Poster abstracts
Poster number 21 submitted by Jonathan Kitzrow
Role of HIV-1 5′UTR RNA structure and conformational dynamics in genome packaging
Jonathan P. Kitzrow (Department of Chemistry and Biochemistry, Center for RNA Biology and Center for Retroviral Research, The Ohio State University, Columbus, OH 43210), Shuohui Lui (Department of Chemistry and Biochemistry, Center for RNA Biology and Center for Retroviral Research, The Ohio State University, Columbus, OH 43210), Shiqin Miao (Department of Chemistry and Biochemistry, Center for RNA Biology and Center for Retroviral Research, The Ohio State University, Columbus, OH 43210), Kevin Jamison (Department of Physics, The Ohio State University, Columbus, OH 43210), Dennis Bong (Department of Chemistry and Biochemistry, Center for RNA Biology and Center for Retroviral Research, The Ohio State University), Michael Poirier (Department of Physics, The Ohio State University, Columbus, OH 43210)
Abstract:
The highly conserved 5′UTR of HIV-1 genomic RNA (gRNA) is central to the regulation of virus replication. Biochemical and NMR experiments support a model in which the 5′UTR can adopt at least two mutually exclusive conformational states. In one state, the genome remains a monomer, as the palindromic dimerization initiation site (DIS) is sequestered via base pairing to upstream sequences. In the second state, the DIS is exposed and the genome is competent for dimerization and packaging into assembling virions. According to this model the conformation of the 5′UTR determines the fate of the genome. The number of 5′ guanosines has also been implicated in the localization of HIV-1 gRNA; transcripts with three 5′ guanosines (3G) are abundant in the cytoplasm, whereas transcripts with a single 5′ guanosine (1G) are preferentially packaged into budding virions.[1, 2] We previously characterized a 238-nt region of the 5′UTR lacking the 5′-TAR/polyA domain using both ensemble and single molecule FRET assays.[3] This study confirmed that the 5′UTR was conformationally dynamic and revealed how the dynamics were modulated by host factor and viral protein binding. The impact of the number of 5′ guanosines on the ensemble of RNA conformations and their dynamics is unknown. Here, we investigate the full 5′UTR using a bifacial peptide nucleic acid strategy to position internal FRET dyes. Native in-gel FRET and RNA structure-probing studies revealed that 1G and 3G 5′UTRs adopt a different ensemble of RNA conformations. Single molecule studies to probe differences in RNA dynamics are underway. These results have implications for the development of RNA-targeted therapeutics that may interfere with selective packaging of HIV-1 gRNA.
References:
1. Masuda, T., et al., Fate of HIV-1 cDNA intermediates during reverse transcription is dictated by transcription initiation site of virus genomic RNA. Sci Rep, 2015. 5: p. 17680.
2. Kharytonchyk, S., et al., Transcriptional start site heterogeneity modulates the structure and function of the HIV-1 genome. Proc Natl Acad Sci U S A, 2016. 113(47): p. 13378-13383.
3. Brigham, B.S., et al., Intrinsic conformational dynamics of the HIV-1 genomic RNA 5'UTR. Proc Natl Acad Sci U S A, 2019. 116(21): p. 10372-10381.
Keywords: RNA Structure, FRET, HIV