sNASP inhibits TLR signaling to regulate immune response in sepsis
Feng-Ming Yang, … , Hui-Ming Chang, Edward T.H. Yeh
Feng-Ming Yang, … , Hui-Ming Chang, Edward T.H. Yeh
Published May 7, 2018
Citation Information: J Clin Invest. 2018;128(6):2459-2472. https://doi.org/10.1172/JCI95720.
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Research Article Immunology Inflammation

sNASP inhibits TLR signaling to regulate immune response in sepsis

Abstract

Many Toll-like receptors (TLRs) signal through TNF receptor–associated factor 6 (TRAF6) to activate innate immune responses. Here, we show that somatic nuclear autoantigenic sperm protein (sNASP) binds to TRAF6 to prevent TRAF6 autoubiquitination in unstimulated macrophages. Following LPS stimulation, a complex consisting of sNASP, TRAF6, IRAK4, and casein kinase 2 (CK2) is formed. CK2 phosphorylates sNASP at serine 158, allowing sNASP to dissociate from TRAF6. Free TRAF6 is then autoubiquitinated, followed by activation of downstream signaling pathways. In sNasp S158A knockin (S158A-KI) mice, LPS-treated macrophages could not phosphorylate sNASP, which remained bound to TRAF6. S158A-KI mice were more susceptible to sepsis due to a marked reduction in IL-1β, TNF-α, and IFN-γ production accompanied by an inability to clear bacteria and recruit leukocytes. Furthermore, phosphorylation-regulated release of sNASP from TRAF6 is observed following activation of TLR-1, -2, -4, -5, and -6. Thus, sNASP is a negative regulator of TLR signaling to modulate the innate immune response.

Authors

Feng-Ming Yang, Yong Zuo, Wei Zhou, Chuan Xia, Bumsuk Hahm, Mark Sullivan, Jinke Cheng, Hui-Ming Chang, Edward T.H. Yeh

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Figure 5

CK2 phosphorylates sNASP on serine 158.

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CK2 phosphorylates sNASP on serine 158. (A) Alignment of sNASP sequence ...
(A) Alignment of sNASP sequence from multiple species revealed that serine 158 is highly conserved and contained in a motif recognized by CK2. X is any residue. (B) THP-1 cells stimulated with LPS in the absence (–) or presence (+) of TBB, an inhibitor of CK2, assessed by IB with antibody against phosphorylated serine (pSerine) or GFP after IP with anti-NASP. (C) THP-1 cells were stimulated with LPS in the presence of siNT or siCK2 and assessed by IB with antibody against phosphorylated serine (pSerine) or anti-GFP after IP with anti-NASP. TCL IB was done with anti-CK2α. (D) HEK293 cells transduced with GFP-tagged WT sNASP, S158A, or empty vector (C3) in the presence of Myc-tagged CK2β combined with HA-tagged CK2 catalytic subunits (CK2α and CK2α’) or CK2 kinase-dead mutants (CK2α K68M and CK2α’ K69M), assessed by IB with antibody against phosphorylated serine (pSerine) from Qiagen or Abcam or anti-GFP after IP with anti-GFP. TCL was IB with anti-HA or anti-Myc. (E) THP-1 cells transduced with GFP-tagged WT sNASP (WT) or S158A mutant in the absence (–) or presence (+) of TBB, assessed by IB with antibody against TRAF6 or GFP after IP with anti-TRAF6. TCL IB was done with anti-NASP and anti–β-actin. Data represent a minimum of 3 independent experiments.