Deciphering the role of the viral non-structural protein 2A in yellow fever vaccine attenuation

• Applicant: Dr. Melina Winkler
• Subject Area: Virology
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 568562045

Arthropod-borne flaviviruses, such as dengue virus (DENV), West Nile virus (WNV) and yellow fever virus (YFV), cause significant human health problems and are an economic burden worldwide. While no vaccine or treatments currently exist to prevent or treat many flavivirus infections, the YFV live-attenuated vaccine strain, YFV-17D, is one of the most effective vaccines ever developed. Although YFV-17D and its virulent parental genome, YFV-Asibi, only differ by less than 1% of nucleotides, the specific viral and/or host factors responsible for YFV virulence vs attenuation have not yet been identified. Using a highly modular approach to generate viral chimeras allowed members of the Ploss lab including myself to map the mutations responsible for viral spread in vitro to the E protein and the mutations responsible for a faster and stronger innate immune response to the non-structural protein 2A (NS2A). We have validated our data in vivo using interferon alpha/beta receptor knock-out and human liver chimeric mice and determined that another virulent YFV strain, Dakar, is attenuated by 17D’s mutations in the E and NS2A regions. Concordantly, 17D exhibits enhanced pathogenicity when the mutations in E and NS2A are reverted to the sequence of its parental pathogenic strain Asibi. In this proposal, I will build on these findings and unravel the mechanisms underlying the phenotypic differences conferred by the mutations in NS2A. I will fine map the mutations in NS2A responsible for immunogenicity. I will further address the role of the NS2A protein in immunogenicity by systematically stimulating individual innate immune pathways and quantifying activation of downstream factors. Finally, I will employ innovative mass spectrometry approaches to determine qualitative and quantitative changes in NS2A’s interactome. Altogether, these data will enhance our mechanistic understanding of YFV virulence vs attenuation. Results stemming from my work hold promise as a blueprint for rational development of live-attenuated vaccines for other virulent flaviviruses.

• DFG Programme: WBP Fellowship
• International Connection: USA
• Host: Professor Alexander Ploss, Ph.D.