Dopamine D₄ receptor ubiquitination

Ubiquitination is a post-translational modification that targets proteins for degradation but can also regulate other cellular processes such as endocytosis, trafficking and DNA repair. We investigate ubiquitination of the dopamine D₄ receptor (D₄R) which belongs to the superfamily of G protein-coupled receptors (GPCR). Several polymorphic variants of the D₄R exist, which differ in the number of 16-amino acid repeats in the third intracellular loop (IC3) of the receptor. The functional role of this polymorphic region is not known but persons with the seven-repeat allele show a predisposition to develop attention deficit hyperactivity disorder (ADHD). We identified a protein, KLHL12, which specifically interacts with this polymorphic region and enhances ubiquitination of the D₄R. We have tested the influence of KLHL12 on the ubiquitination of the most common D₄R polymorphic variants and found that KLHL12 strongly promotes ubiquitination of the two- and four-repeat variant but has hardly any effect on ubiquitination of the seven-repeat D₄R. This suggests that differential ubiquitination of the D₄R may have functional implications. Moreover, we were able to demonstrate that KLHL12-mediated D₄R ubiquitination does not lead to receptor degradation. Next, we aimed to identify specific residues in the sequence of D₄R which undergo ubiquitination and observed that the lysine-less receptor mutant is still ubiquitinated. Subsequently, we have tested the hypothesis whether KLHL12 could promote ubiquitination on non-lysine residues of the D₄R. The importance of the cysteine and serine/threonine residues in the ubiquitination process of the receptor was examined and the obtained results confirmed that D₄R can be ubiquitinated on non-lysine residues. In this review we summarize our data on D₄R ubiquitination and put this in the light of other GPCR ubiquitination studies.