␤-Arrestins are multifunctional adaptors that mediate the desensitization, internalization, and some signaling functions of seventransmembrane receptors (7TMRs). Agonist-stimulated ubiquitination of ␤-arrestin2 mediated by the E3 ubiquitin ligase Mdm2 is critical for rapid ␤2-adrenergic receptor (␤2AR) internalization. We now report the discovery that the deubiquitinating enzyme ubiquitin-specific protease 33 (USP33) binds ␤-arrestin2 and leads to the deubiquitination of ␤-arrestins. USP33 and Mdm2 function reciprocally and favor respectively the stability or lability of the receptor ␤-arrestin complex, thus regulating the longevity and subcellular localization of receptor signalosomes. Receptors such as the ␤2AR, previously shown to form loose complexes with ␤-arrestin (''class A'') promote a ␤-arrestin conformation conducive for binding to the deubiquitinase, whereas the vasopressin V2R, which forms tight ␤-arrestin complexes (''class B''), promotes a distinct ␤-arrestin conformation that favors dissociation of the enzyme. Thus, USP33-␤-arrestin interaction is a key regulatory step in 7TMR trafficking and signal transmission from the activated receptors to downstream effectors.endocytosis ͉ G protein-coupled receptors ͉ ubiquitination ͉ phosphorylation ͉ ERK1/2