Gene-trap mutagenesis identifies mammalian genes contributing to intoxication by Clostridium perfringens ε-toxin.
Gene-trap mutagenesis identifies mammalian genes contributing to intoxication by Clostridium perfringens ε-toxin.
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DOI：
10.1371/journal.pone.0017787
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发表时间：
2011-03-11
期刊：
影响因子：
3.7
通讯作者：
McClain MS
中科院分区：
文献类型：
--
作者：
Ivie SE;Fennessey CM;Sheng J;Rubin DH;McClain MS
产气荚膜梭菌ε-毒素是一种与家养反刍动物致死性毒血症相关的极强毒素，对人类可能有毒性。中毒导致液体在各种组织中积聚，最明显的是在大脑和肾脏中。先前的研究表明，该毒素是一种成孔毒素，导致离子稳态失调，最终导致细胞死亡。然而，哺乳动物宿主因素可能有助于ε-毒素诱导的细胞毒性尚不清楚。利用对ε-毒素致毒高度敏感的插入突变型犬肾（MDCK）细胞文库，筛选对ε-毒素诱导的细胞毒性具有抗性的细胞克隆。在9个存活的耐药细胞克隆中鉴定了突变基因。我们对其中一个已确定的基因进行了额外的实验，作为验证实验方法的一种手段。基因表达微阵列分析显示，其中一个已鉴定的基因，甲型肝炎病毒细胞受体1 (HAVCR1, KIM-1, TIM1)，在人肾细胞系中的表达量比在已知对ε-毒素有抗性的人细胞中的表达量要高。一种人类肾脏细胞系ACHN被发现对毒素敏感，并且比其他抗毒素的人类肾脏细胞系表达的HAVCR1蛋白表达量更大。在MDCK和ACHN细胞中的RNA干扰研究证实，HAVCR1参与了ε-毒素诱导的细胞毒性。此外，ε-毒素在体外被证明与HAVCR1结合。本研究结果表明，利用基因诱变和RNA干扰策略鉴定的HAVCR1等基因是研究ε-毒素诱导细胞死亡过程的重要靶点和潜在治疗干预的新靶点。
The Clostridium perfringens ε-toxin is an extremely potent toxin associated with lethal toxemias in domesticated ruminants and may be toxic to humans. Intoxication results in fluid accumulation in various tissues, most notably in the brain and kidneys. Previous studies suggest that the toxin is a pore-forming toxin, leading to dysregulated ion homeostasis and ultimately cell death. However, mammalian host factors that likely contribute to ε-toxin-induced cytotoxicity are poorly understood. A library of insertional mutant Madin Darby canine kidney (MDCK) cells, which are highly susceptible to the lethal affects of ε-toxin, was used to select clones of cells resistant to ε-toxin-induced cytotoxicity. The genes mutated in 9 surviving resistant cell clones were identified. We focused additional experiments on one of the identified genes as a means of validating the experimental approach. Gene expression microarray analysis revealed that one of the identified genes, hepatitis A virus cellular receptor 1 (HAVCR1, KIM-1, TIM1), is more abundantly expressed in human kidney cell lines than it is expressed in human cells known to be resistant to ε-toxin. One human kidney cell line, ACHN, was found to be sensitive to the toxin and expresses a larger isoform of the HAVCR1 protein than the HAVCR1 protein expressed by other, toxin-resistant human kidney cell lines. RNA interference studies in MDCK and in ACHN cells confirmed that HAVCR1 contributes to ε-toxin-induced cytotoxicity. Additionally, ε-toxin was shown to bind to HAVCR1 in vitro. The results of this study indicate that HAVCR1 and the other genes identified through the use of gene-trap mutagenesis and RNA interference strategies represent important targets for investigation of the process by which ε-toxin induces cell death and new targets for potential therapeutic intervention.
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DOI：
10.1126/science.1193032
发表时间：
2010-08-13
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Genovese G;Friedman DJ;Ross MD;Lecordier L;Uzureau P;Freedman BI;Bowden DW;Langefeld CD;Oleksyk TK;Uscinski Knob AL;Bernhardy AJ;Hicks PJ;Nelson GW;Vanhollebeke B;Winkler CA;Kopp JB;Pays E;Pollak MR
通讯作者：
Pollak MR
影响因子：
3.7
作者：
Goldstein J;Morris WE;Loidl CF;Tironi-Farinati C;McClane BA;Uzal FA;Fernandez Miyakawa ME
通讯作者：
Fernandez Miyakawa ME
影响因子：
64.5
作者：
Gurcel, Laure;Abrami, Laurence;van der Goot, F. Gisou
通讯作者：
van der Goot, F. Gisou
影响因子：
11.4
作者：
Kaplan, G;Totsuka, A;Feinstone, SM
通讯作者：
Feinstone, SM
DOI：
10.1152/ajprenal.00199.2007
发表时间：
2007-09-01
影响因子：
4.2
作者：
Chassin, C.;Bens, M.;Vandewalle, A.
通讯作者：
Vandewalle, A.