Immunobiology of the TLR7, TLR8 and TLR9 subfamily

H. Wagner

Institut für Medizinische Mikrobiologie, Immunologie und Hygiene, Technische Universität München, Trogerstr. 9, 81675 München

According to their genomic organisation and sequence similarities, Toll like receptor (TLR) 7, 8 and 9 are related and may be grouped to a subfamily within the 10 mammalian TLRs known. Furthermore, evidence available indicates that TLR7, 8 and 9 are expressed within innate immune cells at vesicular cytoplasmatic structures while the other TLRs appear cell surface bound. This raises the question whether the physiological ligands of the TLR7, 8, 9 subfamily primarily derive from intracellular pathogens. In support CpG-DNA motifs within bacterial genomic DNA and certain DNA viruses (Herpes-family) stimulate innate immune cells via TLR9 in a MyD88 dependent fashion. While some synthetic small antiviral molecules appear to be recognized via TLR7, the physiological ligands of TLR7 and TLR8 are yet unknown.

Ongoing work with PD Dr. Stefan Bauer and his group implies that uridine rich single stranded (ss) RNA-motifs (derived from U5 within the HIV-LTR) activate both murine plasmacytoid and myeloid dendritic cells (DCs) in a MyD88 dependent fashion: the former producing primarily Type I Interferon, the latter proinflammatory cytokines. Furthermore, human pDC₂ (plasmacytoid precursors) become activated to produce Type I interferon, while cells of myeloid lineage primarily produce TNF-a. By genetic complementation we find that human TLR8 confers responsiveness to ss Oligonucleotides containing the stimulatory RNA motif, but not TLR1 – 7 and TLR9. Based on this, we tentatively propose that human TLR8 recognize as physiological ligand ss RNA motifs rich in uridine. In collaboration with the group of Akira from Japan the responsiveness phenotype to the ss RNA motif in TLR7 and TLR8 deficient mice is presently being analyzed.

Ligands of the TLR family are considered as robust adjuvants in vaccination protocols using rec. antigen (split Ag). Immunostimulatory CpG-DNA bears promising characteristics. In this regard we analyzed the immunogenity of rec. protein covalently linked with CpG-DNA. In this system the CpG-protein conjugate becomes endocytozed by Antigen-presenting cells (APCs) (via a yet poorly defined DNA recognising receptor) and the “loaded” APCs become in parallel activated via endocytozed CpG-DNA. Use of the Tetramer-Technology (in collaboration with the group of Dr. Dirk Busch) revealed an amazing immunogenicity of CpG-protein conjugates which induced clonal expansion of Tetramer Epitope binding CD8 T cells up to 4 – 10 %.