title: Anti-WASP intrabodies inhibit inflammatory responses induced by Toll-like receptors 3, 7, and 9, in macrophages.
abstract: Wiskott-Aldrich syndrome protein (WASP) is an adaptor molecule in immune cells. Recently, we showed that the WASP N-terminal domain interacted with the SH3 domain of Bruton's tyrosine kinase (Btk), and that the complex formed by WASP and Btk was important for TLR2 and TLR4 signaling in macrophages. Several other studies have shown that Btk played important roles in modulating innate immune responses through TLRs in immune cells. Here, we evaluated the significance of the interaction between WASP and Btk in TLR3, TLR7, and TLR9 signaling. We established bone marrow-derived macrophage cell lines from transgenic (Tg) mice that expressed intracellular antibodies (intrabodies) that specifically targeted the WASP N-terminal domain. One intrabody comprised the single-chain variable fragment and the other comprised the light-chain variable region single domain of an anti-WASP N-terminal monoclonal antibody. Both intrabodies inhibited the specific interaction between WASP and Btk, which impaired the expression of TNF-α, IL-6, and IL-1β in response to TLR3, TLR7, or TLR9 stimulation. Furthermore, the intrabodies inhibited the phosphorylation of both nuclear factor (NF)-κB and WASP in response to TLR3, TLR7, or TLR9 stimulation, in the Tg bone marrow-derived macrophages. These results suggested that WASP plays important roles in TLR3, TLR7, and TLR9 signaling by associating with Btk in macrophages.
authors: Animal Immune and Cell Biology Research Unit, National Institute of Agrobiological Sciences, 1-2 Ohwashi, Tsukuba, Ibaraki, 305-8634, Japan.
authors: Chisato
authors: C
authors: Sakuma
authors: Animal Immune and Cell Biology Research Unit, National Institute of Agrobiological Sciences, 1-2 Ohwashi, Tsukuba, Ibaraki, 305-8634, Japan. Electronic address: mitsuru.sato@affrc.go.jp.
authors: Mitsuru
authors: M
authors: Sato
authors: Department of Biological Science and Technology, Graduate School of Faculty of Industrial Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba, 278-8510, Japan.
authors: Takuma
authors: T
authors: Oshima
authors: Animal Immune and Cell Biology Research Unit, National Institute of Agrobiological Sciences, 1-2 Ohwashi, Tsukuba, Ibaraki, 305-8634, Japan.
authors: Takato
authors: T
authors: Takenouchi
authors: Department of Biological Science and Technology, Graduate School of Faculty of Industrial Science and Technology, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba, 278-8510, Japan.
authors: Joe
authors: J
authors: Chiba
authors: Animal Immune and Cell Biology Research Unit, National Institute of Agrobiological Sciences, 1-2 Ohwashi, Tsukuba, Ibaraki, 305-8634, Japan.
authors: Hiroshi
authors: H
authors: Kitani
mesh_terms: D000634:Aminoquinolines / Q000494:pharmacology; D000818:Animals; D000906:Antibodies / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D001854:Bone Marrow Cells / Q000166:cytology; D016207:Cytokines / Q000378:metabolism; D000077271:Imiquimod; D007249:Inflammation / Q000188:drug therapy* / Q000276:immunology / Q000378:metabolism; D008264:Macrophages / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism*; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D016328:NF-kappa B / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011070:Poly I-C / Q000494:pharmacology; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D051196:Toll-Like Receptor 3 / Q000276:immunology / Q000378:metabolism*; D051199:Toll-Like Receptor 7 / Q000276:immunology / Q000378:metabolism*; D051217:Toll-Like Receptor 9 / Q000276:immunology / Q000378:metabolism*; D051316:Wiskott-Aldrich Syndrome Protein / Q000235:genetics / Q000276:immunology*
chemical_list: D000634:Aminoquinolines; D000906:Antibodies; D016207:Cytokines; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D057127:Single-Chain Antibodies; C519703:TLR3 protein, mouse; C477038:Tlr7 protein, mouse; C495370:Tlr9 protein, mouse; D051196:Toll-Like Receptor 3; D051199:Toll-Like Receptor 7; D051217:Toll-Like Receptor 9; C495767:WAS protein, human; D051316:Wiskott-Aldrich Syndrome Protein; D011070:Poly I-C; D000077271:Imiquimod
keywords: Bruton's tyrosine kinase (Btk); Intracellular expressed antibody (intrabody); Macrophage; Toll-like receptors (TLRs); Wiskott-Aldrich syndrome protein (WASP)