SR-BI regulates the synergistic mast cell response by modulating the plasma membrane-associated cholesterol pool

Abstract

The high-affinity IgE receptor Fc{varepsilon}RI is the fundamental mast cell (MC) receptor responsible for the involvement of MCs in IgE-associated allergic disorders. Activation of the Fc{varepsilon}RI is achieved via crosslinking by multivalent antigen (Ag) recognized by IgE, which results in degranulation and pro-inflammatory cytokine production. In comparison to the T and B cell receptor complexes, for which several co-receptors that orchestrate the initial signaling have been described, information is scarce about Fc{varepsilon}RI-associated proteins. Additionally, it is not completely clear how Fc{varepsilon}RI signaling synergizes with input from other receptors and how potential regulators affect this synergistic response. We aimed at identifying new regulators of Fc{varepsilon}RI and found that the HDL receptor SR-BI (gene name: Scarb1/SCARB1) is expressed in MCs, functionally associates with Fc{varepsilon}RI and regulates the local plasma membrane cholesterol content in cholesterol-rich plasma membrane nanodomains as shown by using the cholesterol-sensitive probe GFP-D4. This impacted on the activation of murine MCs upon co-stimulation of the Fc{varepsilon}RI with different receptors known to synergize with Fc{varepsilon}RI-signaling pathways. Amongst them we investigated the co-activation of the Fc{varepsilon}RI with the receptor tyrosine kinase KIT, the IL-33 receptor and GPCRs activated by adenosine or PGE2. Scarb1-deficient bone marrow-derived MCs (BMMCs) showed reduced cytokine secretion in response to these co-stimulation conditions suggesting a role for plasma membrane-associated cholesterol regulating MC-driven inflammation. Mimicking Scarb1 deficiency by membrane cholesterol depletion employing M{beta}CD, we identified PKB and PLC{gamma}1 as cholesterol-sensitive signaling molecules activated downstream of Fc{varepsilon}RI in BMMCs. Specifically, when murine MCs were stimulated with SCF and Ag in combination, PLC{gamma}1 activation appeared to be drastically boosted and this could be mitigated by cholesterol depletion. Inhibiting SR-BI in BMMCs phenocopied this effect. Similarly, SR-BI inhibition also attenuated the synergistic response to PGE2 and anti-IgE in the human ROSAKIT WT mast cell line suggesting that SR-BI is a crucial regulator of synergistic MC activation by regulating the local plasma membrane cholesterol concentration.