Mutational Analysis of The Er-Exit Code in The C-Terminal Domain of Kir6.2

Mutations in the gene encoding the potassium channel subunit Kir6.2 can cause congenital hyperinsulinism (CHI). CHI mutations severely affect channel function and trafficking in the â-cell membrane. The present study reports a novel mutation, E282K, in the Kir6.2 subunit in a patient diagnosed with CHI. The impact of this mutation on channel trafficking is assessed by immunocytochemical studies, and channel function is determined by site-directed mutagenesis. Heterologous expression of Kir6.2 clearly showed that mutations in the DXE code cause the channel to be retained in the ER. Further mutagenic studies revealed that E282K could be a part of the DXE exit code. This raised the possibility that KATP channels may assemble with Sec 23/Sec 24 proteins and exit the ER in COPII vesicles in a process that requires a dominant negative form of Sar1-GTPase (one that was unable to hydrolyse GTP) and prevented the ER exit of wild-type channels. This is due to the mutation in the C-terminal region which prevented the ER exit code. In conclusion, KATP channels use the DXE motif to export newly synthesized proteins from the ER. The functional importance of this code is demonstrated by the severity of CHI, which appears to be due to a genetic mutation in the DXE motif. Thus, the present study provides novel information that Kir6.2 is synthesized in the ER and exported with the aid of the DXE motif in its C-terminal region.