Differential regulation of types-1 and -3 inositol trisphosphate receptors by cytosolic Ca2

• Published in: Biochemical journal Vol. 328 ( Pt 3); no. 3; pp. 785 - 793
• Main Authors: Cardy, T J, Traynor, D, Taylor, C W
• Format: Journal Article
• Language: English
• Published: England 15-12-1997
• Subjects: Amino Acid Sequence; Animals; Baculoviridae - genetics; Binding Sites; Binding, Competitive; Blotting, Western; Calcium - metabolism; Calcium - pharmacology; Calcium Channels - classification; Calcium Channels - genetics; Calcium Channels - metabolism; Cell Line; Cytosol - metabolism; Glycosylation; Inositol 1,4,5-Trisphosphate - metabolism; Inositol 1,4,5-Trisphosphate Receptors; Membrane Proteins - metabolism; Molecular Sequence Data; Protein Conformation; Rats; Receptors, Cytoplasmic and Nuclear - classification; Receptors, Cytoplasmic and Nuclear - genetics; Receptors, Cytoplasmic and Nuclear - metabolism; Recombinant Proteins - metabolism; Spodoptera
• ISSN: 0264-6021; 1470-8728
• DOI: 10.1042/bj3280785

Biphasic regulation of inositol trisphosphate (IP3)-stimulated Ca2+ mobilization by cytosolic Ca2+ is believed to contribute to regenerative intracellular Ca2+ signals. Since cells typically express several IP3 receptor isoforms and the effects of cytosolic Ca2+ are not mediated by a single mechanism, it is important to resolve the properties of each receptor subtype. Full-length rat types-1 and -3 IP3 receptors were expressed in insect Sf9 cells at levels 10-40-fold higher than the endogenous receptors. The expressed receptors were glycosylated and assembled into tetramers, and binding of [3H]IP3 to each subtype was regulated by cytosolic Ca2+. The effects of increased [Ca2+] on native cerebellar and type-1 receptors expressed in Sf9 cells were indistinguishable. A maximally effective increase in [Ca2+] reversibly inhibited [3H]IP3 binding by approx. 50% by decreasing the number of IP3-binding sites (Bmax) without affecting their affinity for IP3. The effects of Ca2+ on type-3 receptors were more complex: increasing [Ca2+] first stimulated [3H]IP3 binding by increasing Bmax, and then inhibited it by causing a substantial decrease in the affinity of the receptor for IP3. The different effects of Ca2+ on the receptor subtypes were not a consequence of limitations in the availability of accessory proteins or of artifactual effects of Ca2+ on membrane structure. We conclude that Ca2+ can inhibit IP3 binding to types-1 and -3 IP3 receptors although by different mechanisms, and that IP3 binding to type-3 receptors is stimulated at intermediate [Ca2+]. A consequence of these differences is that, at resting cytosolic [Ca2+], type-3 receptors are more sensitive than type-1 receptors to IP3, but the situation reverses at higher cytosolic [Ca2+]. Such differences may be important in generating the spatially and temporally complex changes in cytosolic [Ca2+] evoked by receptors linked to IP3 formation.