Structural requirements for uptake and recognition of CpG oligonucleotides

Recognition of infectious danger by innate immune cells is a fundamental requirement to directly combat infections and to activate the adaptive immune response of T and B cells. Pathogen-associated molecular patterns (PAMPs) play a fundamental role in this process. PAMPs are sensed by pattern recognition receptors, among which the Toll-like receptors (TLRs) play an important role. Within the TLR ligands, bacterial CpG DNA is peculiar. CpG DNA is recognized by TLR9 and harbors the outstanding propensity to induce a milieu that favors activation of Th1-dominated immune responses. This is mainly due to activation of dendritic cells and subpopulations, thereby inducing an intense interferon and IL-12 response. Therefore, CpG DNA has become a promising candidate for constructing new vaccines as well as for induction of immune responses in cancer and allergy. CpG DNA can be synthesized with high purity, defined base composition and various chemical modifications. We aimed to understand the structural requirements for cellular uptake and activation of CpG DNA, which will improve our means to enhance the intrinsic activity of CpG for therapeutic use. We show that sequence modifications can be utilized to enhance cellular uptake, and that chemical substitutions can confer new qualities to synthetic CpG DNA. Additionally, we propose a model of CpG DNA recognition which occurs by sensing partial duplex forms instead of single-stranded DNA. Moreover, we provide evidence that the propensity of CpG DNA to induce high amounts of IL-12 is due to its unique ability to trigger epigenetic modifications of the IL-12p40 promoter, including acetylation and nucleosomal remodeling. Hence, CpG DNA represents a new and promising class of adjuvant for vaccination.