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

AbstractThe high-affinity IgE receptor FcεRI is the fundamental mast cell (MC) receptor responsible for the involvement of MCs in IgE-associated allergic disorders. Activation of the FcεRI 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 FcεRI-associated proteins. Additionally, it is not completely clear how FcεRI signaling synergizes with input from other receptors and how potential regulators affect this synergistic response. We aimed at identifying new regulators of FcεRI and found that the HDL receptor SR-BI (gene name:Scarb1/SCARB1) is expressed in MCs, functionally associates with FcεRI 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 FcεRI with different receptors known to synergize with FcεRI-signaling pathways. Amongst them we investigated the co-activation of the FcεRI 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. MimickingScarb1deficiency by membrane cholesterol depletion employing MβCD, we identified PKB and PLCγ1 as cholesterol-sensitive signaling molecules activated downstream of FcεRI in BMMCs. Specifically, when murine MCs were stimulated with SCF and Ag in combination, PLCγ1 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 PGE2and anti-IgE in the human ROSAKITWTmast cell line suggesting that SR-BI is a crucial regulator of synergistic MC activation by regulating the local plasma membrane cholesterol concentration.