The KBTBD6/7-DRD2 axis regulates pituitary adenoma sensitivity to dopamine agonist treatment

Pituitary adenoma (PA) is one of the most common intracranial tumors, and approximately 40% of all PAs are prolactinomas. Dopamine agonists (DAs), such as cabergoline (CAB), have been successfully used in the treatment of prolactinomas. The expression of dopamine type 2 receptor (DRD2) determines the therapeutic effect of DAs, but the molecular mechanisms of DRD2 regulation are not fully understood. In this study, we first demonstrated that DRD2 underwent proteasome-mediated degradation. We further employed the yeast two-hybrid system and identified kelch repeat and BTB (POZ) domain containing 7 (KBTBD7), a substrate adaptor for the CUL3-RING ubiquitin (Ub) ligase complex, as a DRD2-interacting protein. KBTBD6/7 directly interacted with, and ubiquitinated DRD2 at five ubiquitination sites (K221, K226, K241, K251, and K258). CAB, a high-affinity DRD2 agonist, induced DRD2 internalization, and cytoplasmic DRD2 was degraded via ubiquitination under the control of KBTBD6/7, the activity of which attenuated CAB-mediated inhibition of the AKT/mTOR pathway. KBTBD7 knockout (KO) mice were generated using the CRISPR-Cas9 technique, in which the static level of DRD2 protein was elevated in the pituitary gland, thalamus, and heart, compared to that of WT mice. Consistently, the expression of KBTBD6/7 was negatively correlated with that of DRD2 in human pituitary tumors. Moreover, KBTBD7 was highly expressed in dopamine-resistant prolactinomas, but at low levels in dopamine-sensitive prolactinomas. Knockdown of KBTBD6/7 sensitized MMQ cells and primary pituitary tumor cells to CAB treatment. Conversely, KBTBD7 overexpression increased CAB resistance of estrogen-induced in situ rat prolactinoma model. Together, our findings have uncovered the novel mechanism of DRD2 protein degradation and shown that the KBTBD6/7-DRD2 axis regulates PA sensitivity to DA treatment. KBTBD6/7 may thus become a promising therapeutic target for pituitary tumors.