Modulation of D₃R Splicing, Signaling, and Expression by D₁R through PKA→PTB Phosphorylation

The D₁R and D₃R receptors functionally and synergistically interact in striatonigral neurons. Dopaminergic denervation turns this interaction antagonistic, which is correlated with a decrement in D₃nf isoform and an increment in D₃R membranal expression. The mechanisms of such changes in D₃R are attributed to the dysregulation of the expression of their isoforms. The cause and mechanism of this phenomenon remain unknown. Dopaminergic denervation produces a decrement in D₁R and PKA activity; we propose that the lack of phosphorylation of PTB (regulator of alternative splicing) by PKA produces the dysregulation of D₃R splicing and changes D₃R functionality. By using in silico analysis, we found that D₃R mRNA has motifs for PTB binding and, by RIP, co-precipitates with PTB. Moreover, D₁R activation via PKA promotes PTB phosphorylation. Acute and 5-day D₁R blockade decreases the expression of D₃nf mRNA. The 5-day treatment reduces D₃R, D₃nf, and PTB protein in the cytoplasm and increases D₃R in the membrane and PTB in the nucleus. Finally, the blockade of D₁R mimics the effect of dopaminergic denervation in D₁R and D₃R signaling. Thus, our data indicate that through PKA→PTB, D₁R modulates D₃R splicing, expression, and signaling, which are altered during D₁R blockade or the lack of stimulation in dopaminergic denervation.