Fig. 2: E2 mutations that disrupt E2 HS-binding sites also interfere with EEEV-protein receptor interactions.
A Infectivity of K562 cells expressing empty vector (EV), VLDLR, ApoER2 isoform1, or ApoER2 isoform2 (n = 9 from 3 independent experiments, except for EV where n = 12 from 4 independent experiments). B Infectivity of THP-1 cells expressing EV or LDLR (n = 9 from 3 independent experiments). C Binding of chimeric WT and mutant viruses to K562-EV or K562-VLDLR cells (n = 12 from 4 independent experiments for WT, n = 9 from 3 independent experiments for 84–119 and 156–157, and n = 6 from 2 independent experiments for 71–77). Viruses were incubated with cells on ice for 75 min, then cells were washed, and the bound virus was quantified by qPCR, then expressed as a ratio to GAPDH and normalized to EV. D Apparent binding affinity showing the nonlinear fit of WT or mutant SINV/EEEV for VLDLR LA(1-2)-Fc, with KD apparent and Hill slope (h) (n = 6 from 2 independent experiments). E Neutralization of chimeric eGFP reporter WT and mutant viruses by VLDLR LA(1-2)-Fc (100–0.1 µg/mL in tenfold dilutions and 0 µg control) in Vero cells (n = 4 from 2 independent experiments, except for controls where n = 8 for WT and n = 12 for mutants from 2 independent experiments). F Neutralization of chimeric eGFP reporter WT and mutant viruses by VLDLR LA(1-2)-Fc in CHO-K1 and CHO-pgsA-756 cells (n = 4 from 2 independent experiments, except for WT where n = 9 from 3 independent experiments). G Neutralization of chimeric eGFP reporter WT by heparin or BSA control (2000–2 µg/mL in tenfold dilutions and control) in cells overexpressing protein receptors (n = 6 from 2 independent experiments, except for N2a ΔB4galt7-LDLR cells where controls are n = 6 and samples are n = 4 from 2 independent experiments) quantified by flow cytometry. Means ± SD are shown. Significance was determined by two-way ANOVA with Tukey’s post-hoc tests. Exact p-values are indicated. Source data provided as a Source Data file.
