Differential translocation or phosphorylation of alpha B crystallin cannot he detected in ischemically preconditioned rabbit cardiomyocytes
Differential translocation or phosphorylation of alpha B crystallin cannot he detected in ischemically preconditioned rabbit cardiomyocytes
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DOI：
10.1006/jmcc.2000.1164
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发表时间：
2000-07-01
影响因子：
5
通讯作者：
Ganote, CE
中科院分区：
文献类型：
--
作者：
Armstrong, SC;Shivell, LC;Ganote, CE
α B晶体蛋白（alpha BC）是缺血预处理（IPC）的一种推定效应蛋白，其通过ERK 1/2在Ser 45上磷酸化，通过p38 MAPK底物MAPKAPK-2在Ser 59上磷酸化。通过In SDS-PACE和IEF或使用Ser 45和Ser 59磷酸化特异性抗体在以下细胞中测定胞质和细胞骨架组分中α BC的易位和磷酸化：（1）对照兔心肌细胞;（2）通过10分钟体外缺血预处理的细胞;或在用（3）Ser/Thr蛋白磷酸酶1/2A的特异性抑制剂预处理后（calyculin A）;（4）p38 MAPK（SB 203580）;或（5）ERK 1/2（PD 98059）;所有这些均在180分钟缺血之前。缺血诱导BU细胞质向细胞骨架移位，这在所有组中相似。缺血0分钟时，细胞质中存在高度磷酸化的aBC亚型（D1/2），但细胞骨架部分中不存在。缺血60-90 min时，D1/2异构体移位到细胞骨架部分，Calyculin A在整个缺血过程中维持D1/2水平。SB 203580降低α BC磷酸化。PD 98059和IPC均未改变长时间缺血期间的α BC磷酸化。可以得出结论，缺血期间的α BC磷酸化是受调节的。p38 MAPK和ERK 1/2均不表达。无法检测IPC保护与α BC易位或磷酸化之间的相关性表明，本研究中定量的α BC高度磷酸化同工型条带中的蛋白质不是经典IPC的保护性末端效应物。(C)北京大学出版社.
Alpha B Crystallin (alpha BC) is a putative effector protein of ischemic preconditioning (IPC), that is phosphorylated on Ser 45 by ERK1/2 and Ser 59 by the p38 MAPK substrate, MAPKAPK-2. Translocation and phosphorylation of alpha BC was determined in cytosolic and cytoskeletal fractions by In SDS-PACE and IEF or using Ser 45 and Ser 59 phospho-specific antibodies in: (1) control rabbit cardiomyocytes: (2) cells preconditioned by 10 min in vitro ischemia; or after pre-treatment with specific inhibitors of (3) Ser/Thr protein phosphatase 1/2A (calyculin A); (4) p38 MAPK(SB203580); or (5) ERK 1/2 (PD98059); all prior to 180 min ischemia. Ischemia induced a cytosolic to cytoskeletal translocation of BU, which was similar in all the groups, Highly phosphorylated isoforms (D1/2) of aBC were present in cytosolic but not cytoskeletal fractions at 0 min ischemia. By 60-90 min ischemia, D1/2 isoforms had translocated to the cytoskeletal fraction, Calyculin A maintained D1/2 levels throughout prolonged ischemia. SB203580 decreased alpha BC phosphorylation. Neither PD98059 nor IPC altered alpha BC phosphorylation during prolonged ischemia. It is concluded that alpha BC phosphorylation during ischemia is regulated. by p38 MAPK but nut by ERK 1/2. The inability to detect a correlation between IPC protection and either alpha BC translocation or phosphorylation suggests that the proteins in the highly phosphorylated isoform bands of alpha BC quantitated in this study are not protective end effecters of classical IPC. (C) 2000 Academic Press.