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Journal of Virology, August 2002, p. 8058-8068, Vol. 76, No. 16
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.16.8058-8068.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Secondary Structure of the 3' Terminus of Hepatitis C Virus Minus-Strand RNA

Catherine Schuster,1* Catherine Isel,2 Isabelle Imbert,1 Chantal Ehresmann,2 Roland Marquet,2 and Marie Paule Kieny1

U544 INSERM, Institut de Virologie, 67000 Strasbourg,1 CNRS UPR 9002, Institut de Biologie Moléculaire et Cellulaire du CNRS, 67084 Strasbourg Cedex, France2

Received 22 January 2002/ Accepted 10 May 2002

The 3'-terminal ends of both the positive and negative strands of the hepatitis C virus (HCV) RNA, the latter being the replicative intermediate, are most likely the initiation sites for replication by the viral RNA-dependent RNA polymerase, NS5B. The structural features of the very conserved 3' plus [(+)] strand untranslated region [3' (+) UTR] are well established (K. J. Blight and C. M. Rice, J. Virol. 71:7345-7352, 1997). However, little information is available concerning the 3' end of the minus [(-)] strand RNA. In the present work, we used chemical and enzymatic probing to investigate the conformation of that region, which is complementary to the 5' (+) UTR and the first 74 nucleotides of the HCV polyprotein coding sequence. By combining our experimental data with computer predictions, we have derived a secondary-structure model of this region. In our model, the last 220 nucleotides, where initiation of the (+) strand RNA synthesis presumably takes place, fold into five stable stem-loops, forming domain I. Domain I is linked to an overall less stable structure, named domain II, containing the sequences complementary to the pseudoknot of the internal ribosomal entry site in the 5' (+) UTR. Our results show that, even though the (-) strand 3'-terminal region has the antisense sequence of the 5' (+) UTR, it does not fold into its mirror image. Interestingly, comparison of the replication initiation sites on both strands reveals common structural features that may play key functions in the replication process.

* Corresponding author. Mailing address: U544 INSERM, Institut de Virologie, 3 rue Koeberlé, 67000 Strasbourg, France. Phone: 33 (0)3.90.24.37.41. Fax: 33 (0)3.90.24.37.23. E-mail: Catherine.schuster{at}viro-ulp.u-strasbg.fr.

Journal of Virology, August 2002, p. 8058-8068, Vol. 76, No. 16
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.16.8058-8068.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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