title: The N-terminal Ca2+-independent calmodulin-binding site on the inositol 1,4,5-trisphosphate receptor is responsible for calmodulin inhibition, even though this inhibition requires Ca2+.
abstract: Calmodulin (CaM) is a ubiquitous Ca(2+)-sensor protein that plays an important role in regulating a large number of Ca(2+) channels, including the inositol 1,4,5-trisphosphate receptor (IP(3)R). CaM binds to the IP(3)R at Ca(2+)-dependent as well as at Ca(2+)-independent interaction sites. In this study, we have investigated the Ca(2+)-independent CaM-binding site for its role in the regulation of the Ca(2+)-dependent bell-shaped activation curve of the IP(3)R. Suramin, a polysulfonated napthylurea, displaced CaM in both the presence and the absence of Ca(2+). Suramin competed with CaM for binding to different peptides representing the previously identified CaM-binding sites on IP(3)R1. By interacting with the N-terminal Ca(2+)-independent CaM-binding site, suramin mimicked the functional effect of CaM and induced an allosteric but competitive inhibition of IP(3) binding. Therefore, suramin also potently inhibited IP(3)-induced Ca(2+) release (IICR) from permeabilized cells predominantly expressing IP(3)R1 (L15 fibroblasts) or IP(3)R3 (Lvec fibroblasts), even though the IP(3)R3 does not contain Ca(2+)-dependent CaM-binding sites. Furthermore, we have found that CaM(1234), a CaM mutated in its four EF hands, inhibited IICR in a Ca(2+)-dependent way with the same potency as CaM. We conclude that CaM inhibits IICR via the N-terminal binding site. The inhibition requires Ca(2+) but CaM itself is not the Ca(2+) sensor for the inhibition of the IP(3)R.
authors.affiliation: Laboratorium voor Fysiologie, Katholieke Universiteit Leuven Campus Gasthuisberg O/N, Leuven, Belgium.
authors.forename: Nael Nadif
authors.initials: NN
authors.lastname: Kasri
authors.forename: Geert
authors.initials: G
authors.lastname: Bultynck
authors.forename: Jeremy
authors.initials: J
authors.lastname: Smyth
authors.forename: Karolina
authors.initials: K
authors.lastname: Szlufcik
authors.forename: Jan B
authors.initials: JB
authors.lastname: Parys
authors.forename: Geert
authors.initials: G
authors.lastname: Callewaert
authors.forename: Ludwig
authors.initials: L
authors.lastname: Missiaen
authors.forename: Rafael A
authors.initials: RA
authors.lastname: Fissore
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Mikoshiba
authors.forename: Humbert
authors.initials: H
authors.lastname: de Smedt
mesh_terms: D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism*; D002147:Calmodulin / Q000037:antagonists & inhibitors* / Q000378:metabolism; D002478:Cells, Cultured; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism*; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007313:Insecta; D017434:Protein Structure, Tertiary; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D013498:Suramin / Q000378:metabolism
chemical_list: D015220:Calcium Channels; D002147:Calmodulin; D053496:Inositol 1,4,5-Trisphosphate Receptors; D018160:Receptors, Cytoplasmic and Nuclear; D013498:Suramin; D015544:Inositol 1,4,5-Trisphosphate; D002118:Calcium