APLP1 and Rab5A interact with the II-III loop of the voltage-gated Ca-channel Ca(v)2.3 and modulate its internalization differently.

Background: Voltage gated calcium channels (VGCCs) regulate cellular activity in response to membrane depolarization by altering calcium homeostasis. Because calcium is the most versatile second messenger, regulation of the amount of VGCCs at the plasma membrane is highly critical for several essential cellular processes. Among the different types of VGCCs, the Cav2.3 calcium channel and its regulation mechanisms are least understood due to Cav2.3’s resistance to most pharmacological agents. Methods: In order to study regulation and surface expression of Cav2.3, a yeast two hybrid (Y2H) screen with the II-III loop of human Cav2.3 as bait, was performed. APLP1, a member of the APP gene family and Rab5A, an endocytotic catalyst were identified as putative interaction partners. The interaction were confirmed by immunoprecipitation. To study the functional importance of the interaction, patch-clamp recordings in Cav2.3 stably transfected HEK-293 cells (2C6) and surface biotin endocytosis assays were performed. Results: We are able to show that the II-III loop of the Cav2.3 calcium channel binds APLP1 and that this binding promotes internalization of the channel. In addition, Rab5A also binds to the same loop of the channel and exerts an inhibitory effect on APLP1 mediated channel internalization. Conclusions: This study identifies a regulation mechanism of Cav2.3’s surface expression, which implicates APLP1 as a regulator of calcium homeostasis. Thus APLP1 may play a crucial role in neuropathological mechanisms, which involve modulation of surface expression of voltage-gated Ca2+ channels.