RNA virus pathogenicity is regulated in significant part by complex higher-order RNA structures across their genomes. Structured RNA elements clearly play important roles in viral replication, protein synthesis, packaging, evasion of host immune factors, and the hijacking of host-cell machinery, but the full extent to which viral RNA genomes contain true higher-order structures whose functions are critical to viral processes is poorly understood. Dengue virus (DENV) is a serious mosquito-borne pathogen that threatens more than one- third of the world?s population, and there are currently no broadly effective vaccines or therapeutics. The goals for this research and training proposal are to discover the extent and conservation of higher-order RNA genome structure across all four major DENV serotypes and to characterize the functional importance of conserved RNA structural elements. Training will emphasize the development of new expertise in novel chemical RNA structure probing methods, next-generation sequencing, bioinformatics analyses, and cell culture-based functional assays. RNA secondary and tertiary structure across the entire genomes of all four major DENV serotypes will be interrogated at single-nucleotide resolution using state-of-the-art chemical probing methods read out by massively parallel sequencing. The trainee will devise new computational analysis strategies to compare diverse DENV serotype structural models to identify conserved RNA structural elements. The functional importance of these elements will be assessed in cell culture-based virus functional assays evaluating DENV replication and infectivity. Novel RNA regulatory motifs discovered in DENV may be exploited in the development of critically needed and broadly effective anti-dengue virus therapeutics and vaccination strategies. The proposed multidisciplinary training in innovative RNA structure probing, next generation sequencing, bioinformatics, and modern virological methods will thoroughly prepare the trainee to be a dynamic leader in the field of RNA viruses and biotechnology.
Dengue virus, a serious mosquito-borne pathogen that threatens more than one-third of the world?s population, uses higher-order structure in its RNA genome to regulate critical steps in its viral life cycle. This research and training project involves the discovery and characterization of functional higher-order RNA structural elements conserved across the RNA genomes of multiple dengue virus serotypes. Novel RNA regulatory motifs discovered in dengue virus may be exploited in the development of critically needed and broadly effective anti- dengue virus therapeutics and vaccination strategies.
|Dethoff, Elizabeth A; Boerneke, Mark A; Gokhale, Nandan S et al. (2018) Pervasive tertiary structure in the dengue virus RNA genome. Proc Natl Acad Sci U S A 115:11513-11518|