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Overexpression and function of small conductance Ca 2+ ‐sensitive K + channel (SK Ca ) isoform 3.1 in HL‐1 myocyte mitochondria (648.12)
Author(s) -
Yang Meiying,
Camara Amadou,
Bienengraeber Martin,
Kwok WaiMeng,
Stowe David
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.648.12
Subject(s) - mitochondrion , myocyte , microbiology and biotechnology , chemistry , transfection , membrane potential , biology , biochemistry , gene
SK Ca agonists protect against cardiac ischemia and reperfusion injury. SK Ca channels are present in cell membranes of vascular and cardiac atrial cells, but not in cell membranes of cardiac ventricular cells. Of the 3 known SK Ca isoforms, we found SK Ca isoform 3 of splice variant 1 (SK Ca 3.1) in the inner mitochondrial (m) membrane of guinea pig ventricular cells. To determine its mitochondrial targeting sequence and function, we cloned full length (FL) and truncated SK Ca 3.1 cDNA for overexpression in HL‐1 myocytes. SK Ca 3.1 localization was examined by immunocytochemistry and western blotting and channel function was evaluated by K + uptake into HL‐1 myocyte mitochondria with the K + ‐selective fluorescent probe PBFI AM. We found that SK3 FL ‐EGFP, N‐terminus truncated SK3.1 (SK3 Δ1‐277 ‐EGFP), but not C‐terminus (includes calmodulin binding motif) truncated SK3.1 (SK3 Δ626‐720 ‐EGFP), localized into mitochondria when transfected into HL‐1 cells. Overexpression of SK3 FL ‐EGFP and SK3 Δ1‐277 ‐EGFP, but not SK3 Δ626‐720 ‐EGFP, increased the rate of K + uptake in HL‐1 cell mitochondria in response to added KCl and CaCl 2 . K + uptake by overexpressed SK3 FL ‐EGFP was sensitive to the SK Ca blocker apamin. Our results show that mSK Ca 3.1 requires its C‐terminus not only for mitochondrial targeting but also for Ca 2+ ‐stimulated K + uptake into mitochondria. Grant Funding Source : R01 HL 089514