Requirements for effective inhibition of immunostimulatory CpG motifs by neutralizing motifs

Antisense Nucleic Acid Drug Dev. 2000 Oct;10(5):381-9. doi: 10.1089/oli.1.2000.10.381.

Abstract

The DNA of bacteria and many viruses contain unmethylated CpG dinucleotides in particular sequence contexts that activate vertebrate immune cells. A subset of these CpG motifs was previously found to oppose the effects of immunostimulatory (CpG-S) motifs and has been termed neutralizing (CpG-N) motifs. Here we show that oligodeoxynucleotides (ODNs) composed of clusters of CpG-N motifs could partially inhibit the induction of interleukin-12 (IK-12) from mouse spleen cells by ODN containing CpG-S motifs. However, non-CpG-containing ODN were also inhibitory, suggesting that neutralization of CpG-S ODNs by CpG-N ODNs in trans was nonspecific. Neutralization of CpG-S motifs by CpG-N motifs in cis was specific, but the degree of inhibition was strongly dependent on the particular CpG-S motif being neutralized, with motifs having an A residue 5' to the CG being much more resistant to inhibition than motifs having a T residue 5' to the CG. The degree of inhibition was dependent on the spacing between the CpG-S and CpG-N motifs, with the ability to neutralize inversely correlating with distance. In addition, whereas ODNs containing extended clusters of CpG-N motifs were nonstimulatory, isolated CpG-N motifs remained stimulatory in most sequence contexts. Finally, CpG-N ODNs were shown to be nonstimulatory when instilled into the lungs of BALB/c mice, but the ability of CpG-N motifs to neutralize CpG-S motifs in cis was not observed. These results show that there are precise and fairly complex interactions between immunostimulatory and inhibitory sequence motifs that govern whether a given DNA is able to activate the vertebrate immune system.

MeSH terms

  • Adjuvants, Immunologic / genetics*
  • Administration, Inhalation
  • Animals
  • Base Sequence
  • Cells, Cultured
  • CpG Islands / genetics*
  • CpG Islands / immunology*
  • Female
  • Genetic Therapy
  • Interleukin-12 / biosynthesis
  • Interleukin-12 / immunology
  • Lung / metabolism
  • Mice
  • Mice, Inbred BALB C
  • Oligodeoxyribonucleotides / chemistry
  • Oligodeoxyribonucleotides / genetics
  • Oligodeoxyribonucleotides / immunology*
  • Oligodeoxyribonucleotides / pharmacology*
  • Spleen / metabolism

Substances

  • Adjuvants, Immunologic
  • Oligodeoxyribonucleotides
  • Interleukin-12