Cleavage of Toll-Like Receptor 9 Ectodomain Is Required for In Vivo Responses to Single Strand DNA

Ryutaro Fukui; Chikako Yamamoto; Fumi Matsumoto; Masahiro Onji; Takuma Shibata; Yusuke Murakami; Atsuo Kanno; Takuto Hayashi; Natsuko Tanimura; Nobuaki Yoshida; Kensuke Miyake

doi:10.3389/fimmu.2018.01491

Cleavage of Toll-Like Receptor 9 Ectodomain Is Required for In Vivo Responses to Single Strand DNA

Front Immunol. 2018 Jun 27:9:1491. doi: 10.3389/fimmu.2018.01491. eCollection 2018.

Authors

Ryutaro Fukui¹, Chikako Yamamoto¹, Fumi Matsumoto¹, Masahiro Onji^{1

2}, Takuma Shibata¹, Yusuke Murakami^{1

3}, Atsuo Kanno¹, Takuto Hayashi¹, Natsuko Tanimura¹, Nobuaki Yoshida⁴, Kensuke Miyake^{1

5}

Affiliations

¹ Division of Innate Immunity, Department of Microbiology and Immunology, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
² Institute of Molecular Biotechnology of the Austrian Academy of Sciences, Vienna, Austria.
³ Department of Pharmacotherapy, Research Institute of Pharmaceutical Sciences, Musashino University, Tokyo, Japan.
⁴ Laboratory of Developmental Genetics, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
⁵ Laboratory of Innate Immunity, Center for Experimental Medicine and Systems Biology, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.

Abstract

Mouse toll-like receptor 9 (TLR9) is an endosomal sensor for single-stranded DNA. TLR9 is transported from the endoplasmic reticulum to endolysosomes by a multiple transmembrane protein Unc93 homolog B1, and proteolytically cleaved at its ectodomain. The structure of TLR9 and its biochemical analyses have shown that the proteolytic cleavage of TLR9 ectodomain enables TLR9-dimerization and TLR9 activation. However, the requirement of TLR9 cleavage in vivo has not been studied. We here show that the 13 amino acids deletion at the cleavage site made TLR9 resistant to proteolytic cleavage. The deletion mutation in the Tlr9 gene impaired TLR9-dependent cytokine production in conventional dendritic cells from the mutant mice. Not only in vitro, in vivo production of inflammatory cytokines (TNF-α and IL-12p40), chemokine (CCR5/RANTES), and type I interferon (IFN-α) induced by administration of TLR9 ligand was also impaired. These results demonstrate that the TLR9 cleavage is required for TLR9 responses in vivo.

Keywords: CpG-ODN; Unc93 homolog B1; in vivo response; primary immune cells; proteolytic cleavage; toll-like receptor 9.