Toll-like receptor 4 signaling is required for clusterin-induced tumor necrosis factor-α secretion in macrophage

Young Jun Shim, Yoo Keung Tae, Byeong Ho Kang, Jin Sung Park, Sol Yi Jeon, Bon Hong Min

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Clusterin is a secretory glycoprotein that is up-regulated in areas of inflammation and under increased levels of oxidative stress. Previously, we demonstrated that clusterin activates NF-κB, and up-regulates the expression of MMP-9 and TNF-α. In this research, we extend our previous findings by reporting that such clusterin-induced macrophage response is mediated via TLR4 signaling. Specifically, we found that TNF-α induced by clusterin was significantly abrogated by pretreatment of TLR4-signaling inhibitors and anti-TLR4 neutralizing antibody. Additionally, a primary culture of macrophages derived from TLR4-signal defective and knockout mice were unresponsive to clusterin, resulting in no TNF-α secretion, whereas macrophages carrying wild-type TLR4 responded to clusterin and induced TNF-α. Moreover, clusterin increased NF-κB promoter activity in HEK-Blue hTLR4 cells, but not in HEK-Blue Null2 cells. To confirm that clusterin elicits TLR4 signal transduction, recombinant clusterin was generated and purified from cell culture. Interestingly, we found that the recombinant clusterin with C-terminal HA-tag induces TNF-α secretion at a significantly lower level compared to an intact form of clusterin without C-terminal HA-tag. Removal of HA-tag from the recombinant clusterin restored its activity, suggesting that C-terminal HA-tag partially masks the domain involved in TLR4 signaling. Furthermore, clusterin enhanced TLR4 mobilization into lipid raft of plasma membrane, and TNF-α and MMP-9 secretion stimulated by clusterin was diminished by pretreatment with methyl-β-cyclodextrin (MβCD), which was used to disrupt lipid raft. In conclusion, clusterin-induced TNF-α and MMP-9 up-regulation is most likely mediated via TLR4 recruitment into lipid rafts, and these data describe a novel role of clusterin as an endogenous regulator for TLR4 signaling.

Original languageEnglish
Pages (from-to)1407-1412
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume482
Issue number4
DOIs
Publication statusPublished - 2017 Jan 22

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Keywords

  • Clusterin
  • Lipid raft
  • Macrophage
  • NF-κB
  • TLR4
  • TNF

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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