Physiological functions of the regulatory domains of the cardiac Na(+)/Ca(2+) exchanger NCX1.
Physiological functions of the intracellular regulatory domains of the Na(+)/Ca(2+) exchanger NCX1 were studied by examining Ca(2+) handling in CCL39 cells expressing a low-affinity Ca(2+) regulatory site mutant (D447V/D498I), an exchanger inhibitory peptide (XIP) region mutant displaying no Na(+) inactivation (XIP-4YW), or a mutant lacking most of the central cytoplasmic loop (Delta246-672). We found that D447V/D498I was unable to efficiently extrude Ca(2+) from the cytoplasm, particularly during a small rise in intracellular Ca(2+) concentration induced by the physiological agonist alpha-thrombin or thapsigargin. The same mutant took up Ca(2+) much less efficiently than the wild-type NCX1 in Na(+)-free medium when transfectants were not loaded with Na(+), although it appeared to take up Ca(2+) normally in transfectants preloaded with Na(+). XIP-4YW and, to a lesser extent, Delta246-672, but not NCX1 and D447V/D498I, markedly accelerated the loss of viability of Na(+)-loaded transfectants. Furthermore, XIP-4YW was not activated by phorbol ester, whereas XIP-4YW and D447V/D498I were resistant to inhibition by ATP depletion. The results suggest that these regulatory domains play important roles in the physiological and pathological Ca(2+) handling by NCX1, as well as in the regulation of NCX1 by protein kinase C or ATP depletion.
Department of Molecular Physiology, National Cardiovascular Center Research Institute, Suita, Osaka 565-8565, Japan.
Y
Y
Pan
T
T
Iwamoto
A
A
Uehara
T Y
TY
Nakamura
I
I
Imanaga
M
M
Shigekawa
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000494:pharmacology / Q000502:physiology; D006224:Cricetinae; D003412:Cricetulus; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D008168:Lung / Q000187:drug effects / Q000378:metabolism; D011493:Protein Kinase C / Q000187:drug effects / Q000378:metabolism*; D012964:Sodium / Q000494:pharmacology / Q000502:physiology; D019831:Sodium-Calcium Exchanger / Q000187:drug effects / Q000378:metabolism*
D019831:Sodium-Calcium Exchanger; C110374:sodium-calcium exchanger 1; D000255:Adenosine Triphosphate; D012964:Sodium; D011493:Protein Kinase C; D002118:Calcium