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Transmembrane TNF (pro-TNF) is palmitoylated
T Utsumi1, T Takeshige, K Tanaka
1Department of Biological Chemistry, Faculty of Agriculture, Yamaguchi University, Yamaguchi 753-8515, Japan. utsumi@agr.yamaguchi-u.ac.jp
FEBS Letters
|July 4, 2001
Summary
Human tumor necrosis factor (pro-TNF) undergoes palmitoylation, a lipid modification. This process attaches palmitic acid to pro-TNF via a thioester bond at Cys-47, anchoring it to the plasma membrane.
Area of Science:
- Biochemistry
- Cell Biology
- Molecular Biology
Background:
- Tumor necrosis factor (TNF) is a key cytokine involved in inflammation and immunity.
- Transmembrane TNF (tmTNF) exists as a precursor form (pro-TNF) before processing.
- Post-translational modifications like acylation can regulate protein function and localization.
Purpose of the Study:
- To investigate protein acylation of human transmembrane tumor necrosis factor (pro-TNF).
- To identify the specific type of acylation and the residue involved in pro-TNF modification.
- To elucidate the mechanism by which pro-TNF interacts with the plasma membrane.
Main Methods:
- Expression of human pro-TNF cDNA in COS-1 and Sf9 cells.
- Metabolic labeling using [(3)H]myristic acid and [(3)H]palmitic acid.
- Hydroxylamine treatment to assess thioester bond involvement.
- Site-directed mutagenesis of cysteine residues in the pro-TNF leader sequence.
Main Results:
- The 26 kDa pro-TNF precursor was specifically labeled with [(3)H]palmitic acid, while the 17 kDa mature TNF was not.
- Palmitoylation occurred via a thioester bond, as indicated by hydroxylamine sensitivity.
- Mutagenesis identified Cys-47 as the sole site of palmitoylation, located at the transmembrane/cytoplasmic domain boundary.
Conclusions:
- Human pro-TNF undergoes specific palmitoylation at Cys-47.
- This lipid modification via a thioester bond contributes to pro-TNF's interaction with the plasma membrane.
- Pro-TNF utilizes both its transmembrane domain and a lipid anchor for membrane association.