Related Experiment Video
Updated: Aug 6, 2026

Myeloid Innate Signaling Pathway Regulation by MALT1 Paracaspase Activity
Published on: January 7, 2019
Clathrin heavy chain is required for TNF-induced inflammatory signaling
Guillermo A Escobar1, Robert C McIntyre, Ernest E Moore
1University of Colorado Health Sciences Center, Denver, CO 80262, USA.
Background:
Tumor necrosis factor receptor I recruits tumor necrosis factor receptor-associated death domain (TRADD) and multiple kinases that ultimately phosphorylate inhibitor kappa B (IKB alpha). Degradation of phospho-IKB alpha (p-IKB alpha) frees nuclear factor kappa B (NFKB) to be active and phosphorylated. Many receptors require clathrin-mediated endocytosis to provide the scaffolds necessary for signaling. Therefore, we investigated the role of clathrin heavy chain (CHC) in tumor necrosis factor alpha (TNF-alpha)-induced IKB alpha phosphorylation and NFKB activation. We hypothesized that CHC was required for TNF-alpha-induced inflammatory signaling.
Methods:
We treated human pulmonary epithelial cells with small interfering RNA to knock down intracellular CHC (CHCsil). TRADD and scrambled (noncoding) small interfering RNA sequences were used as positive and negative controls, respectively. Treatment groups were exposed to 10 ng/mL of TNF-alpha. Total I kappaB alpha, p-I kappaB alpha, and phosphorylated P65 (a subunit of NFKB) were determined by immunoblot staining. Densitometry was normalized to controls for the analysis of the stains. TNF-alpha-induced release of monocyte chemoattractant protein 1 (MCP-1) was determined by enzyme-linked immunosorbent assay. Statistical analyses were determined by analysis of variance or paired t test as appropriate.
Results:
TNF-alpha-induced I kappaB alpha phosphorylation and degradation at 5 and 30 minutes, respectively, and induced P65 phosphorylation. CHCsil diminished p-I kappaB alpha by 91% (P < .03); however, I kappaB alpha degradation was not affected. CHC knockdown caused a 66% decrease in P65 phosphorylation after 3 minutes of TNF-alpha. CHCsil decreased TNF-alpha-induced MCP-1 by 46% (P < .05), compared with control.
Conclusions:
CHCsil significantly impairs phosphorylation of both I kappaB alpha and P65. CHCsil also significantly decreased MCP-1 production. These data suggest that CHC is required for certain TNF-alpha-induced, inflammatory signaling pathways.
Related Concept Videos
NF-κB-dependent Signaling Pathway
NF-κB-dependent Signaling Mechanism
The heterodimer of NF-κB...
T Cell Activation and Clonal Selection
Naive T cells that have not yet encountered an antigen express two primary CD...
Drugs for Treatment of Crohn's Disease in IBD Using Biologic Agents: Anti-TNF
Intracellular Signaling Affects Focal Adhesions
Some...
NF-kB-dependent Signaling Pathway
NF-κB-dependent Signaling Mechanism
The heterodimer of NF-κB...
TGF - β Signaling Pathway
