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(Circulation Research. 2005;96:91.)
© 2005 American Heart Association, Inc.

Cellular Biology

Cardiac Sodium-Calcium Exchanger Is Regulated by Allosteric Calcium and Exchanger Inhibitory Peptide at Distinct Sites

Christoph Maack, Anand Ganesan, Agnieszka Sidor, Brian O’Rourke

From the Johns Hopkins University, Institute of Molecular Cardiobiology, Division of Cardiology, Baltimore, Md.

Correspondence to Brian O’Rourke, PhD, Institute of Molecular Cardiobiology, Division of Cardiology, 720 Rutland Ave, 844 Ross Bldg, Baltimore, MD 21205-2195. E-mail bor{at}jhmi.edu

The sarcolemmal Na⁺-Ca²⁺ exchanger (NCX) is the main Ca²⁺ extrusion mechanism in cardiac myocytes and is thus essential for the regulation of Ca²⁺ homeostasis and contractile function. A cytosolic region (f-loop) of the protein mediates regulation of NCX function by intracellular factors including inhibition by exchanger inhibitory peptide (XIP), a 20 amino acid peptide matching the sequence of an autoinhibitory region involved in allosteric regulation of NCX by intracellular Na⁺, Ca²⁺, and phosphatidylinositol-4,5-biphosphate (PIP₂). Previous evidence indicates that the XIP interaction domain can be eliminated by large deletions of the f-loop that also remove activation of NCX by intracellular Ca²⁺. By whole-cell voltage clamping experiments, we demonstrate that deletion of residues 562–679, but not 440– 456, 498–510, or 680–685 of the f-loop selectively eliminates XIP-mediated inhibition of NCX expressed either heterologously (HEK293 and A549 cells) or in guinea pig cardiac myocytes. In contrast, by plotting I_NCX against reverse-mode NCX-mediated Ca²⁺ transients in myocytes, we demonstrate that Ca²⁺-dependent regulation of NCX is preserved in 562–679, but significantly reduced in the other three deletion mutants. The findings indicate that f-loop residues 562–679 may contain the regulatory site for endogenous XIP, but this site is distinct from the Ca²⁺-regulatory domains of the NCX. Because regulation of the NCX by Na⁺ and PIP₂ involves the endogenous XIP region, the 562–679 mutant NCX may be a useful tool to investigate this regulation in the context of the whole cardiac myocyte.

Key Words: cardiac myocytes • adenovirus • mutation

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