Weller M, Malipiero U, Groscurth P, Fontana A
Department of Internal Medicine, University Hospital Zürich, Switzerland.
Exp Cell Res. 1995 Dec;221(2):395-403. doi: 10.1006/excr.1995.1390.
The phosphatase inhibitors okadaic acid and calyculin A were used to examine the role of phosphorylation processes in T cell apoptosis induced by interleukin-2 (IL-2) deprivation or transforming growth factor-beta 2 (TGF-beta 2). Okadaic acid and calyculin A inhibited IL-2-driven T cell proliferation and induced apoptosis at concentrations known to inhibit protein phosphatase 1. High concentrations of both agents caused toxic changes of prominent cellular swelling and dilatation of rough endoplasmic reticular profiles. When the T cells were induced to undergo apoptosis by IL-2 deprivation, okadaic acid and calyculin A delayed loss of membrane integrity, nucleosomal size DNA fragmentation, and loss of bcl-2 mRNA. However, T cells deprived of IL-2 in the presence of okadaic acid or calyculin A revealed DNA breaks by in situ DNA end labeling and apoptotic morphology by electron microscopy and failed to show enhanced survival after reexposure to IL-2. Although TGF-beta-mediated signaling is thought to involve the dephosphorylation of specific substrates, okadaic acid and calyculin A not only failed to inhibit, but actually augmented, TGF-beta 2-induced inhibition of T cell proliferation and induction of apoptosis. Exposure to either TGF-beta 2 or the phosphatase inhibitors prevented phosphorylation of the retinoblastoma protein RB. In summary, okadaic acid and calyculin A: (i) induce T cell apoptosis in the presence of IL-2, (ii) allow us to distinguish essential from epiphenomenal features of T cell apoptosis after IL-2 deprivation, and (iii) cooperate with TGF-beta 2 in inducing growth arrest and apoptosis of murine T cells via intracellular cascades that converge in the prevention of RB phosphorylation.
使用磷酸酶抑制剂冈田酸和花萼海绵诱癌素A来研究磷酸化过程在白细胞介素-2（IL-2）缺乏或转化生长因子-β2（TGF-β2）诱导的T细胞凋亡中的作用。冈田酸和花萼海绵诱癌素A在已知可抑制蛋白磷酸酶1的浓度下，抑制IL-2驱动的T细胞增殖并诱导凋亡。两种药物的高浓度都会导致明显的细胞肿胀和粗面内质网轮廓扩张等毒性变化。当T细胞因IL-2缺乏而被诱导发生凋亡时，冈田酸和花萼海绵诱癌素A会延迟膜完整性丧失、核小体大小的DNA片段化以及bcl-2 mRNA的丧失。然而，在冈田酸或花萼海绵诱癌素A存在的情况下缺乏IL-2的T细胞，通过原位DNA末端标记显示出DNA断裂，通过电子显微镜显示出凋亡形态，并且在重新暴露于IL-2后未能显示出存活率提高。尽管TGF-β介导的信号传导被认为涉及特定底物的去磷酸化，但冈田酸和花萼海绵诱癌素A不仅未能抑制，反而实际上增强了TGF-β2诱导的T细胞增殖抑制和凋亡诱导。暴露于TGF-β2或磷酸酶抑制剂均可阻止视网膜母细胞瘤蛋白RB的磷酸化。总之，冈田酸和花萼海绵诱癌素A：（i）在IL-2存在的情况下诱导T细胞凋亡，（ii）使我们能够区分IL-2缺乏后T细胞凋亡的本质特征和现象特征，以及（iii）通过在预防RB磷酸化中汇聚的细胞内级联反应，与TGF-β2协同诱导小鼠T细胞的生长停滞和凋亡。
