Caspase-3-induced truncation of type 1 inositol trisphosphate receptor accelerates apoptotic cell death and induces inositol trisphosphate-independent calcium release during apoptosis
Caspase-3-induced truncation of type 1 inositol trisphosphate receptor accelerates apoptotic cell death and induces inositol trisphosphate-independent calcium release during apoptosis
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DOI：
10.1074/jbc.m403872200
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发表时间：
2004-10-08
影响因子：
4.8
通讯作者：
De Smedt, H
中科院分区：
文献类型：
--
作者：
Assefa, Z;Bultynck, G;De Smedt, H
本研究采用1，4，5-三磷酸肌醇受体缺陷型（IP 3R-KO）B淋巴细胞，探讨了1型1，4，5-三磷酸肌醇受体（IP（3）R1）及其被caspase-3裂解在凋亡中的功能相关性。我们发现，肌醇1，4，5-三磷酸受体缺陷的细胞在很大程度上抵抗由星形孢菌素（STS）和B细胞受体（BCR）刺激诱导的凋亡。野生型IP（3）R1或缺乏肌醇1，4，5-三磷酸诱导的Ca 2+释放活性的N-末端缺失突变体（Delta 1 -225）的表达恢复了对细胞凋亡的敏感性和随之而来的游离胞质Ca 2+浓度的升高（[Ca 2 +]（i））。然而，caspase-3-不可切割的突变体受体的表达，大大减缓了细胞凋亡的速率，并防止了Ca 2+超载和继发性坏死。相反，IP（3）R1的“仅通道”结构域（caspase-3裂解产生的受体片段）的表达强烈增加了细胞凋亡的倾向。与这些观察结果一致，半胱天冬酶抑制剂阻碍了细胞凋亡和相关的[Ca 2 +]升高（i）。星形孢菌素和B细胞受体诱导的细胞凋亡和[Ca 2 +]（i）的增加都可以在名义上无Ca 2+和无血清的培养基中诱导，这表明与细胞凋亡相关的[Ca 2 +]（i）的升高主要是因为从内部储存释放，而不是通过质膜流入。总之，我们的研究结果表明，IP（3）R1在细胞凋亡中起着关键作用，细胞凋亡过程中[Ca 2 +]（i）的增加主要是IP（3）R1被caspase-3切割的结果。这些观察结果还表明，功能性IP（3）R1本身的表达不足以产生快速执行细胞凋亡所需的显著水平的胞质Ca 2+，但需要预先激活caspase-3并导致IP（3）R1的截短。
Inositol 1,4,5-trisphosphate receptor-deficient (IP3R-KO) B-lymphocytes were used to investigate the functional relevance of type 1 inositol 1,4,5-trisphosphate receptor (IP(3)R1) 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 IP(3)R1 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 IP(3)R1, 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 IP(3)R1 plays a pivotal role in apoptosis and that the increase in [Ca2+](i) during apoptosis is mainly the consequence of IP(3)R1 cleavage by caspase-3. These observations also indicate that expression of a functional IP(3)R1 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 IP(3)R1 are required.