A novel role for calmodulin: Ca2+-independent inhibition of type-1 inositol trisphosphate receptors.
Calmodulin inhibits both inositol 1,4,5-trisphosphate (IP3) binding to, and IP3-evoked Ca2+ release by, cerebellar IP3 receptors [Patel, Morris, Adkins, O'Beirne and Taylor (1997) Proc. Natl. Acad. Sci. U. S.A. 94, 11627-11632]. In the present study, full-length rat type-1 and -3 IP3 receptors were expressed at high levels in insect Spodoptera frugiperda 9 cells and the effects of calmodulin were examined. In the absence of Ca2+, calmodulin caused a concentration-dependent and reversible inhibition of [3H]IP3 binding to type-1 IP3 receptors by decreasing their apparent affinity for IP3. The effect was not reproduced by high concentrations of troponin C, parvalbumin or S-100. Increasing the medium free [Ca2+] ([Ca2+]m) inhibited [3H]IP3 binding to type-1 receptors, but the further inhibition caused by a submaximal concentration of calmodulin was similar at each [Ca2+]m. In the absence of Ca2+, 125I-calmodulin bound to a single site on each type-1 receptor subunit and to an additional site in the presence of Ca2+. There was no detectable binding of 125I-calmodulin to type-3 receptors and binding of [3H]IP3 was insensitive to calmodulin at all [Ca2+]m. Both peptide and conventional Ca2+-calmodulin antagonists affected neither [3H]IP3 binding directly nor the inhibitory effect of calmodulin in the absence of Ca2+, but each caused a [Ca2+]m-dependent reversal of the inhibition of [3H]IP3 binding caused by calmodulin. Camstatin, a peptide that binds to calmodulin equally well in the presence or absence of Ca2+, reversed the inhibitory effects of calmodulin on [3H]IP3 binding at all [Ca2+]m. We conclude that calmodulin specifically inhibits [3H]IP3 binding to type-1 IP3 receptors: the first example of a protein regulated by calmodulin in an entirely Ca2+-independent manner. Inhibition of type-1 IP3 receptors by calmodulin may dynamically regulate their sensitivity to IP3 in response to the changes in cytosolic free calmodulin concentration thought to accompany stimulation of neurones.
Cardy
T J
TJ
Department of Pharmacology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QJ, UK.
Taylor
C W
CW
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology*; D015220:Calcium Channels / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002147:Calmodulin / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D002460:Cell Line; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007700:Kinetics; D008969:Molecular Sequence Data; D010320:Parvalbumins / Q000494:pharmacology; D010446:Peptide Fragments / Q000737:chemistry; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D018411:Spodoptera; D014162:Transfection; D019209:Troponin C / Q000494:pharmacology
D015220:Calcium Channels; D002147:Calmodulin; D053496:Inositol 1,4,5-Trisphosphate Receptors; D010320:Parvalbumins; D010446:Peptide Fragments; D018160:Receptors, Cytoplasmic and Nuclear; D011994:Recombinant Proteins; D019209:Troponin C; D015544:Inositol 1,4,5-Trisphosphate; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002118:Calcium