title: Evidence for proteasomal degradation of Kv1.5 channel protein.
abstract: BACKGROUND
  The voltage-gated potassium channel Kv1.5 plays a critical role in the maintenance of the membrane potential. While protein degradation is one of the major mechanisms for the regulation of channel functions, little is known on the degradation mechanism of Kv1.5.
  
  
  RESULTS
  Kv1.5 was expressed in COS cells and its degradation, intracellular localization, and channel activities were assessed by pulse-chase analysis, immunofluorescence, and patch clamp techniques, respectively. Expressed Kv1.5 had a half-life time of approximately 6.7 h, which was prolonged by the proteasome inhibitors of MG132, ALLN, proteasomal inhibitor 1, or lactacystine, but not by a lysosomal inhibitor chloroquine. MG132 increased the protein level of Kv1.5, as well as the level of its ubiquitinated form in a dose-dependent manner. Similar effects of MG132 on endogenous Kv1.5 were seen in cultured rat atrial cells. Within a cell, Kv1.5 was mainly localized in both the endoplasmic reticulum and Golgi apparatus. MG132 increased the immunoreactivity of Kv1.5 in these compartments and also increased Ik(ur) currents through the cell-surface Kv1.5. Pretreatment with either brefeldin A or colchicine abolished MG132-induced increase in Ik(ur) currents.
  
  
  CONCLUSIONS
  Kv1.5 is degraded by the proteasome. The inhibition of the proteasome increased Ik(ur) currents secondary to stabilization of the channel protein in the endoplasmic reticulum/Golgi apparatus.
authors.affiliation: Department of Cardiovascular Medicine, Tottori University Faculty of Medicine, Japan.
authors.forename: Masaru
authors.initials: M
authors.lastname: Kato
authors.forename: Kazuyoshi
authors.initials: K
authors.lastname: Ogura
authors.forename: Junichiro
authors.initials: J
authors.lastname: Miake
authors.forename: Norihito
authors.initials: N
authors.lastname: Sasaki
authors.forename: Shin-Ichi
authors.initials: S
authors.lastname: Taniguchi
authors.forename: Osamu
authors.initials: O
authors.lastname: Igawa
authors.forename: Akio
authors.initials: A
authors.lastname: Yoshida
authors.forename: Yoshiko
authors.initials: Y
authors.lastname: Hoshikawa
authors.forename: Mitsunobu
authors.initials: M
authors.lastname: Murata
authors.forename: Eiji
authors.initials: E
authors.lastname: Nanba
authors.forename: Yasutaka
authors.initials: Y
authors.lastname: Kurata
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Kawata
authors.forename: Haruaki
authors.initials: H
authors.lastname: Ninomiya
authors.forename: Takayuki
authors.initials: T
authors.lastname: Morisaki
authors.forename: Masafumi
authors.initials: M
authors.lastname: Kitakaze
authors.forename: Ichiro
authors.initials: I
authors.lastname: Hisatome
mesh_terms: D000818:Animals; D019556:COS Cells; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004553:Electric Conductivity; D004721:Endoplasmic Reticulum / Q000737:chemistry; D006056:Golgi Apparatus / Q000737:chemistry; D051666:Kv1.5 Potassium Channel; D007976:Leupeptins / Q000494:pharmacology; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism; D009842:Oligopeptides; D018408:Patch-Clamp Techniques; D010455:Peptides / Q000235:genetics; D024642:Potassium Channels, Voltage-Gated / Q000032:analysis / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000032:analysis; D014452:Ubiquitins / Q000378:metabolism
chemical_list: D015853:Cysteine Proteinase Inhibitors; C496983:Kcna5 protein, rat; D051666:Kv1.5 Potassium Channel; D007976:Leupeptins; D009842:Oligopeptides; D010455:Peptides; D024642:Potassium Channels, Voltage-Gated; D011993:Recombinant Fusion Proteins; D014452:Ubiquitins; C085706:FLAG peptide; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde