Intracellular retention of the two isoforms of the D-2 dopamine receptor promotes endoplasmic reticulum disruption

The dopamine D-2 receptor exists as a long (D-2a) and a short (D-2b) isofor m generated by alternative splicing of the corresponding transcript, which modifies the length of the third cytoplasmic loop implicated in heterotrime ric G-protein-coupling. Anatomical data suggested that this segment regulat es the intracellular traffic and localization of the receptor. To directly address this question we used a combination of tagging procedures and immun ocytochemical techniques to detect each of the two D-2 receptor isoforms. S urprisingly, most of the newly synthesized receptors accumulate in large in tracellular compartments, the plasma membrane being only weakly labeled, wi thout significant difference between the two receptor isoforms. Double labe ling experiments showed that this localization corresponded neither to endo somal compartments nor to the Golgi apparatus. The D-2 receptor is mostly r etained in the endoplasmic reticulum (ER), the long isoform more efficientl y than the short one. It is accompanied by a striking vacuolization of the ER, roughly proportional to the expression levels of the two receptor isofo rms. This phenomenon is partly overcome by treatment with pertussis toxin. In addition, an intrinsic activity of the D-2 receptor isoforms is revealed by [S-35]- GTP gammaS binding and cAMP assay, which suggested that express ion of weakly but constitutively active D-2 receptors promotes activation o f heterotrimeric G protein inside the secretory pathway. This mechanism may participate in the regulation of the cellular traffic of the D-2 receptors isoforms.