Molecular Cloning and Characterization of CALP/KChIP4, a Novel EF-hand Protein Interacting with Presenilin 2 and Voltage-gated Potassium Channel Subunit Kv4

Yuichi Morohashi,Noriyuki Hatano,Susumu Ohya,Rie Takikawa,Tomonari Watabiki,Nobumasa Takasugi,Yuji Imaizumi,Taisuke Tomita,Takeshi Iwatsubo

doi:10.1074/jbc.m200897200

Abstract

Presenilin (PS) genes linked to early-onset familial Alzheimer's disease encode polytopic membrane proteins that are presumed to constitute the catalytic subunit of gamma-secretase, forming a high molecular weight complex with other proteins. During our attempts to identify binding partners of PS2, we cloned CALP (calsenilin-like protein)/KChIP4, a novel member of calsenilin/KChIP protein family that interacts with the C-terminal region of PS. Upon co-expression in cultured cells, CALP was directly bound to and co-localized with PS2 in endoplasmic reticulum. Overexpression of CALP did not affect the metabolism or stability of PS complex, and gamma-cleavage of betaAPP or Notch site 3 cleavage was not altered. However, co-expression of CALP and a voltage-gated potassium channel subunit Kv4.2 reconstituted the features of A-type K(+) currents and CALP directly bound Kv4.2, indicating that CALP functions as KChIPs that are known as components of native Kv4 channel complex. Taken together, CALP/KChIP4 is a novel EF-hand protein interacting with PS as well as with Kv4 that may modulate functions of a subset of membrane proteins in brain.

Highlights

Presenilin (PS) genes linked to early-onset familial Alzheimer’s disease encode polytopic membrane proteins that are presumed to constitute the catalytic subunit of ␥-secretase, forming a high molecular weight complex with other proteins
Multiple sequence alignment and BLAST analysis indicated that CALP had high amino acid identity at its C-terminal region with KChIP2 (79.6%) and calsenilin/KChIP3 (77.6%), whereas its N terminus was very divergent from any KChIPs
We cloned a cDNA encoding mouse CALP from a mouse brain cDNA library by the 5Ј- and 3Ј-RACE method, and we autoradiogram of glutathione S-transferase (GST)-fusioned full-length- and EF-hand mutant (EFmt; Asp and Gly residues in EF-hand motifs being replaced with Ala, see under “Results”) CALP, showing calcium binding through the EF-hand motifs

Summary

Introduction

Presenilin (PS) genes linked to early-onset familial Alzheimer’s disease encode polytopic membrane proteins that are presumed to constitute the catalytic subunit of ␥-secretase, forming a high molecular weight complex with other proteins. Co-expression of CALP and a voltagegated potassium channel subunit Kv4.2 reconstituted the features of A-type K؉ currents and CALP directly bound Kv4.2, indicating that CALP functions as KChIPs that are known as components of native Kv4 channel complex. CALP/KChIP4 is a novel EFhand protein interacting with PS as well as with Kv4 that may modulate functions of a subset of membrane proteins in brain. Inherited as an autosomal dominant trait, and mutations in three different genes have far been linked to early-onset autosomal dominant forms of familial AD (FAD). PS1 and PS2 genes encode polytopic integral membrane proteins that are predominantly localized in intracellular membranes and span the membrane six to eight times

Methods

Results

Conclusion