to DR1 and complementary to epsilon influences hepatitis B virus replication efficiency. Virology 303, 199-210 https://doi.org/10.1006/viro.2002.1645
  25. Tavis, J.E., Perri, S., and Ganem, D. (1994). Hepadnavirus reverse transcription initiates within the stem-loop of the RNA packaging signal and employs a novel strand transfer. J. Virol. 68, 3536-3543
  26. Wang, G.H., and Seeger, C. (1993). Novel mechanism for reverse transcription in hepatitis B viruses. J. Virol. 67, 6507-6512