Rottlerin induces calcium influx and protein degradation in cultured lenses independent of effects on protein kinase C delta.
Rottlerin induces calcium influx and protein degradation in cultured lenses independent of effects on protein kinase C delta.
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DOI：
10.1111/j.1742-7843.2007.00143.x
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发表时间：
2007-12
影响因子：
3.1
通讯作者：
Shang-zhi Xu
中科院分区：
文献类型：
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作者：
Shang-zhi Xu
Rottlerin已被广泛接受为蛋白激酶C δ（PKC δ）的特异性抑制剂;然而，最近的数据表明，这种化合物的特异性成为一个问题。在此，我们使用透镜器官培养系统来解决这个问题，因为PKC δ可能调节透镜中差距连接通透性。有趣的是，我们发现rottlerin诱导的connexin 50的降解比PKC δ更快。此外，将rottlerin与质子载体羰基氰-4-（三氟甲氧基）-苯腙（FCCP）进行比较，发现rottlerin和FCCP都随时间显著增加了透镜重量，所述质子载体与rottlerin具有许多共同特征。这种透镜重量的增加部分逆转的细胞外钙与乙二醇四乙酸（EGTA）的耗竭或通过阻断L-型钙通道与维拉帕米，表明罗特勒可能会诱导钙内流。事实上，由rottlerin和FCCP诱导的连接蛋白50（而不是PKC δ）的快速降解被EGTA阻断。此外，rottlerin和FCCP还诱导Cx46、filensin、vimentin和CP 49的降解。为了确定这种蛋白质降解是否与由于rottlerin解偶联线粒体而导致的ATP减少有关，检查了具有不同处理的晶状体中的ATP含量。结果表明EGTA对透镜ATP含量无影响。综上所述，这些数据表明，rottlerin，像FCCP，诱导钙内流，导致蛋白质降解和裂解的透镜，这种效果是无关的PKC δ的抑制。因此，在考虑使用rottlerin作为PKC δ抑制剂时必须非常谨慎。
Rottlerin has been widely accepted as a specific inhibitor of protein kinase C delta (PKC delta); however, recent data suggest that the specificity of this compound become a question. Herein, we address this issue using a lens organ culture system, as PKC delta might regulate the gap junction permeability in lens. Interestingly, we found that rottlerin induced the degradation of connexin50 more rapidly than that of PKC delta. Furthermore, comparison of rottlerin with a protonophore, carbonylcyanide-4-(trifluoromethoxy)-phenylhydrazone (FCCP) that shares many characteristics with rottlerin, showed that both rottlerin and FCCP dramatically increased lens weight over time. This increase in lens weight was partially reversed by depletion of extracellular calcium with ethyleneglycoltetraacetic acid (EGTA) or by blocking L-type calcium channels with verapamil, suggesting rottlerin may induce calcium influx. Indeed, the rapid degradation of connexin50 (but not PKC delta) induced by rottlerin and FCCP was blocked by EGTA. In addition, rottlerin and FCCP also induced degradation of connexin46, filensin, vimentin and CP49. In order to determine whether this protein degradation is associated with the decrease of ATP due to uncoupling mitochondria by rottlerin, ATP content in lenses with different treatments were examined. The result indicated that EGTA had no effect on lens ATP content. Taken together, these data suggest that rottlerin, like FCCP, induces calcium influx, leading to protein degradation and cleavage in the lens, and that this effect is unrelated to the inhibition of PKC delta. Thus, extreme caution must be taken when considering use of rottlerin as a PKC delta inhibitor.