HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 283, Issue 25, 17485-17493, June 20, 2008

This Article Free via Author's Choice: AC AC Full Text Full Text (PDF) AC All Versions of this Article: 283/25/17485    most recent M802300200v1 Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Wahid, A. M. Articles by Conn, G. L. Search for Related Content PubMed PubMed Citation Articles by Wahid, A. M. Articles by Conn, G. L. Social Bookmarking                      What's this?

Systematic Deletion of the Adenovirus-associated RNA_I Terminal Stem Reveals a Surprisingly Active RNA Inhibitor of Double-stranded RNA-activated Protein Kinase^*

Ahmed M. Wahid¹, Veronica K. Coventry¹², and Graeme L. Conn³

From the Manchester Interdisciplinary Biocentre, Faculty of Life Sciences, University of Manchester, Manchester M1 7DN, United Kingdom

Adenoviruses use the short noncoding RNA transcript virus-associated (VA) RNA_I to counteract two critical elements of the host cell defense system, innate cellular immunity and RNA interference, mediated by the double-stranded RNA-activated protein kinase (PKR) and Dicer/RNA-induced silencing complex, respectively. We progressively shortened the VA RNA_I terminal stem to examine its necessity for inhibition of PKR. Each deletion, up to 15 bp into the terminal stem, resulted in a cumulative decrease in PKR inhibitory activity. Remarkably, however, despite significant apparent destabilization of the RNA structure, the final RNA mutant that lacked the entire terminal stem (TS21 RNA) efficiently bound PKR and exhibited wild-type inhibitory activity. TS21 RNA stability was strongly influenced by solution pH, indicating the involvement of a protonated base within the VA RNA_I central domain tertiary structure. Gel filtration chromatography and isothermal titration calorimetry analysis indicated that wild-type VA RNA_I and TS21 RNA form similar 1:1 complexes with PKR but that the latter lacks secondary binding site(s) that might be provided by the terminal stem. Although TS21 RNA bound PKR with wild-type K_d, and overall change in free energy (G), the thermodynamics of binding (H and S) were significantly altered. These results demonstrate that the VA RNA_I terminal stem is entirely dispensable for inhibition of PKR. Potentially, VA RNA_I is therefore a truly bi-functional RNA; Dicer processing of the VA RNA_I terminal stem saturates the RNA interference system while generating a "mini-VA RNA_I" molecule that remains fully active against PKR.

Received for publication, March 24, 2008 , and in revised form, April 21, 2008.

^* This work was supported by Research Career Development Fellowship 061444 from The Wellcome Trust (to G. L. C.) and studentship support from the Egyptian Ministry for Higher Education (to A. M. W.) and Medical Research Council, UK (to V. K. C.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Author's Choice—Final version full access.

¹ Both authors contributed equally to this work.

² Present address: Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK.

Author's Choice

Creative Commons Attribution Non-Commercial License applies to Author Choice Articles

³ To whom correspondence should be addressed: Manchester Interdisciplinary Biocentre, 131 Princess St., Manchester M1 7DN, UK. E-mail: Graeme.L.Conn{at}manchester.ac.uk.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				J. A.L. Short Viral evasion of interferon stimulated genes Bioscience Horizons, June 1, 2009; 2(2): 212 - 224. [Abstract] [Full Text] [PDF]