Gene overexpression/suppression analysis of candidate virulence factors of Candida albicans
Gene overexpression/suppression analysis of candidate virulence factors of Candida albicans
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DOI：
10.1128/ec.00445-07
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发表时间：
2008-03-01
期刊：
影响因子：
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通讯作者：
Ibrahim, Ashraf S.
中科院分区：
文献类型：
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作者：
Fu, Yue;Luo, Guanpingsheng;Ibrahim, Ashraf S.
我们在白色念珠菌中开发了一种有条件的过表达/抑制遗传策略，以便能够同时检测功能的获得或丧失，从而确定新的毒力因子。该策略涉及在感兴趣的基因前插入一个强的四环素调控的启动子。为了验证该策略，在体外筛选了编码糖基磷脂酰肌醇（GPI）锚定表面蛋白的基因库的毒力表型。在筛选过程中，发现IFF 4的过表达增加了C.白色念珠菌对塑料和人类上皮细胞，但不内皮细胞。与体外结果一致，IFF 4过表达适度增加了小鼠阴道念珠菌病期间的组织真菌负荷。除了体外筛选测试外，还发现IFF 4过表达会增加C。白念珠菌对嗜热链球菌介导的杀伤的易感性。此外，IFF 4过表达降低了正常小鼠的血源性播散性念珠菌病的严重程度，但在血小板减少症小鼠中没有，再次与体外表型一致。12种其他GPI蛋白的过表达不影响正常GPI蛋白细胞表面积累，表明过表达策略不影响细胞产生此类蛋白的能力。这些数据表明，相同的基因可以增加或减少念珠菌的毒力在不同的感染模型，强调在不同的解剖学背景下研究毒力基因的重要性。最后，这些数据验证了使用条件性过表达/抑制遗传策略来鉴定念珠菌毒力因子。
We developed a conditional overexpression/suppression genetic strategy in Candida albicans to enable simultaneous testing of gain or loss of function in order to identify new virulence factors. The strategy involved insertion of a strong, tetracycline-regulated promoter in front of the gene of interest. To validate the strategy, a library of genes encoding glycosylphosphatidylinositol (GPI)-anchored surface proteins was screened for virulence phenotypes in vitro. During the screening, overexpression of IFF4 was found to increase the adherence of C. albicans to plastic and to human epithelial cells, but not endothelial cells. Consistent with the in vitro results, IFF4 overexpression modestly increased the tissue fungal burden during murine vaginal candidiasis. In addition to the in vitro screening tests, IFF4 overexpression was found to increase C. albicans susceptibility to neutrophil-mediated killing. Furthermore, IFF4 overexpression decreased the severity of hematogenously disseminated candidiasis in normal mice, but not in neutropenic mice, again consistent with the in vitro phenotype. Overexpression of 12 other GPI proteins did not affect normal GPI protein cell surface accumulation, demonstrating that the overexpression strategy did not affect the cell capacity for making such proteins. These data indicate that the same gene can increase or decrease candidal virulence in distinct models of infection, emphasizing the importance of studying virulence genes in different anatomical contexts. Finally, these data validate the use of a conditional overexpression/suppression genetic strategy to identify candidal virulence factors.