Inhibition of TRAF6 ubiquitin-ligase activity by PRDX1 leads to inhibition of NFKB activation and autophagy activation
TRAF6 (TNF receptor associated factor 6) plays a pivotal role in NFKB activation and macroautphagy/autophagy activation induced by TLR4 (toll like receptor 4) signaling. The objective of this study was to determine the functional role of PRDX1 (peroxiredoxin 1) in NFKB activation and autophagy activation. PRDX1 interacted with the ring finger domain of TRAF6 and inhibited its ubiquitin-ligase activity. The inhibition on TRAF6 ubiquitin-ligase activity by PRDX1 induced the suppression of ubiquitination of an evolutionarily conserved signaling intermediate in Toll pathways (ECSIT) essential for NFKB activation and BECN1 (beclin 1) required for autophagy activation. An inhibitory effect of PRDX1 on TRAF6 was clearly evidenced in PRDX1-knockdown (PRDX1KD) THP-1, PRDX1KD MDA-MB-231, and PRDX1KD SK-HEP-1 cells. PRDX1KD THP-1 cells showed increases of NFKB activation, pro-inflammatory cytokine production, NFKB-dependent gene expression induced by TLR4 stimulation, and resistance against Salmonella typhimurium infection. Additionally, migration and invasion abilities of PRDX1KD MDA-MB-231 and PRDX1KD SK-HEP-1 cancer cells were significantly enhanced compared to those of control cancer cells. Taken together, these results suggest that PRDX1 negatively regulates TLR4 signaling for NFKB activation and autophagy functions such as bactericidal activity, cancer cell migration, and cancer cell invasion by inhibiting TRAF6 ubiquitin-ligase activity.
Abbreviations: 3-MA: 3-methyladenine; BECN1: beclin 1; CHUK/IKKA: conserved helix-loop-helix ubiquitous kinase; ECSIT: ECSIT signalling integrator; ELISA: enzyme-linked immunosorbent assay; NFKB: nuclear factor kappa-light-chain-enhancer of activated B cells; IB: immunoblotting; IKBKB/IKKB: inhibitor of nuclear factor kappa B kinase subunit beta; IL1B: interleukin 1 beta; IL6: interleukin 6; IP: immunoprecipitation; LPS: lipopolysaccharide; MAP1LC3/LC3: microtuble associated protein 1 light chain 3; MAP3K7/TAK1: mitogen-activated protein kinase kinase kinase 7; MAPK14/p38: mitogen-activated protein kinase 14; mROS: mitochondrial reactive oxygen species; PRDX1: peroxiredoxin 1; PRDX6: peroxiredoxin 6; RELA/p65: RELA proto-oncogene, NF-kB subunit; TRAF6 TNF: receptor associated factor 6.