Potent and selective inhibitors of the inositol-requiring enzyme 1 endoribonuclease.
Potent and selective inhibitors of the inositol-requiring enzyme 1 endoribonuclease.
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DOI：
10.1074/jbc.m110.199737
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发表时间：
2011-04-08
期刊：
影响因子：
--
通讯作者：
Patterson JB
中科院分区：
文献类型：
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作者：
Volkmann K;Lucas JL;Vuga D;Wang X;Brumm D;Stiles C;Kriebel D;Der-Sarkissian A;Krishnan K;Schweitzer C;Liu Z;Malyankar UM;Chiovitti D;Canny M;Durocher D;Sicheri F;Patterson JB
肌醇需求酶 1 (IRE1) 是未折叠蛋白反应 (UPR) 中最高度保守的信号传导节点，是许多与内质网应激相关疾病的潜在治疗靶点。 IRE1 通过定点切割 mRNA 内的两个发夹环来激活 XBP-1 转录因子，以促进其非常规剪接和选择性翻译。我们使用含有人 IRE1α (hIRE1α-细胞) 的独特胞质激酶和核糖核酸内切酶结构域的构建体以及 mini-XBP-1 茎环 RNA 作为底物筛选抑制剂。一类化合物是来自文库中水杨醛亚胺水解产物的水杨醛类似物。水杨醛类似物能够以剂量依赖性方式有效抑制多个 mini-XBP-1 茎环 RNA 的位点特异性切割。水杨醛类似物也能有效抑制酵母 Ire1，但几乎没有抑制 RNase L 或不相关的 RNase A 和 T1 的活性。动力学分析表明，一种有效的水杨醛类似物 3-乙氧基-5,6-二溴水杨醛是 XBP-1 RNA 底物的非竞争性抑制剂。表面等离子共振研究证实该化合物以特异性、可逆且剂量依赖性的方式与 IRE1 结合。水杨醛可抑制人体细胞中药理学诱导的 XBP-1 剪接。这些化合物还阻断已知 XBP-1 靶标以及 IRE1 降解的 mRNA 的转录上调。最后，水杨醛类似物 3-甲氧基-6-溴水杨醛在急性内质网应激体内模型中强烈抑制 XBP-1 剪接。据我们所知，水杨醛类似物是第一个报道的特异性 IRE1 核糖核酸内切酶抑制剂。
Inositol-requiring enzyme 1 (IRE1) is the most highly conserved signaling node of the unfolded protein response (UPR) and represents a potential therapeutic target for a number of diseases associated with endoplasmic reticulum stress. IRE1 activates the XBP-1 transcription factor by site-specific cleavage of two hairpin loops within its mRNA to facilitate its nonconventional splicing and alternative translation. We screened for inhibitors using a construct containing the unique cytosolic kinase and endoribonuclease domains of human IRE1α (hIRE1α-cyto) and a mini-XBP-1 stem-loop RNA as the substrate. One class compounds was salicylaldehyde analogs from the hydrolyzed product of salicylaldimines in the library. Salicylaldehyde analogs were active in inhibiting the site-specific cleavage of several mini-XBP-1 stem-loop RNAs in a dose-dependent manner. Salicyaldehyde analogs were also active in inhibiting yeast Ire1 but had little activity inhibiting RNase L or the unrelated RNases A and T1. Kinetic analysis revealed that one potent salicylaldehyde analog, 3-ethoxy-5,6-dibromosalicylaldehyde, is a non-competitive inhibitor with respect to the XBP-1 RNA substrate. Surface plasmon resonance studies confirmed this compound bound to IRE1 in a specific, reversible and dose-dependent manner. Salicylaldehydes inhibited XBP-1 splicing induced pharmacologically in human cells. These compounds also blocked transcriptional up-regulation of known XBP-1 targets as well as mRNAs targeted for degradation by IRE1. Finally, the salicylaldehyde analog 3-methoxy-6-bromosalicylaldehyde strongly inhibited XBP-1 splicing in an in vivo model of acute endoplasmic reticulum stress. To our knowledge, salicylaldehyde analogs are the first reported specific IRE1 endoribonuclease inhibitors.