Caspase-3-induced truncation of type 1 inositol trisphosphate receptor accelerates apoptotic cell death and induces inositol trisphosphate-independent calcium release during apoptosis.
Inositol 1,4,5-trisphosphate receptor-deficient (IP3RKO) B-lymphocytes were used to investigate the functional relevance of type 1 inositol 1,4,5-trisphosphate receptor (IP3R1) and its cleavage by caspase-3 in apoptosis. We showed that inositol 1,4,5-trisphosphate receptor-deficient cells were largely resistant to apoptosis induced by both staurosporine (STS) and B-cell receptor (BCR) stimulation. Expression of either the wild-type IP3R1 or an N-terminal deletion mutant (Delta1-225) that lacks inositol 1,4,5-trisphosphate-induced Ca2+ release activity restored sensitivity to apoptosis and the consequent rise in free cytosolic Ca2+ concentration ([Ca2+]i). Expression of caspase-3-non-cleavable mutant receptor, however, dramatically slowed down the rate of apoptosis and prevented both Ca2+ overload and secondary necrosis. Conversely, expression of the "channel-only" domain of IP3R1, a fragment of the receptor generated by caspase-3 cleavage, strongly increased the propensity of the cells to undergo apoptosis. In agreement with these observations, caspase inhibitors impeded apoptosis and the associated rise in [Ca2+]i. Both the staurosporine- and B-cell receptor-induced apoptosis and increase in [Ca2+]i could be induced in nominally Ca2+-free and serum-free culture media, suggesting that the apoptosis-related rise in [Ca2+]i was primarily because of the release from internal stores rather than of influx through the plasma membrane. Altogether, our results suggest that IP3R1 plays a pivotal role in apoptosis and that the increase in [Ca2+]i during apoptosis is mainly the consequence of IP3R1 cleavage by caspase-3. These observations also indicate that expression of a functional IP3R1 per se is not enough to generate the significant levels of cytosolic Ca2+ needed for the rapid execution of apoptosis, but a prior activation of caspase-3 and the resulting truncation of the IP3R1 are required.
Assefa
Zerihun
Z
Afdeling Fysiologie , Katholieke Universiteit Leuven, Campus Gasthuisberg O/N, Herestraat 49, 3000 Leuven, Belgium.
Bultynck
Geert
G
Szlufcik
Karolina
K
Nadif Kasri
Nael
N
Vermassen
Elke
E
Goris
Jozef
J
Missiaen
Ludwig
L
Callewaert
Geert
G
Parys
Jan B
JB
De Smedt
Humbert
H
D000818:Animals; D017209:Apoptosis; D001402:B-Lymphocytes / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000737:chemistry*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D016923:Cell Death; D002462:Cell Membrane / Q000378:metabolism; D002531:Cerebellum / Q000378:metabolism; D002645:Chickens; D016895:Culture Media, Serum-Free / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D017353:Gene Deletion; D015544:Inositol 1,4,5-Trisphosphate / Q000737:chemistry*; D053496:Inositol 1,4,5-Trisphosphate Receptors; D051379:Mice; D008861:Microsomes / Q000378:metabolism; D009154:Mutation; D009336:Necrosis; D017434:Protein Structure, Tertiary; D011947:Receptors, Antigen, B-Cell / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D019311:Staurosporine / Q000494:pharmacology; D013997:Time Factors; D014162:Transfection
D015220:Calcium Channels; D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors; D053496:Inositol 1,4,5-Trisphosphate Receptors; D011947:Receptors, Antigen, B-Cell; D018160:Receptors, Cytoplasmic and Nuclear; D011994:Recombinant Proteins; D015544:Inositol 1,4,5-Trisphosphate; D004247:DNA; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases; D019311:Staurosporine; D002118:Calcium