Serine phosphorylation of p60 tumor necrosis factor receptor by PKC-δ in TNF-α-activated neutrophils

Abstract

Tumor necrosis factor-α (TNF-α) triggers degranulation and oxygen radical release in adherent neutrophils. The p60TNF receptor (p60TNFR) is responsible for proinflammatory signaling, and protein kinase C (PKC) is a candidate for the regulation of p60TNFR. Both TNF-α and the PKC-activator phorbol 12-myristate 13-acetate triggered phosphorylation of p60TNFR. Receptor phosphorylation was on both serine and threonine but not on tyrosine residues. The PKC-δ isotype is a candidate enzyme for serine phosphorylation of p60TNFR. Staurosporine and the PKC-δ inhibitor rottlerin inhibited TNF-α-triggered serine but not threonine phosphorylation. Serine phosphorylation was associated with receptor desensitization, as inhibition of PKC resulted in enhanced degranulation (elastase release). After neutrophil activation, PKC-δ was the only PKC isotype that associated with p60TNFR within the correct time frame for receptor phosphorylation. In vitro, only PKC-δ, but not the α-, βI-, βII-, or ζ-isotypes, was competent to phosphorylate the receptor, indicating that p60TNFR is a direct substrate for PKC-δ. These findings suggest a selective role for PKC-δ in negative regulation of the p60TNFR and of TNF-α-induced signaling.