PT  - JOURNAL ARTICLE
AU  - Zara Y Weinberg
AU  - Amanda S Zajac
AU  - Tiffany Phan
AU  - Daniel J Shiwarski
AU  - Manojkumar A Puthenveedu
TI  - Sequence-specific regulation of endocytic lifetimes modulates arrestin-mediated signaling at the μ opioid receptor
AID  - 10.1124/mol.116.106633
DP  - 2017 Jan 01
TA  - Molecular Pharmacology
PG  - mol.116.106633
4099  - http://molpharm.aspetjournals.org/content/early/2017/02/02/mol.116.106633.short
4100  - http://molpharm.aspetjournals.org/content/early/2017/02/02/mol.116.106633.full
AB  - Functional selectivity at the μ opioid receptor (μR), a prototypical GPCR that is a physiologically relevant target for endogenous opioid neurotransmitters and analgesics, has been a major focus for drug discovery in the recent past. The cellular mechanisms that mediate functional selectivity, however, are still being fully elucidated. The present work tested the hypothesis that lifetimes of agonist-induced receptor-arrestin clusters at the cell surface controls the magnitude of arrestin signaling, and therefore functional selectivity, on μR. We show that endomorphin-2 (EM2), an arrestin-biased ligand for μR, lengthens surface lifetimes of receptor-arrestin clusters significantly compared to morphine. The lengthening of lifetimes required two specific leucines on the C-terminal tail of μR. Mutation of these leucines to alanines decreased arrestin-mediated signaling by EM2 without affecting G-protein signaling, suggesting that lengthened endocytic lifetimes were required for arrestin-biased signaling by EM2. Lengthening surface lifetime by pharmacologically slowing endocytosis was sufficient to increase arrestin-mediated signaling by both EM2 and the clinically relevant agonist morphine. Our findings show that distinct ligands can leverage specific sequence elements on the μR to regulate receptor endocytic lifetimes and the magnitude of arrestin-mediated signaling, and implicate these sequences as important determinants of functional selectivity in the opioid system.