A novel molecular determinant for cAMP-dependent regulation of the frog heart Na+-Ca2+ exchanger.
Na+-Ca2+ exchanger is one of the major sarcolemmal Ca2+ transporters of cardiac myocytes. In frog ventricular myocytes the exchanger is regulated by isoproterenol via a beta-adrenoreceptor/adenylate-cyclase/cAMPdependent signaling pathway providing a molecular mechanism for the relaxant effect of the hormone. Here, we report on the presence of a novel exon of 27-base pair insertion, which generates a nucleotide binding motif (P-loop) in the frog cardiac Na+-Ca2+ exchanger. To examine the functional role of this motif, we constructed a full-length frog heart Na+-Ca2+ exchanger cDNA (fNCX1a) containing this exon. The functional expression of fNCX1a in oocytes showed characteristic voltage dependence, divalent (Ni2+, Cd2+) inhibition, and sensitivity to cAMP in a manner similar to that of native exchanger in frog myocytes. In oocytes expressing the dog heart NCX1 or the frog mutant (DeltafNCX1a) lacking the 9-amino acid exon, cAMP failed to regulate Na+-dependent Ca2+ uptake. We suggest that this motif is responsible for the observed cAMP-dependent functional differences between the frog and the mammalian hearts.
Departments of Pharmacology and Medicine, Georgetown University Medical Center, Washington, DC 20007, USA.
Y M
YM
Shuba
T
T
Iwata
V G
VG
Naidenov
M
M
Oz
K
K
Sandberg
A
A
Kraev
E
E
Carafoli
M
M
Morad
D000262:Adenylyl Cyclases / Q000378:metabolism; D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000818:Animals; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D004285:Dogs; D004594:Electrophysiology; D005091:Exons; D006321:Heart / Q000502:physiology; D007545:Isoproterenol / Q000494:pharmacology; D009206:Myocardium / Q000378:metabolism*; D009865:Oocytes / Q000187:drug effects / Q000502:physiology; D011943:Receptors, Adrenergic, beta / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D019831:Sodium-Calcium Exchanger / Q000235:genetics / Q000378:metabolism*; D013873:Thionucleotides / Q000494:pharmacology; D014982:Xenopus laevis
D000318:Adrenergic beta-Agonists; D011943:Receptors, Adrenergic, beta; D011994:Recombinant Proteins; D019831:Sodium-Calcium Exchanger; D013873:Thionucleotides; C110374:sodium-calcium exchanger 1; C015929:8-((4-chlorophenyl)thio)cyclic-3',5'-AMP; D000242:Cyclic AMP; D000262:Adenylyl Cyclases; D007545:Isoproterenol