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Background: Desensitization of G protein–coupled receptors (GPCRs) refers to the attenuation of receptor responsiveness by prolonged or intermittent exposure to agonists. The binding of β-arrestin to the cytoplasmic cavity of the phosphorylated receptor, which competes with the G protein, has been widely accepted as an extensive model for explaining GPCRs desensitization. However, studies on various GPCRs, including dopamine D2-like receptors (D2R, D3R, D4R), have suggested the existence of other desensitization mechanisms. The present study employed D2R/D3R variants with different desensitization properties and utilized loss-of-function approaches to uncover the mechanisms underlying GPCRs homologous desensitization, focusing on the signaling cascade that regulates the ubiquitination of AKT. Results: AKT undergoes K8/14 ubiquitination by TRAF6, which occurs in the nucleus and promotes its membrane recruitment, phosphorylation and activation under receptor desensitization conditions. The nuclear entry of TRAF6 relies on the presence of the importin complex. Src regulates the nuclear entry of TRAF6 by mediating the interaction between TRAF6 and importin β1. Ubiquitinated AKT translocates to the plasma membrane where it associates with Mdm2 to phosphorylate it at the S166 and S186 residues. Thereafter, phosphorylated Mdm2 is recruited to the nucleus, resulting in the deubiquitination of β-Arr2. The deubiquitinated β-Arr2 then forms a complex with Gβγ, which serves as a biomarker for GPCRs desensitization. Like in D3R, ubiquitination of AKT is also involved in the desensitization of β2 adrenoceptors. Conclusion: Our study proposed that the property of a receptor that causes a change in the subcellular localization of TRAF6 from the cytoplasm to the nucleus to mediate AKT ubiquitination could initiate the desensitization of GPCRs. [ABSTRACT FROM AUTHOR]

The insulin and dopaminergic systems in the brain are associated with schizophrenia and Parkinson's disease with respect to etiology and treatment. The present study investigated the crosstalk between the insulin receptor (IR) and dopamine receptor and found that insulin stimulation selectively inhibits signaling of D3R in a PKCβII‐dependent manner. Upon insulin stimulation, E3 ligase enzyme Mdm2 moves out of the nucleus to ubiquitinate PKCβII. Subsequently, ubiquitinated PKCβII translocates to the cell membrane and interacts with D3R in a phosphorylation‐dependent manner at S229/257, resulting in the attenuation of D3R signaling and initiating clathrin‐mediated endocytosis and downregulation. Considering that both IR and D3R are closely related to some neuropsychosis, this study could provide new molecular insight into the etiology of the disorder. [ABSTRACT FROM AUTHOR]

A class-A GPCR dopamine D2 receptor (D2R) plays a critical role in the proper functioning of neuronal circuits through the downstream activation of both G-protein- and β-arrestin-dependent signaling pathways. Understanding the signaling pathways downstream of D2R is critical for developing effective therapies with which to treat dopamine (DA)-related disorders such as Parkinson's disease and schizophrenia. Extensive studies have focused on the regulation of D2R-mediated extracellular-signal-regulated kinase (ERK) 1/2 signaling; however, the manner in which ERKs are activated upon the stimulation of a specific signaling pathway of D2R remains unclear. The present study conducted a variety of experimental techniques, including loss-of-function experiments, site-directed mutagenesis, and the determination of protein interactions, in order to investigate the mechanisms underlying β-arrestin-biased signaling-pathway-mediated ERK activation. We found that the stimulation of the D2R β-arrestin signaling pathway caused Mdm2, an E3 ubiquitin ligase, to move from the nucleus to the cytoplasm and interact with tyrosine phosphorylated G-protein-coupled receptor kinase 2 (GRK2), which was facilitated by Src, a non-receptor tyrosine kinase. This interaction led to the ubiquitination of GRK2, which then moved to the plasma membrane and interacted with activated D2R, followed by the phosphorylation of D2R as well as the mediation of ERK activation. In conclusion, Mdm2-mediated GRK2 ubiquitination, which is selectively triggered by the stimulation of the D2R β-arrestin signaling pathway, is necessary for GRK2 membrane translocation and its interaction with D2R, which in turn mediates downstream ERK signaling. This study is primarily novel and provides essential information with which to better understand the detailed mechanisms of D2R-dependent signaling. [ABSTRACT FROM AUTHOR]