Kchip1 In In Kv4.3 Membrane Transport

KChIP are several NCS proteins that were screened out with N termini of Kv4 by means of yeast two-hybrid system. Increasing data show that physiology interactions between KChIP and Kv4.2, which is a member of Kv4 subfamily, exist in mammal. The interaction between KChlPl and Kv4.3 in mammal, however, has not been demonstrated.In this research, the interaction between KChlPl and Kv4.3 in mammal was demonstrated, since the co-immunoprecipitation of KChlPl and Kv4.3 was found in rat brain membrane fraction.To investigate the biological meaning of the interaction between KChlPl and Kv4.3, KChlPl and Kv4.3L, which is the longer variant Kv4.3, were cloned. And the cloned KChIPl-pcDNA3.1(+) and Kv4.3L-pCMV-Tag4A mammalian expression vectors were transfected COS-7 cells, then their sub-cellular localization was observed by means of immunofluorescence staining. It showed that KChlPl and Kv4.3L were co-localized on the plasma membrane in the co-transfected cells, while KChlPl or Kv4.3L diffused the cytoplasm in the cells transfected the either plasmids alone, respectively. This result indicated that KChlPl can facilitate trafficking of Kv4.3L to plasma membrane.To further study the effect of KChlPl on the trafficking of Kv4.3L tothe plasma membrane, we truncated Kv4.3L with the N terminal 2~39aa which mediate the interaction with KChlPl, and the truncated mutant was transfected COS-7 cells. It showed that this mutant Kv4.3L was localized on the plasma membrane when expressed alone, unlike the wild-type Kv4.3L diffused the cytoplasm when expressed alone. But when fused the N terminal 39aa to EGFP, the fusion protein, NKv4.3-EGFP, distributed the whole cell. And when KChlPl and Kv4.3L were fused, KChIPl-Kv4.3 could express on the plasma membrane.All our results indicated that N terminal 39aa may act as a signal inhibiting the localization of Kv4.3L on plasma membrane. And this inhibiting signal can be eliminated when the fragment interacts or fuses with KChlPl. Therefore, Kv4.3L can be trafficked to membrane efficiently with co-expressing of KChlPl.