MI2025 – Donald Ting

Title:
N153-linked glycans on the envelope protein of orthoflaviviruses antagonize antibody-mediated host defenses

Abstract:
Dengue virus (DENV) poses a significant disease burden globally with an estimated 390 million infections annually. The envelope (E) protein of DENV is glycosylated at two highly conserved asparagine (N) sites (N67 and N153). While the functions of these glycans have been extensively studied in vitro, their biological roles have largely been overlooked in vivo.

A DENV2 mutant lacking N153-linked glycans (N153Q mutant) was engineered and found to be mildly impaired in vitro but drastically attenuated in a symptomatic mouse model of severe dengue, as evidenced by accelerated viral clearance. B cell depletion and knockout (KO) in mouse model restored N153Q parental virulence, suggesting the involvement of B cells in N153Q mutant attenuation. Homologous passive transfer of purified IgM from infected B-cell proficient mice into B cell-KO mice cleared N153Q mutant from blood circulation. In vitro neutralization assay using mouse sera showed that WT and N153Q viruses were not significantly neutralized. These data suggest that N153Q mutant attenuation was due to non-neutralizing IgM-mediated viral clearance mechanism. Interestingly, heterologous passive transfer of purified IgM from WT-infected mice into N153Q-infected B cell-KO mice did not clear the mutant, suggesting that infection with WT DENV and N153Q mutant induced distinct antibody repertoires.

Furthermore, using plasma samples from convalescent dengue patients and monoclonal antibodies, in vitro neutralization assays showed that N153Q DENV1-4 and N154Q ZIKV mutants were more susceptible than WT to IgG-mediated neutralization. Glycoproteomics combined with molecular dynamics (MD) simulations revealed that the presence of the glycan and its composition on E protein heavily influenced IgG binding.

These findings represent a novel immune evasion strategy for orthoflaviviruses and have important implications for the development of antibodies, live attenuated vaccine and antiviral.

Biography:
Donald carried out his PhD studies under the guidance of A/P Sylvie Alonso from the Infectious Diseases Translational Research Programme (IDTRP), and Department of Microbiology & Immunology, Yong Loo Lin School of Medicine, National University of Singapore (NUS). He completed his undergraduate studies at NUS and graduated with a Bachelor of Science (Hons) in Life Sciences. His interest in virology arose from his fascination with these simple yet complex infectious agents.