Endocrine Abstracts

The actin binding protein filamin A (FLNA) is required for somatostatin receptor 2 (SST2) and dopamine receptor 2 (DRD2) expression, intracellular localization and signaling in GH- and PRL-secreting pituitary tumors, respectively, playing a role in tumor responsiveness to somatostatin analogs and dopaminergic drugs. FLNA functions are tightly regulated by several mechanisms, including FLNA phosphorylation. It has recently been shown that in GH-secreting pituitary tumors FLNA phosphorylation on Ser2152 switches FLNA function from a scaffold that allows SST2 signal transduction, to a signal termination protein that hampers all SST2 antitumoral effects. Aims of the present study were to evaluate in PRL- and ACTH-secreting pituitary tumor cell lines MMQ and AtT-20, endogenously expressing DRD2: 1) the effects of cAMP pathway activation and DRD2 agonist on FLNA phosphorylation; 2) the impact of FLNA phosphorylation on DRD2 intracellular signal transduction.

We found that forskolin increased (+2.87 ± onefold, P < 0.05 in MMQ; +1.92 ± 0.8 fold, P < 0.05 in AtT-20), and DRD2 agonist BIM53097 reduced (–77.3 ± 2%, P < 0.001 in MMQ; –52.7 ± 3%, P < 0.05 in AtT-20), FLNA phosphorylation on Ser2152. The overexpression of a phosphomimetic (S2152D) FLNA mutant in both cell lines completely prevented DRD2 antiproliferative effects, that were comparable in cells transfected with empty vector, wild type FLNA as well as phosphodeficient FLNA mutant (S2152A) (–20.6 ± 5%, P < 0.001 in MMQ; –36.6 ± 12%, P < 0.01 in AtT-20). Accordingly, S2152D FLNA expression abolished the expected ability of BIM53097 to increase (in MMQ) or decrease (in AtT20) ERK phosphorylation, an effect that was maintained in S2152A FLNA expressing cells (+178 ± 65%, P < 0.05 in MMQ; –55 ± 13%, P < 0.01 in AtT-20).

In addition, the inhibitory effects of DRD2 on PRL secretion (–19 ± 1%, P < 0.05 in MMQ expressing S2152A FLNA) were completely lost in S2152D FLNA transfected cells. In conclusion, our data demonstrated that cAMP pathway and DRD2 agonist regulated FLNA activity by increasing or decreasing, respectively, FLNA phosphorylation on Ser2152. Moreover, we found that FLNA phosphorylation prevented DRD2 signaling in PRL- and ACTH-secreting tumoral pituitary cell lines, suggesting that this posttranslational FLNA modification might represent a new GPCRs regulatory mechanism. In pituitary tumors expressing DRD2, modulation of P-FLNA might suggest new pharmacological strategies to overcome drug resistance, and P-FLNA might represent a new biomarker for tumor responsiveness to dopaminergic drugs.