Lipid rafts constrain basal alpha(1A)-adrenergic receptor signaling by maintaining receptor in an inactive conformation.
We have reported that the alpha(1A)-adrenergic receptor (alpha(1A)AR) in rat-1 fibroblasts is a lipid raft protein. Here we examined whether disrupting lipid rafts by methyl-beta-cyclodextrin (MCD) sequestration of cholesterol affects alpha(1A)AR signaling. Unexpectedly, MCD increased alpha(1A)AR-dependent basal inositol phosphate formation and p38 mitogen-activated protein kinase activation in a cholesterol-dependent manner. It also initiated internalization of surface alpha(1A)AR, which was partially blocked by receptor inhibition. Binding assays revealed MCD-mediated increases in receptor agonist affinity as well as reciprocal decreases in inverse agonist affinity, a behavior that is usually interpreted as a shift toward the active receptor conformation. In untreated cells a fraction of the receptor was found to be present in preassociated receptor/G protein complexes, which rapidly dissociate upon receptor stimulation. Consistent with MCD-induced signaling, raft disruption resulted in an increase in receptor/G protein complexes. These results strongly suggest that lipid rafts constrain basal alpha(1A)AR activity; however, preassembled receptor/G protein complexes could still provide a mechanism for accelerating alpha(1A)AR signaling following stimulation.
Lei, B; Morris, DP; Smith, MP; Schwinn, DA
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