Abstract:

A line of Chinese hamster ovary (CHO) cells called CK1.4 was produced by transfection with the gene for the bovine cardiac Na(+)-Ca2+ exchanger. CK1.4 cells stably expressed substantial exchange activity and exchanger protein as shown by immunoprecipitation. Exchange activity was quantified as 45Ca2+ influx that depended on both increasing intracellular Na+ and lowering the concentration of external Na+. Replacing external Na+ with K+ slightly increased 45Ca2+ uptake by CK1.4 cells with basal Na+ and greatly increased 45Ca2+ uptake by Na(+)-loaded cells. Neither exchange activity nor exchanger protein was detected in the nontransfected parental line. By contrast to CK1.4 cells, replacing external Na+ with K+ decreased 45Ca2+ uptake in the nontransfected cells whether or not they were Na+ loaded. Changes in cytosolic free Ca2+ determined with fura-2 were consistent with the 45Ca2+ uptake data. Analysis of poly(A)(+)-RNA by Northern blot confirmed that CK1.4 cells, but not the parental line, expressed the exchanger. Expression of the exchanger was also observed in aortic myocytes and a renal epithelial cell line (LLC-MK2) but not in other lines of renal epithelial cells (MDCK, LLC-PK1) or human dermal fibroblasts. The cardiac exchanger produced substantial 45Ca2+ efflux from CK1.4 cells in response to hormone-evoked release of stored Ca2+. CK1.4 cells are an attractive model for studies of the regulation of the cardiac exchanger because they stably express sufficient exchanger for biochemical and immunological analysis

Notes:

DA - 19930517 IS - 0002-9513 (Print) IS - 0002-9513 (Linking) LA - eng PT - Journal Article PT - Research Support, U.S. Gov't, P.H.S RN - 0 (Calcium Radioisotopes) RN - 0 (Carrier Proteins) RN - 0 (Sodium-Calcium Exchanger) RN - 5ACL011P69 (Ouabain) RN - 9NEZ333N27 (Sodium) RN - AE28F7PNPL (Methionine) RN - RWP5GA015D (Potassium) RN - SY7Q814VUP (Calcium) SB - IM

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