Janus kinase 2 activation by the platelet-activating factor receptor (PAFR): roles of Tyk2 and PAFR C terminus.
Platelet-activating factor (PAF) is a phospholipid with multiple physiological and pathological actions. The PAF receptor (PAFR) belongs to the G protein-coupled, heptahelical receptor superfamily. Recently, we have shown that PAF signals through the Janus kinase (Jak)/STAT pathway and that Tyk2 plays an essential role in PAF-induced PAFR promoter 1 activation. ... In the present study we found that PAF stimulated Jak2 tyrosine phosphorylation in the monocytic cell line MonoMac-1 as well as in COS-7 cells transfected with PAFR and Jak2 cDNAs. The use of a G protein-uncoupled PAFR (D289A) mutant indicated that Jak2 activation was G protein independent. Interestingly, following PAF stimulation, Jak2 coimmunoprecipitated with PAFR in the presence of active Tyk2, but not with a kinase-inactive Tyk2 mutant, K930I. Moreover, Tyk2-K930I completely blocked PAF-stimulated Jak2 phosphorylation. Gradual deletion of C-terminal residues of the PAFR resulted in progressively decreased Jak2 activation. Deletion of 12 C-terminal residues in mutant V330Stop diminished Jak2 tyrosine phosphorylation by 17%. Further deletions of 25-37 residues from the PAFR C-tail (C317Stop, M311Stop, and T305Stop) resulted in a 50% decrease in Jak2 phosphorylation compared with the wild-type receptor. Complete removal of the C tail resulted in a mutant (K298Stop) that failed to activate Jak2, suggesting that the receptor C-terminal region contains important domains for Jak2 activation. Finally, the coexpression of a minigene encoding the C terminus of PAFR partially inhibited PAF-induced kinase activation. Taken together, our results indicate that PAF activates Jak2 and that Tyk2 and the C-terminal tail of PAFR are of critical importance for PAF-induced Jak2 activation.
Mesh Terms:
Animals, COS Cells, Catalysis, Cell Line, Cercopithecus aethiops, DNA, Complementary, Enzyme Activation, Humans, Janus Kinase 2, Monocytes, Peptide Fragments, Phosphorylation, Platelet Activating Factor, Platelet Membrane Glycoproteins, Protein-Tyrosine Kinases, Proteins, Proto-Oncogene Proteins, Receptors, G-Protein-Coupled, Sequence Deletion, Signal Transduction, TYK2 Kinase, Transfection
Animals, COS Cells, Catalysis, Cell Line, Cercopithecus aethiops, DNA, Complementary, Enzyme Activation, Humans, Janus Kinase 2, Monocytes, Peptide Fragments, Phosphorylation, Platelet Activating Factor, Platelet Membrane Glycoproteins, Protein-Tyrosine Kinases, Proteins, Proto-Oncogene Proteins, Receptors, G-Protein-Coupled, Sequence Deletion, Signal Transduction, TYK2 Kinase, Transfection
J. Immunol.
Date: Oct. 01, 2003
PubMed ID: 14500680
View in: Pubmed Google Scholar
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