title: The critical DNA flanking sequences of a CpG oligodeoxynucleotide, but not the 6 base CpG motif, can be replaced with RNA without quantitative or qualitative changes in Toll-like receptor 9-mediated activity.
abstract: Double- and single-stranded oligodeoxynucleotides containing unmethylated cytosine-guanosine (CpG) dinucleotides (CpG-ODN) activate immune cells via TLR9. In this report we synthesized hybrid DNA-RNA molecules (HDR) in order to further explore the structure-immune function relationship of CpG-ODN in TLR9 signaling and the potential immunomodulatory properties of RNA. We demonstrate that replacement of the deoxyadenosine flanking sequences, critical for the immune activating properties of CpG-ODN, with a similar number of adenosines, although not guanosines, cytosines, or uracils, maintains complete immunostimulatory activity of the hybrid oligonucleotide in vitro, whereas a similar RNA replacement of even 1 base of the required unmethylated 6 base DNA motif (purine-purine-CpG-pyrimidine-pyrimidine) results in a complete loss of activity. Regardless of whether the critical flanking sequence was RNA or DNA there was no significant change in the quantitative or qualitative immune-stimulating activity, or TLR-specificity of the resulting sequences, thus underscoring the relatively permissive functional role of the flanking sequence, and the more specific role of the motif in mediating TLR9 signaling. These data further support a potential role for RNA in immunomodulation.
authors.affiliation: Institute for Vaccine Research, Department of Pathology, Uniformed Services University of the Health Sciences, 4301 Jones Bridge Road, Bethesda, MD 20814, USA.
authors.forename: Goutam
authors.initials: G
authors.lastname: Sen
authors.forename: Michael
authors.initials: M
authors.lastname: Flora
authors.forename: Gouri
authors.initials: G
authors.lastname: Chattopadhyay
authors.forename: Dennis M
authors.initials: DM
authors.lastname: Klinman
authors.forename: Andrew
authors.initials: A
authors.lastname: Lees
authors.forename: James J
authors.initials: JJ
authors.lastname: Mond
authors.forename: Clifford M
authors.initials: CM
authors.lastname: Snapper
mesh_terms: D000818:Animals; D020029:Base Pairing; D001483:Base Sequence; D049109:Cell Proliferation / Q000187:drug effects; D016207:Cytokines / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D004247:DNA / Q000737:chemistry* / Q000276:immunology; D004268:DNA-Binding Proteins / Q000187:drug effects* / Q000276:immunology; D004305:Dose-Response Relationship, Drug; D007700:Kinetics; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D009692:Nucleic Acid Heteroduplexes / Q000138:chemical synthesis / Q000276:immunology / Q000494:pharmacology; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000276:immunology / Q000494:pharmacology*; D012313:RNA / Q000737:chemistry* / Q000276:immunology; D011956:Receptors, Cell Surface / Q000187:drug effects* / Q000276:immunology; D015398:Signal Transduction / Q000235:genetics / Q000276:immunology; D013154:Spleen / Q000166:cytology / Q000187:drug effects / Q000276:immunology; D013329:Structure-Activity Relationship; D051217:Toll-Like Receptor 9
chemical_list: C408982:CPG-oligonucleotide; D016207:Cytokines; D004268:DNA-Binding Proteins; D009692:Nucleic Acid Heteroduplexes; D009838:Oligodeoxyribonucleotides; D011956:Receptors, Cell Surface; C495370:Tlr9 protein, mouse; D051217:Toll-Like Receptor 9; D012313:RNA; D004247:DNA