Ribosome depurination is not sufficient for ricin-mediated cell death in Saccharomyces cerevisiae
Ribosome depurination is not sufficient for ricin-mediated cell death in Saccharomyces cerevisiae
复制标题
DOI：
10.1128/iai.01295-06
复制
发表时间：
2007-01-01
影响因子：
3.1
通讯作者：
Tumer, Nigun E.
中科院分区：
文献类型：
--
作者：
Li, Xiao-Ping;Baricevic, Marianne;Tumer, Nigun E.
植物毒素蓖麻毒素是已知的最有效和致命的物质之一。蓖麻毒素通过从大rRNA中高度保守的α-八叠球菌毒素/蓖麻毒素环中去除特定的腺嘌呤来抑制蛋白质合成。人们对蓖麻毒素如何与核糖体相互作用以及它杀死细胞的分子机制知之甚少。为了深入了解蓖麻毒素诱导细胞死亡的机制，我们建立了酵母（酿酒酵母）作为一个简单的和遗传上易于处理的系统，以分离细胞毒性缺陷的突变体。在表达蓖麻毒素A链前体形式（前RTA）的酵母细胞中，核糖体被脱嘌呤，这些细胞显示凋亡标记，如核碎裂、染色质凝聚和活性氧的积累。我们进行了大规模的前RTA诱变，并分离出一组无毒RTA突变体的基础上，他们无法杀死酵母细胞。几个无毒RTA突变体脱嘌呤核糖体和抑制翻译的野生型RTA在体内相同的程度。从酵母中分离的突变蛋白在体外脱嘌呤核糖体，表明它们具有催化活性。然而，表达这些突变体的细胞没有显示凋亡的标志。这些结果提供了第一个证据，证明核糖体脱嘌呤和抑制翻译的能力并不总是与蓖麻毒素介导的细胞死亡相关，表明核糖体脱嘌呤和翻译抑制并不完全解释蓖麻毒素的细胞毒性。
The plant toxin ricin is one of the most potent and lethal substances known. Ricin inhibits protein synthesis by removing a specific adenine from the highly conserved alpha-sarcin/ricin loop in the large rRNA. Very little is known about how ricin interacts with ribosomes and the molecular mechanism by which it kills cells. To gain insight to the mechanism of ricin-induced cell death, we set up yeast (Saccharomyces cerevisiae) as a simple and genetically tractable system to isolate mutants defective in cytotoxicity. Ribosomes were depurinated in yeast cells expressing the precursor form of the A chain of ricin (pre-RTA), and these cells displayed apoptotic markers such as nuclear fragmentation, chromatin condensation, and accumulation of reactive oxygen species. We conducted a large-scale mutagenesis of pre-RTA and isolated a panel of nontoxic RTA mutants based on their inability to kill yeast cells. Several nontoxic RTA mutants depurinated ribosomes and inhibited translation to the same extent as wild-type RTA in vivo. The mutant proteins isolated from yeast depurinated ribosomes in vitro, indicating that they were catalytically active. However, cells expressing these mutants did not display hallmarks of apoptosis. These results provide the first evidence that the ability to depurinate ribosomes and inhibit translation does not always correlate with ricin-mediated cell death, indicating that ribosome depurination and translation inhibition do not account entirely for the cytotoxicity of ricin.