siRNA Screening Identifies the Host Hexokinase 2 (HK2) Gene as an Important Hypoxia-Inducible Transcription Factor 1 (HIF-1) Target Gene in Toxoplasma gondii-Infected Cells.
siRNA Screening Identifies the Host Hexokinase 2 (HK2) Gene as an Important Hypoxia-Inducible Transcription Factor 1 (HIF-1) Target Gene in Toxoplasma gondii-Infected Cells.
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siRNA 筛选将宿主己糖激酶 2 (HK2) 基因鉴定为弓形虫感染细胞中重要的缺氧诱导转录因子 1 (HIF-1) 靶基因。
DOI：
10.1128/mbio.00462-15
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发表时间：
2015-06-23
期刊：
影响因子：
6.4
通讯作者：
Blader IJ
中科院分区：
文献类型：
--
作者：
Menendez MT;Teygong C;Wade K;Florimond C;Blader IJ
虽然已经确定氧的可利用性调节细胞的代谢和生长，但关于细胞内病原体如何利用宿主因子在生理氧水平下生长的知之甚少。因此，进行大规模的人小干扰RNA筛选，以确定重要的细胞内原生动物寄生虫弓形虫在组织氧分压下的生长宿主基因。在通过该筛选鉴定的基因中，我们集中于己糖激酶2（HK 2）基因，因为其表达受低氧诱导转录因子1（HIF-1）调节，这对弓形虫生长很重要。弓形虫以HIF-1依赖的方式增加宿主HK 2转录物和蛋白水平。此外，在用HK 2表达质粒转染的HIF-1缺陷细胞中，在3%氧气下的寄生虫生长得以恢复。HIF-1激活和HK 2表达都伴随着宿主糖酵解通量的增加，这表明在寄生虫感染的细胞中增强的HK 2表达在功能上是重要的。寄生虫对宿主HK 2和HIF-1表达的依赖性不限于转化细胞系，因为两者都是寄生虫在非转化C2 C12成肌细胞中生长所需的，并且HK 2在感染后体内上调。虽然HK 2通常在生理O2水平下与线粒体外膜的细胞质面相关，但HK 2在感染后重新定位到宿主细胞质，这是寄生虫在3%氧气下生长所需的过程。总之，我们的研究结果表明，HIF-1依赖的表达和重新定位的HK 2代表了一种新的机制，弓形虫建立其复制生态位在组织氧张力。关于宿主细胞如何在模拟组织中发现的氧水平下对细胞内寄生虫弓形虫的存活做出贡献，我们知之甚少。我们以前的工作表明，弓形虫激活了生长所需的氧调节转录因子的表达。在这里，我们报告说，弓形虫调节的丰度和活性的一个关键的宿主代谢酶，己糖激酶2，通过激活HIF-1和促进解离的己糖激酶2从线粒体膜。总的来说，我们的数据揭示HIF-1/己糖激酶2作为细胞内病原体的新靶点，其通过重新编程宿主细胞的代谢来产生有利于寄生虫在生理氧水平下复制的环境。
Although it is established that oxygen availability regulates cellular metabolism and growth, little is known regarding how intracellular pathogens use host factors to grow at physiological oxygen levels. Therefore, large-scale human small interfering RNA screening was performed to identify host genes important for growth of the intracellular protozoan parasite Toxoplasma gondii at tissue oxygen tensions. Among the genes identified by this screen, we focused on the hexokinase 2 (HK2) gene because its expression is regulated by hypoxia-inducible transcription factor 1 (HIF-1), which is important for Toxoplasma growth. Toxoplasma increases host HK2 transcript and protein levels in a HIF-1-dependent manner. In addition, parasite growth at 3% oxygen is restored in HIF-1-deficient cells transfected with HK2 expression plasmids. Both HIF-1 activation and HK2 expression were accompanied by increases in host glycolytic flux, suggesting that enhanced HK2 expression in parasite-infected cells is functionally significant. Parasite dependence on host HK2 and HIF-1 expression is not restricted to transformed cell lines, as both are required for parasite growth in nontransformed C2C12 myoblasts and HK2 is upregulated in vivo following infection. While HK2 is normally associated with the cytoplasmic face of the outer mitochondrial membrane at physiological O2 levels, HK2 relocalizes to the host cytoplasm following infection, a process that is required for parasite growth at 3% oxygen. Taken together, our findings show that HIF-1-dependent expression and relocalization of HK2 represent a novel mechanism by which Toxoplasma establishes its replicative niche at tissue oxygen tensions. Little is known regarding how the host cell contributes to the survival of the intracellular parasite Toxoplasma gondii at oxygen levels that mimic those found in tissues. Our previous work showed that Toxoplasma activates the expression of an oxygen-regulated transcription factor that is required for growth. Here, we report that Toxoplasma regulates the abundance and activity of a key host metabolic enzyme, hexokinase 2, by activating HIF-1 and by promoting dissociation of hexokinase 2 from the mitochondrial membrane. Collectively, our data reveal HIF-1/hexokinase 2 as a novel target for an intracellular pathogen that acts by reprograming the host cell’s metabolism to create an environment conducive for parasite replication at physiological oxygen levels.