Mitochondrial ROMK channel is a molecular component of mitoK(ATP).
BACKGROUND
Activation of the mitochondrial ATP-sensitive potassium channel (mitoK(ATP)) has been implicated in the mechanism of cardiac ischemic preconditioning, yet its molecular composition is unknown.


OBJECTIVE
To use an unbiased proteomic analysis of the mitochondrial inner membrane to identify the mitochondrial K(+) channel underlying mitoK(ATP).


RESULTS
Mass spectrometric analysis was used to identify KCNJ1(ROMK) in purified bovine heart mitochondrial inner membrane and ROMK mRNA was confirmed to be present in neonatal rat ventricular myocytes and adult hearts. ROMK2, a short form of the channel, is shown to contain an N-terminal mitochondrial targeting signal, and a full-length epitope-tagged ROMK2 colocalizes with mitochondrial ATP synthase β. The high-affinity ROMK toxin, tertiapin Q, inhibits mitoK(ATP) activity in isolated mitochondria and in digitonin-permeabilized cells. Moreover, short hairpin RNA-mediated knockdown of ROMK inhibits the ATP-sensitive, diazoxide-activated component of mitochondrial thallium uptake. Finally, the heart-derived cell line, H9C2, is protected from cell death stimuli by stable ROMK2 overexpression, whereas knockdown of the native ROMK exacerbates cell death.


CONCLUSIONS
The findings support ROMK as the pore-forming subunit of the cytoprotective mitoK(ATP) channel.
Division of Cardiology, The Johns Hopkins University School of Medicine, Baltimore, MD, USA.
D Brian
DB
Foster
Alice S
AS
Ho
Jasma
J
Rucker
Anders O
AO
Garlid
Ling
L
Chen
Agnieszka
A
Sidor
Keith D
KD
Garlid
Brian
B
O'Rourke
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis; D001514:Bee Venoms / Q000494:pharmacology; D016466:CHO Cells; D002417:Cattle; D006224:Cricetinae; D003412:Cricetulus; D019610:Cytoprotection; D003981:Diazoxide / Q000494:pharmacology; D005786:Gene Expression Regulation; D006801:Humans; D013058:Mass Spectrometry; D008929:Mitochondria, Heart / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D051336:Mitochondrial Membranes / Q000187:drug effects / Q000378:metabolism*; D025261:Mitochondrial Proton-Translocating ATPases / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D009336:Necrosis; D026902:Potassium Channel Blockers / Q000494:pharmacology; D015221:Potassium Channels / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D013793:Thallium / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
D001514:Bee Venoms; C510692:Kcnj1 protein, rat; D026902:Potassium Channel Blockers; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D012333:RNA, Messenger; C117780:mitochondrial K(ATP) channel; C029738:tertiapin; D000255:Adenosine Triphosphate; D013793:Thallium; D025261:Mitochondrial Proton-Translocating ATPases; D003981:Diazoxide