Interaction of DJ-1 with Lyn is essential for IgE-mediated stimulation of human mast cells.

ABSTRACT: BACKGROUND:DJ-1 is a redox-sensitive protein with multiple roles in cell homeostasis, levels of which are altered in patients with mast cell (MC)-related disorders. However, whether DJ-1 can regulate human MC function is unknown. OBJECTIVE:We sought to investigate the potential role of DJ-1 in the responses of human MCs to antigen stimulation. METHODS:DJ-1 was silenced in human CD34+-derived MCs and in the LAD2 MC line by using lentiviral short hairpin RNA constructs. Release of ?-hexosaminidase, prostaglandin D2, and GM-CSF and changes in reactive oxygen species levels were measured after Fc?RI engagement. Enzymatic assays, sucrose density gradient centrifugation, immunoprecipitation, dot and Western blotting, and confocal imaging were performed for signaling, cellular localization, and coassociation studies. RESULTS:DJ-1 knockdown substantially reduced mediator release, as well as Lyn kinase and spleen tyrosine kinase activation and signaling through mechanisms that appeared largely unrelated to DJ-1 antioxidant activity. Following Fc?RI activation, nonoxidized rather than oxidized DJ-1 translocated to lipid rafts, where it associated with Lyn, an interaction that appeared critical for maximal Lyn activation and initiation of signaling. Using purified recombinant proteins, we demonstrated that DJ-1 directly bound to Lyn but not to other Src kinases, and this interaction was specific for human but not mouse proteins. In addition, DJ-1 reduced Src homology 2 domain-containing phosphatase 2 phosphatase activity by scavenging reactive oxygen species, thus preventing spleen tyrosine kinase dephosphorylation and perpetuating MC signaling. CONCLUSION:We demonstrate a novel role for DJ-1 in the early activation of Lyn by Fc?RI, which is essential for human MC responses and provides the basis for an alternative target in allergic disease therapy.

SUBMITTER: Kim DK 

PROVIDER: S-EPMC5899056 | biostudies-literature | 2018 Jul

REPOSITORIES: biostudies-literature