Membrane-permeable tastants amplify beta 2-adrenergic receptor signaling and delay receptor desensitization via intracellular inhibition of GRK2's kinase activity

作者：Malach Einav; Shaul Merav E; Peri Irena; Huang Liquan; Spielman Andrew I; Seger Rony; Naim Michael^*

来源：Biochimica et Biophysica Acta-General Subjects, 2015, 1850(7): 1375-1388.

DOI：10.1016/j.bbagen.2015.03.015

• Membrane-permeable
• Tastants
• beta(2)AR
• GRK2
• Signaling
• Desensitization

Background: Amphipathic sweet and bitter tastants inhibit purified forms of the protein kinases GRK2, GRK5 and PKA activities. Here we tested whether membrane-permeable tastants may intracellularly interfere with GPCR desensitization at the whole cell context.
Methods: beta(2)AR-transfected cells and cells containing endogenous beta(2)AR were preincubated with membrane-permeable or impermeable tastants and then stimulated with isoproterenol (ISO). cAMP formation, beta(2)AR phosphorylation and beta(2)AR internalization were monitored in response to ISO stimulation. IBMX and H89 inhibitors and GRK2 silencing were used to explore possible roles of PDE, PICA, and GRK2 in the tastants-mediated amplification of cAMP formation and the tastant delay of beta(2)AR phosphorylation and internalization.
Results: Membrane-permeable but not impermeable tastants amplified the ISO-stimulated cAMP formation in a concentration- and time-dependent manner. Without ISO stimulation, amphipathic tastants, except caffeine, had no effect on cAMP formation. The amplification of ISO-stimulated cAMP formation by the amphipathic tastants was not affected by PDE and PICA activities, but was completely abolished by GRK2 silencing. Amphipathic tastants delayed the ISO-induced GRK-mediated phosphorylation of beta(2)ARs and GRK2 silencing abolished it. Further, tastants also delayed the ISO-stimulated beta(2)AR internalization.
Conclusion: Amphipathic tastants significantly amplify beta(2)AR signaling and delay its desensitization via their intracellular inhibition of GRK2.
General Significance: Commonly used amphipathic tastants may potentially affect similar GPCR pathways whose desensitization depends on GRK2's kinase activity. Because GRK2 also modulates phosphorylation of non-receptor components in multiple cellular pathways, these gut-absorbable tastants may permeate into various cells, and potentially affect GRK2-dependent phosphorylation processes in these cells as well.

• 出版日期2015-7