Dopamine D3 receptors are down-regulated following heterologous endocytosis by a specific interaction with G protein-coupled receptor-associated sorting protein-1

Abstract

The D3 dopamine receptor is endocytosed through a heterologous mechanism mediated by phorbol esters. Here, we show that following this endocytosis the D3 dopamine receptors fail to recycle and are instead targeted for degradation through an interaction with the G protein-coupled receptor (GPCR)-associated sorting protein-1 (GASP-1). Furthermore, we identified a specific binding motif in the C terminus common to the D 3 and D2 that confers GASP-1 binding. shRNA knockdown of GASP-1 delayed post-endocytic degradation of both the D2 and D 3 dopamine receptors. In addition, mutation of the D2 and D3 receptor C termini to resemble the D4, which does not interact with GASP-1, not only inhibited GASP-1 binding but slowed degradation after endocytosis. Conversely, mutation of the C terminus of the D4 to resemble that of the D2 and D3 facilitated GASP-1 binding and promoted post-endocytic degradation of the mutant D4 receptor. Thus, we have identified a motif that is both necessary and sufficient to promote GASP-1 binding and receptor degradation. In addition, these data demonstrated that GASP-1 can mediate post-endocytic degradation of dopamine receptors that have been endocytosed not only as a consequence of dopamine activation but also as a consequence of activation by phorbol esters. © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.