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.
Department of Pharmacology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QJ, UK.
T J
TJ
Cardy
C W
CW
Taylor
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