In our previous study by Gupta et al, dominant T-cell epitopes of SARS CoV-N(N) protein were predicted by software. The spectrum of interferon (IFN)-γ responses of Balb/c mice immunized against two different forms of SARS CoV-N plasmid was then analyzed. A cluster of dominant T-cell epitopes of SARS CoV-N protein was found in the N-terminus (amino acids 76–114). On the basis of this study, four different plasmids were constructed: (i) DNA encoding the unmodified N (p-N) or N(70−122) (p-N(70−122)) as an endogenous cytoplasmic protein or (ii) DNA encoding a lysosome-associated membrane protein (LAMP) chimera with N (p-LAMP/N) or N(70−122) (p-LAMP/N(70−122)). The immune responses of mice to these four constructs were evaluated. The results showed marked differences in the responses of the immunized mice. A single priming immunization with the p-LAMP/N construct was sufficient to elicit an antibody response. Enzyme-linked immunospot (ELISpot) assay indicated that p-LAMP/N(70−122) and p-LAMP/N plasmids both elicited a greater IFN-γ response than p-N. p-N and p-N(70−122) constructs induced low or undetectable levels of cytokine secretion. We also found that the p-LAMP/N(70−122) construct promoted a long-lasting T-cell memory response without an additional boost 6 months after three immunizations. These findings show that DNA vaccines, even epitope-based DNA vaccines using LAMP as chimera, can elicit both humoral and cellular immune responses.
Immune responses to T-cell epitopes of SARS CoV-N protein are enhanced by N immunization with a chimera of lysosome-associated membrane protein