Complement-Mediated Neutralization of Dengue Virus Requires Mannose-Binding Lectin

Avirutnan, Panisadee; Hauhart, Richard E.; Marovich, Mary A.; Garred, Peter; Atkinson, John P.; Diamond, Michael S.

A previous study showed that serum from wild-type C57BL/6 mice neutralized DENV-2 generated in both C6/36 insect cells and BHK21-15 mammalian cells (25) . In that study, 80% of the neutralizing activity was lost when serum from MBL-A Ϫ/Ϫ ϫ MBL-C Ϫ/Ϫ (MBL-A/C Ϫ/Ϫ ) mice was used, suggesting that the lectin pathway contributed to complement-mediated neutralization of DENV serotype 2 (DENV-2) (25) . However, that study did not address whether neutralization of DENV-2 required activation of other complement activation pathways or downstream components. To evaluate this, we pretreated insect cell (C6/36)-derived and mammalian cell (Vero)-derived DENV-2 with naïve sera from the wild type and several congenic mouse strains deficient in different complement components. Analogous to results with insect cell-derived WNV (25) , neutralization of both insect cell-and mammalian cell-derived DENV-2 was dependent on MBL and MBL-associated serine protease 2 (MASP-2), but not C1q or C5 ( Fig. 1A and C) . Thus, the classical complement activation pathway and the formation of membrane attack complex were not necessary for neutralization of DENV-2 by serum. Because an absence of both C3 and C4 (C3 Ϫ/Ϫ ϫ C4 Ϫ/Ϫ double knockout [DKO]) almost abrogated the inhibitory effect of serum, MBL-dependent neutralization of DENV-2 required activation of the complement system; however, deficiencies in either C3 or C4 resulted in only partial loss of neutralization phenotypes. This suggests that DENV-2 neutralization by MBL occurs through both the canonical lectin (C4-dependent) pathway and the C4 and C2 bypass pathways (11) (12) (13) of complement activation (Fig. 1E ).