Inhibition of Hsp70 Enhances A-Type Kv4 Current by Reducing Degradation of Auxiliary KChP4A

Abstract

We have previously shown that KChIP4s reduces surface expression of Kv4 through its N-terminal KID (Kv4 inhibitory domain) that causes endoplasmic reticulum retention of the channel complex. In this study we found that treatment of HEK 293 cells expressing KChIP4a with cycloheximide that inhibits protein synthesis causes a significant degradation of KChIP4a. This accelerated degradation of KChIP4a was reversed by application of a proteasome inhibitor MG132, indicating the degradation of KChIP4a proteins through proteasome pathway. Functional dissection revealed a key domain consisting of eight hydrophobic and aliphatic residues in the N-terminus that is critical for degradation. Using mass spectrum analysis and co-immunoprecipitation assay, we further identified Hsp70 protein (heat shock protein 70) that can specifically interact with auxiliary KChIP4a. Inhibition of Hsp70 function by inhibitors Pifithrin-μ and VER-155008 can recover the reduction of surface Kv4 channels induced by KChIP4a, suggesting that Hsp70 is necessary for degradation of KChIP4a. In hippocampal neurons, inhibition of Hsp70 resulted in an increase of A-type current, suggesting a role of Hsp70 in functional Kv4 channel complexes. Further investigations of interactions between Hsp70 and KChIP4a and in vivo effects of Hsp70 inhibition on neurological functions in animal models are currently underway.