Role of CpG motifs in immunostimulation and gene expression
The first demonstration that bacterial DNA was in fact immunostimulatory came in 1984 when investigators discovered that the anti-tumor activity of the Mycobacterium bovis bacillus Calmette-Guerin (BCG) could be isolated to its deoxyribonucleic acid fraction (Tokunaga et al., 1984). Subsequently, in 1991, investigators demonstrated that single stranded DNA (ssDNA) from E. coli, but not mammalian DNA, was capable of stimulating murine splenocytes in vitro in a dose dependent manner (Messina et al., 1991). This stimulation was blocked following treatment with DNAse but was not affected when using cells from the endotoxin-resistant C3H/HeJ mouse, therefore ruling out a possible role for endotoxin contamination. Further, the elimination of T cell populations did not reduce the extent of splenocyte proliferation, suggesting that the predominant cell type induced to proliferate were B cells. In 1992 investigators discovered that the immunostimulatory properties of bacterial DNA could be reproduced using synthetically derived oligodeoxynucleotides (ODNs). The sequences of these ODNs contained palindromes (e.g., AACGTT) that were thought to be responsible for the immune stimulatory activity (Kataoka et al., 1992; Tokunaga et al., 1992; Yamamoto et al., 1992).