Figure 5.

Inhibition of direct presentation prevents CD8⁺ T-cell priming to cytosolic transgene devoid of functional MHC class II epitope. (a) Secreted sOVA construct comprises the full-length OVA cDNA while cytoplasmic cOVA construct is deleted from 126 amino acids at the Cter OVA cDNA. For both constructs, additional constructs bearing four repeats of the miR142.3p target sequence in the 3′ untranslated region were generated. (b–d) C57Bl/6 female mice were injected i.m. at day 0 with rAAV1-cOVA, rAAV1-cOVA-miR, rAAV1-sOVA, or rAAV1-sOVA-miR. (b) At day 14 or 35, mice were sacrificed and their splenocytes tested by IFN-γ ELISPOT against the OVA₂₅₇ peptide. Data represent six mice per group per time point pooled from two independent experiments. ND, not detected. At (c) day 14 or (d) day 35, tibialis anterior muscles from mice injected with rAAV1-cOVA or rAAV1-cOVA-miR were frozen and 7 µm thick transversal cryosections were stained with hematoxilin and eosin. Field size is 720 × 720 micron and inserts size is 300 × 300 micron. One representative muscle out of three is shown. (e) Rag2^−/− or C57Bl/6 female mice were injected i.m. at day 0 with rAAV1-cOVA or rAAV1-cOVAmiR. On day 35, tibialis anterior muscles from sacrificed mice were removed and OVA mRNA was quantified by Q-RT-PCR. Data represent four mice per group. One of two different experiments is shown.