Phosphorylation of tumor necrosis factor receptor CD120a (p55) by p42(mapk/erk2) induces changes in its subcellular localization

The interaction of tumor necrosis factor-alpha (TNF alpha) with its recepto r sets in motion downstream signaling events including the activation of me mbers of the mitogen-activated protein kinase (MAPK) family. In this study, we show that p42(mapk/erk2) phosphorylates sequences present within the cy toplasmic domain of CD120a (p55). By using a GST-CD120a-(207-425) fusion pr otein as substrate, phosphorylation was induced following stimulation of mo use macrophages with TNF alpha, granulocyte-macrophage colony-stimulating f actor, macrophage colony-stimulating factor, and zymosan particles and was blocked by immunodepletion of p42(mapk/erk2) and by specific inhibition of p42(mapk/erk2) activation with PD098059. Transfection of COS-7 cells with C D120a (p55), wild type p42(mapk/erk2), and constitutively active MEK-1 foll owed by metabolic labeling with [P-32]orthophosphate indicated that p42(map k/erk2) phosphorylated the cytoplasmic domain of CD120a (p55) in intact cel ls. As a consequence of phosphorylation, CD120a (p55) expression at the pla sma membrane and Golgi apparatus was lost and the receptor accumulated in i ntracellular tubular structures associated with the endoplasmic reticulum. Mutation of the four Ser and Thr ERK consensus phosphorylation sites to Ala residues inhibited the ability of the receptor to redistribute to intracel lular tubules in a p42(mapk/erk2)-dependent fashion; whereas mutation of th e phosphorylation sites to Asp and Glu residues mimicked the effect of rece ptor phosphorylation. These findings thus indicate that the phosphorylation of CD120a (p55) alters the subcellular localization of the receptor and ma y thereby result in changes in its signaling properties.