Differential Use of the Signaling Molecules BTK and PLC-γ in Mast Cells Stimulated Via the Receptors FcεRI and KIT

Abstract

Mast cells belong to the myeloid lineage of the hemopoietic system, are localized to the tissue-environment interphase, and execute regulatory and protective roles in both innate and adaptive immunity. However, mast cells also are involved in pathophysiological processes ranging from allergic diseases to autoimmune diseases and tumors. Two receptors expressed on the surface of mast cells and prominently involved in physiology and pathophysiology of mast cell biology are the high-affinity receptor for IgE (FcεRI) and the receptor tyrosine kinase KIT (a.k.a. CD117), the receptor for stem cell factor (SCF). Whereas antigen (Ag)-triggered activation of the IgE-bound FcεRI results in immediate release of preformed mediators (degranulation) and production of arachidonic acid metabolites and cytokines/chemokines, SCF-induced mast cell activation is only causing the latter. On the other side, SCF is inducing mast cell proliferation and chemotaxis, two responses that are not caused by FcεRI activation. Intriguingly, the signal transduction downstream of these receptors involves activation of the same signaling pathways and engagement of comparable sets of signaling proteins, raising the question, how differential mast cell effector functions are controlled by these receptors. Therefore, it is essential to learn more about the molecular pathways leading to FcεRI and KIT-mediated activation of mast cells. In the presented work, primary murine mast cells were stimulated either with Ag via the IgE-bound FcεRI or with SCF via its receptor KIT and differential signal transduction was analyzed. We put our focus on two central signaling elements in mast cells, the cytosolic tyrosine kinase BTK and its target, phospholipase C-γ, and also addressed differences between using suboptimal and optimal ligand concentrations. Cell responses like degranulation, proinflammatory cytokine production, activating protein phosphorylation, and intracellular calcium mobilization were analyzed making use of mast cells from wild-type and transgenic (knock-out) mice as well as selective pharmacological inhibitors. Interestingly, marked differences were found with respect to pathway activation, importance of particular signaling proteins, and induction of effector responses. Ag-triggered degranulation is thought to be dependent on BTK/PLC-γ-mediated calcium mobilization and thus it was unexpected to observe only a weak effect of PLC-γ inhibition. On the other side, SCF-induced calcium mobilization was strictly dependent on PLC-γ, however, only a minor involvement of BTK could be observed. In combination with the additional results obtained in our study, future analyses of differential pathway organization by multiprotein assemblies as well as kinetic control mechanisms appear to be important. Detailed knowledge of these mechanisms will help to precisely promote beneficial and strictly control detrimental mast cell responses. DisclosuresNo relevant conflicts of interest to declare.