FIGURE 7.

Roles of ubiquitination in the life cycle of agonist-stimulated β₂AR. Step 1, within seconds of agonist exposure, β₂ARs stimulate G_s and adenylyl cyclase, increasing cellular cAMP. Step 2, agonist-occupied receptors are also phosphorylated by GRKs on cytoplasmic domain seryl and/or threonyl residues, within seconds to minutes of agonist exposure. Step 3, cytosolic β-arrestin2 (βarr2) translocates to phosphorylated receptors within 1–5 min after agonist treatment. Agonist-dependent β-arrestin ubiquitination (U) occurs immediately upon β-arrestin recruitment and is mediated by Mdm2 that is bound to β-arrestin. β-Arrestin recruitment prevents further G protein coupling and β-arrestin ubiquitination allows it to form signaling and endocytic complexes, facilitating both receptor endocytosis and MAPK signaling. Step 4, β-arrestin conformational changes that occur upon receptor binding allow its interaction with Nedd4, which probably displaces Mdm2 from β-arrestin (5–15 min after agonist treatment). Step 5, by interacting simultaneously with β₂AR, clathrin, and AP-2, β-arrestin2 facilitates β₂AR endocytosis. Step 6, β-arrestin2 is deubiquitinated by an as-yet unidentified process, leading to its disengagement from the receptor complex (10–15 min after agonist treatment). Nedd4 mediates ubiquitination of the endosomally located β₂AR. Step 7, ubiquitinated β₂ARs move on into early endosomes (at >15 min after activation). Step 8, β₂AR ubiquitination persists until about 6 h after agonist stimulation, when β₂ARs move into late endosomal/lysosomal compartments. Step 9, level of ubiquitinated β₂ARs decreases, as ubiquitinated receptors are degraded in lysosomes (6–24 h or more after agonist stimulation). Step 10, from the early endosomes, receptors may take up an alternate path and enter recycling endosomes (<15–30 min after activation), in which β₂ARs become dephosphorylated and perhaps deubiquitinated, and return to the plasma membrane as “naïve receptors” (Step 11).