Figure 2.

Lentiviral-Vector-Transduced DCs Raise a Protective Response to LCMV Infection

(A) On the left is shown the structure of lentiviral vectors expressing GFP, mCD40L, and mCD40L fused to GP_33–41 separated by a picornavirus P2A self-cleaving motif. On the right, the biosynthesis of the CD40L:GP_33–41 fusion protein in the ER is diagrammed. Because of the type II topology of CD40L, the peptide is released into the endoplasmic reticulum upon synthesis to assemble with MHC class I proteins. (B) SAMHD1 knockout BMDCs were transduced with lentiviral vectors and 1 × 10⁶ were injected into C57BL/6 mice (n = 5). After 5 days, CD8⁺ splenic T cells were stained with GP_33–41 MHC class I tetramer and analyzed by flow cytometry. (C) Mice were injected with 1 × 10⁶-transduced BMDCs and after 7 days, challenged with LCMV Armstrong (n = 5). After 4 days, the virus load in the spleen was measured by plaque assay. The dotted line represents the lower level of detection. (D) Mice were injected with a serial 5-fold dilution of BMDCs transduced with the mCD40L-GP33 vector or control mCD40L lentiviral vector and then challenged with LCMV Armstrong (n = 4). After 4 days, virus loads were measured by plaque assay. The dotted line represents the lower level of detection. (E) Mice were injected with BMDCs transduced with the lentiviral vector expressing mCD40L or mCD40L and GP_33–41. 7 days after immunization, the mice were challenged by intracerebral injection of LCMV Armstrong (n = 5) and monitored for 1 year. After 300 days, the mice were rechallenged with LCMV Armstrong. Data represent mean ± SD. Significance determined by Mann-Whitney U tests.