Identification of 30 kDa calsequestrin-binding protein, which regulates calcium release from sarcoplasmic reticulum of rabbit skeletal muscle

Abstract

In a previous study [Yamaguchi, Kawasaki and Kasai (1995) Biochem. Biophys. Res. Commun. 210, 648-653], we showed that the stilbene derivative 4,4'-di-isothiocyanostilbene-2,2'-disulphonic acid activates the Ca2+ channel in the sarcoplasmic reticulum (SR) in rabbit skeletal muscle, and it does not bind to the channel protein itself but to the SR 30 kDa protein. Furthermore, the 30 kDa protein was shown to bind to calsequestrin (CSQ), which is one of the regulators of the Ca2+ release channel in the SR. In the present study, we determined the partial amino acid sequence of the CSQ-binding 30 kDa protein and, consequently, this protein was proved to be highly similar to ADP/ATP translocase (AAT) expressed in the mitochondria in a variety of cells. By Western-blotting analysis, the CSQ-binding 30 kDa protein was recognized by the antibody raised against bovine cardiac AAT and, furthermore, depolarization-induced Ca2+ release monitored in the rabbit skeletal muscle triads was significantly activated by the antibody. As a result of cloning and sequencing of the cDNA encoding AAT of the rabbit skeletal muscle, the amino acid sequence was found to be the same as that of the CSQ-binding 30 kDa protein determined above. Furthermore, the expressed product of the cDNA encoding AAT in Escherichia coli was proved to bind to CSQ. These results suggest that AAT itself is expressed in the rabbit skeletal muscle SR and regulates the Ca2+ release from the SR; that is, excitation-contraction coupling of the skeletal muscle cell.

Early impairment of calcium handling and altered expression of junctin in hearts of mice overexpressing the β 1 − adrenergic receptor

“…The physiological functions of these proteins are still largely unknown. Their role in human heart failure has not yet been characterized, but data from in vitro studies suggest a role for junctin in cardiomyocyte calcium handling (13,35) and junctin-transgenic mice display structural alterations of cardiac calcium release units (36).…”

Generation, Control, and Processing of Cellular Calcium Signals

“…It has been proposed that the significant structural changes observed after binding of Ca 2+ to calsequestrin (increase of α-helical content, decrease of hydophobicity [Ostwald and MacLennan, 1974]) affect the ryanodine receptor Ca 2+ -channel, stimulating the release of Ca 2+ from the SR lumen (Szegedi et al, 1999). The functional connection between calsequestrin and the receptor has been ascribed tentatively to triadin, a 95-kDa protein that is thought to bind the ryanodine receptor (Caswell et al, 1991;Guo and Campbell, 1995;MacPherson and Campbell, 1993), and junctin, a 26-kDa protein with sequence homology to triadin (Yamaguchi and Kasai, 1998). Both proteins are anchored to the SR membrane by a single transmembrane domain and bind calsequestrin to their luminal domains, probably through regions rich in basic amino acids.…”

Aldolase potentiates DIDS activation of the ryanodine receptor in rabbit skeletal sarcoplasmic reticulum

“…The effects of DIDS on RyRs make this compound a potentially useful probe to elucidate the mechanism of RyR function. Two studies have already used this compound to probe its interaction with the RyR1 and two RyR1-associated proteins, calmodulin [14] and a 30 kDa protein [15][16][17][18]. The 30 kDa protein was identified as a calsequestrin-binding protein in rabbit skeletal SR and was postulated to be a receptor for DIDS, implying that DIDS is bound to RyR1 via the 30 kDa protein in rabbit skeletal muscle [17].…”

“…Two studies have already used this compound to probe its interaction with the RyR1 and two RyR1-associated proteins, calmodulin [14] and a 30 kDa protein [15][16][17][18]. The 30 kDa protein was identified as a calsequestrin-binding protein in rabbit skeletal SR and was postulated to be a receptor for DIDS, implying that DIDS is bound to RyR1 via the 30 kDa protein in rabbit skeletal muscle [17]. On the other hand, the direct DIDS binding to RyR and covalent modification of RyR by DIDS have been postulated to be the underlying molecular mechanisms for DIDS activation of RyRs [9][10][11][12].…”