Abstract

The expression of HIV Pol genes is entirely dependent upon a programmed -1 translational frameshift. Frameshifting is mediated by an RNA element that includes a slippery site and a highly conserved, downstream structure. The downstream RNA structure stimulates frameshifting through an unknown mechansim. The objective is to develop an atomic-level understanding of the structure and function of the frameshift site RNA. Dr. Butcher will determine the NMR structures of the frameshift site RNAs from HIV-1 group M, as well as from HIV-2 and SIV-SMM. These structures are broadly recognized to be attractive targets for the development of new therapeutics. Therefore, as a step towards the rational development of second generation RNA binding ligands with enhanced specificity, the structure of the HIV-1 frameshift site RNA bound to a novel small molecule with anti-HIV activity, guanidino neomycin B will be determined. Finally, in order to more broadly understand the structure and function of the frameshift site RNA, photoreactive RNA modifications will be used to probe its interactions with the translational machinery by laser-induced cross-linking. The resulting data will be mapped to the ribosomal and frameshift site RNA structures to produce the first view of the frameshifting interaction.
The specific aims are: 1. Determine the NMR structure of the entire HIV-1 frameshift site (FS) RNA. 2. Investigate the structural and thermodynamic effects of guanidino glycoside binding. 3. Determine the structure of the HIV-2 and SIV-SMM FS RNAs. 4. Identify sites of interaction between the HIV FS RNA and translational machinery.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM072447-04
Application #
7391689
Study Section
AIDS Molecular and Cellular Biology Study Section (AMCB)
Program Officer
Preusch, Peter C
Project Start
2005-04-01
Project End
2010-03-31
Budget Start
2008-04-01
Budget End
2010-03-31
Support Year
4
Fiscal Year
2008
Total Cost
$219,391
Indirect Cost

  Institution

Name
University of Wisconsin Madison
Department
Biochemistry
Type
Schools of Earth Sciences/Natur
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715

  Publications

Butcher, Samuel E; Jan, Eric (2016) tRNA-mimicry in IRES-mediated translation and recoding. RNA Biol 13:1068-1074
Garcia-Miranda, Pablo; Becker, Jordan T; Benner, Bayleigh E et al. (2016) Stability of HIV Frameshift Site RNA Correlates with Frameshift Efficiency and Decreased Virus Infectivity. J Virol 90:6906-6917
Vander Meulen, Kirk A; Horowitz, Scott; Trievel, Raymond C et al. (2016) Measuring the Kinetics of Molecular Association by Isothermal Titration Calorimetry. Methods Enzymol 567:181-213
Hilimire, Thomas A; Bennett, Ryan P; Stewart, Ryan A et al. (2016) N-Methylation as a Strategy for Enhancing the Affinity and Selectivity of RNA-binding Peptides: Application to the HIV-1 Frameshift-Stimulating RNA. ACS Chem Biol 11:88-94
Au, Hilda H; Cornilescu, Gabriel; Mouzakis, Kathryn D et al. (2015) Global shape mimicry of tRNA within a viral internal ribosome entry site mediates translational reading frame selection. Proc Natl Acad Sci U S A 112:E6446-55
Mouzakis, Kathryn D; Dethoff, Elizabeth A; Tonelli, Marco et al. (2015) Dynamic motions of the HIV-1 frameshift site RNA. Biophys J 108:644-54
Yin, Kaifeng; Hacia, Joseph G; Zhong, Zhe et al. (2014) Genome-wide analysis of miRNA and mRNA transcriptomes during amelogenesis. BMC Genomics 15:998
Low, Justin T; Garcia-Miranda, Pablo; Mouzakis, Kathryn D et al. (2014) Structure and dynamics of the HIV-1 frameshift element RNA. Biochemistry 53:4282-91
Moser, Ann B; Hey, Jody; Dranchak, Patricia K et al. (2013) Diverse captive non-human primates with phytanic acid-deficient diets rich in plant products have substantial phytanic acid levels in their red blood cells. Lipids Health Dis 12:10
Mouzakis, Kathryn D; Lang, Andrew L; Vander Meulen, Kirk A et al. (2013) HIV-1 frameshift efficiency is primarily determined by the stability of base pairs positioned at the mRNA entrance channel of the ribosome. Nucleic Acids Res 41:1901-13

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