TLR9 activation is required for cytotoxic response elicited by baculovirus capsid display

MARÍA INÉS CRESPO; PAULA MOLINARI; GUIDO N. MOLINA; BELKYS A. MALETTO; OSCAR TABOGA; IGNACIO CEBRIAN; G MORON

Mar del Plata

Congreso; LXX Reunión de la Sociedad Argentina de Inmunología. 3ra Congreso Franco-Argentino de Inmunología; 2022

Sociedad Argentina de Inmunología

The baculovirus (BV) infects lepidopteran as natural hosts and representsan efficient vector for vaccine development. We showed thatBV expressing OVA in its capsid (BVOVA) improved the immuneresponse by eliciting CD8+ T cell activation. BVs enter intact intosubcellular compartments of dendritic cells (DC) where both viralsurface proteins and capsid follow separate intracellular routes, allowingthe viral capsid containing the foreign Ag to reach the cytosol.In this study, we analyzed the intracellular mechanisms involvedin CD8+ T cell activation by BVOVA. We first evaluated if the BVGP64-fusogenic protein is required for OVA cross-presentation bypretreating BVOVA with an anti-GP64 neutralizing antibody. In thiscase, infected splenic DCs failed to activate the B3Z CD8+ T cellhibridoma, specific for the MHCI(Kb)-OVA257–264. Moreover, T cell activationwas inhibited when DCs were treated with lactacystin, indicatingthat proteasome is required for efficiently cross-presentation.We then analyzed the dependence of TLR9 and STING signaling byinfecting DCs with BVOVA in the presence of H151, a STING inhibitor,or IRS869, a TLR9 signaling inhibitor. Only the incubation withIRS869 blocked B3Z activation by DCs. We also evaluated whetherTLR9 is required to induce proliferation of naïve CD8+ T cellsusing BVOVA-infected DCs from WT and TLR9-/- mice. We found that TLR9-/- DCs cannot activate naïve CD8+ T cells efficiently. Also,TLR9-/- mice fail to generate an OVA-specific cytotoxic responseafter BVOVA injection. Finally, using IFNAR-/- mice we observedtype I IFN are not required for cytotoxic response induction. In thiswork, we found that the viral envelope fuses with the endosomalmembrane allowing the viral capsid exits to cytosol for proteasomaldegradation. During this BVOVA intracellular trafficking, endosomalTLR9 signaling in DCs is triggered, thereby stimulating the presentationof the MHCI/OVA257-264 complex to CD8+ T cells, which activateinto CTL.