Fas-induced arachidonic acid release is mediated by Ca2+-independent phospholipase A2 but not cytosolic phospholipase A2 which undergoes proteolytic inactivation
Fas-induced arachidonic acid release is mediated by Ca2+-independent phospholipase A2 but not cytosolic phospholipase A2 which undergoes proteolytic inactivation
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DOI：
10.1074/jbc.273.22.13870
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发表时间：
1998-05-29
影响因子：
4.8
通讯作者：
Kudo, I
中科院分区：
文献类型：
--
作者：
Atsumi, G;Tajima, M;Kudo, I
Fas介导的人白血病U937细胞凋亡伴随着膜甘油磷脂中花生四烯酸（AA）和油酸释放的增加，表明磷脂酶A（2）（PLA（2））激活。在细胞凋亡过程中，IV型胞质PLA（2）（cPLA（2）），一种PLA（2）同工酶，其表观分子量为110 kDa，对刺激偶联AA释放至关重要，转化为78 kDa片段，伴随着催化活性的丧失。cPLA（2）的裂解与凋亡细胞中caspase-3样蛋白酶活性的增加相关，并可被caspase-3抑制剂消除。突变型cPLA（2）蛋白中Asp（522）被Asn取代，与caspase-3切割位点的共有序列（DXXD向下箭头X）对齐，对凋亡相关蛋白水解具有抗性。此外，cPLA（2）的COOH-末端缺失突变体（在Asp（522）处截短）与78-kDa片段一起迁移，并且没有表现出酶活性。因此，caspase-3介导的cPLA（2）裂解最终导致cPLA活性所必需的催化三联体的破坏，从而终止其AA释放功能。相反，VI型Ca2+非依赖性PLA（2）（iPLA（2））（一种涉及磷脂重塑的PLA（2）同工酶）的活性在细胞凋亡期间保持完整。iPLA（2）的抑制剂，但cPLA（2）和分泌型PLA（2）抑制剂均不显著抑制AA释放，重要的是，延迟Fas诱导的细胞死亡。我们的结论是iPLA（2）介导的脂肪酸释放在Fas刺激的细胞中被促进，并且在凋亡性细胞死亡过程中起着修饰性的但不是必需的作用。
Fas-mediated apoptosis of human leukemic U937 cells was accompanied by increased arachidonic acid (AA) and oleic acid release from membrane glycerophospholipids, indicating phospholipase A(2) (PLA(2)) activation. During apoptosis, type IV cytosolic PLA(2) (cPLA(2)), a PLA(2) isozyme with an apparent molecular mass of 110 kDa critical for stimulus-coupled AA release, was converted to a 78-kDa fragment with concomitant loss of catalytic activity. Cleavage of cPLA(2) correlated with increased caspase-3-like protease activity in apoptotic cells and was abrogated by a caspase-3 inhibitor. A mutant cPLA(2) protein in which Asp(522) was replaced by Asn, which aligns with the consensus sequence of the caspase-3 cleavage site (DXXD down arrow X), was resistant to apoptosis-associated proteolysis. Moreover, a COOH-terminal deletion mutant of cPLA(2) truncated at Asp(522) comigrated with the 78-kDa fragment and exhibited no enzymatic activity. Thus, caspase-3-mediated cPLA(2) cleavage eventually leads to destruction of a catalytic triad essential for cPLA, activity, thereby terminating its AA-releasing function. In contrast, the activity of type VI Ca2+-independent PLA(2) (iPLA(2)), a PLA(2) isozyme implicated in phospholipid remodeling, remained intact during apoptosis, Inhibitors of iPLA(2), but neither cPLA(2) nor secretory PLA(2) inhibitors, suppressed AA release markedly and, importantly, delayed cell death induced by Fas, Therefore, we conclude that iPLA(2)-mediated fatty acid release is facilitated in Fas-stimulated cells and plays a modifying although not essential role in the apoptotic cell death process.