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Title: Anti-WASP intrabodies inhibit inflammatory responses induced by Toll-like receptors 3, 7, and 9, in macrophages

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 bymore » associating with Btk in macrophages. - Highlights: • The interaction between WASP and Btk is critical for TLR3, TLR7, and TLR9 signaling. • Anti-WASP intrabodies inhibited several TLR pathways that led to cytokine expression. • Phosphorylation of NF-κB via TLR signaling was inhibited by anti-WASP intrabodies. • WASP phosphorylation via several TLR ligands was inhibited by anti-WASP intrabodies.« less
Authors:
 [1] ;  [1] ;  [2] ;  [1] ;  [2] ;  [1]
  1. Animal Immune and Cell Biology Research Unit, National Institute of Agrobiological Sciences, 1-2 Ohwashi, Tsukuba, Ibaraki, 305-8634 (Japan)
  2. 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)
Publication Date:
OSTI Identifier:
22458499
Resource Type:
Journal Article
Resource Relation:
Journal Name: Biochemical and Biophysical Research Communications; Journal Volume: 458; Journal Issue: 1; Other Information: Copyright (c) 2015 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; BONE MARROW; INFLAMMATION; LIGANDS; MACROPHAGES; MICE; MONOCLONAL ANTIBODIES; PHOSPHORYLATION; RECEPTORS; SIGNALS; TYROSINE