Tumor necrosis factor α modifies agonist-dependent responses in human neutrophils by inducing the synthesis and myristoylation of a specific protein kinase C substrate

Marcus Thelen, Antony Rosen, Angus C. Nairn, Alan Aderem

Research output: Contribution to journalArticle

Abstract

Tumor necrosis factor α (TNF-α) and bacterial lipopolysaccharide (LPS) induce the synthesis and cotranslational myristoylation of an 82-kDa specific protein kinase C substrate in human neutrophils. The myristic acid is covalently bound via a hydroxylamine-resistant amide linkage to the N-terminal glycine of the protein. The isoelectric point of the protein is at pH 4.6. The protein is rapidly phosphorylated when neutrophils are stimulated with chemotactic agonists or with phorbol 12-myristate 13-acetate, an activator of protein kinase C, and displays two characteristic phosphopeptides in one- and two-dimensional separation systems. Identical phosphopeptides were detected when the 82-kDa protein was phosphorylated in vitro with purified kinase C. The 82-kDa protein was immunoprecipitated by a polyclonal antiserum raised against the 87-kDa specific protein kinase C substrate from bovine brain. From these biochemical and immunological criteria it is concluded that the 82-kDa protein is the human neutrophil homolog of MARCKS, the myristoylated, alaninerich C kinase substrate previously described in bovine and rat brain and in murine fibroblasts and macrophages. TNF-α and LPS prime human neutrophils for potentiated protein kinase C-dependent responses such as the respiratory burst and exocytosis. Consistent with this, these mediators do not induce the phosphorylation of MARCKS but prime the neutrophils for enhanced phosphorylation of this protein when the cells subsequently encounter activators of protein kinase C. This increase in MARCKS phosphorylation can be explained by the elevated levels of the protein observed in TNF-α- or LPS-treated neutrophils. Indeed, MARCKS constitutes 90% of all proteins synthesized in response to TNF-α or LPS. These data strongly suggest that MARCKS acts as a critical effector molecule in the transduction pathway of these important inflammatory mediators.

Original languageEnglish (US)
Pages (from-to)5603-5607
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume87
Issue number15
Publication statusPublished - 1990
Externally publishedYes

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Keywords

  • N-myristoylglycine
  • Signal transduction

ASJC Scopus subject areas

  • Genetics
  • General

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