Mutants of Mycobacterium tuberculosis lacking three of the five rpf-like genes are defective for growth in vivo and for resuscitation in vitro
Mutants of Mycobacterium tuberculosis lacking three of the five rpf-like genes are defective for growth in vivo and for resuscitation in vitro
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DOI：
10.1128/iai.73.5.3038-3043.2005
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发表时间：
2005-05-01
影响因子：
3.1
通讯作者：
Mizrahi, V
中科院分区：
文献类型：
--
作者：
Downing, KJ;Mischenko, VV;Mizrahi, V
结核分枝杆菌含有5个基因，rpfA到rpfE，它们与黄色微球菌的复苏促进因子(RPF)基因有显著的同源性，其产物是使休眠的黄色微球菌复苏生长所必需的，对这种生物的生长是必不可少的。先前的研究表明，5个RPF样基因中的任何一个的缺失都不会影响结核分枝杆菌在体外的生长或存活。结合全基因组表达谱的结果，这一发现表明它们的功能冗余。在这项研究中，我们证明了单缺失突变体在体内的表型与野生型结核分枝杆菌H37Rv相似。单个RPF样基因的缺失对液体培养中结核分枝杆菌的生长或长期存活没有明显的影响，而且在最可能的数量分析中，三个被测菌株(Delta rpfB、Delta rpfD和Delta rpfE株)从非培养状态自发复苏的能力也没有受到影响。相比之下，两个多重毒株KDT8(Delta rpfA突变Delta rpfC Delta rpfB)和KDT9(Delta RpfA Delta RpfC Delta RpfD)缺乏五个RPF样基因中的三个，在小鼠感染模型中显著但不同地减弱。这些突变体也不能在体外自发复苏，这表明结核分枝杆菌的RPF样蛋白在从不可培养状态复苏中的重要性。这些结果有力地表明，结核分枝杆菌的五个RPF样基因的生物学功能并不是完全多余的，并强调了这些蛋白作为治疗干预靶点的潜在用途。
Mycobacterium tuberculosis contains five genes, rpfA through rpfE, that bear significant homology to the resuscitation-promoting factor (rpf) gene of Micrococcus luteus, whose product is required to resuscitate the growth of dormant cultures of M. luteus and is essential for the growth of this organism. Previous studies have shown that deletion of any one of the five rpf-like genes did not affect the growth or survival of M. tuberculosis in vitro. In conjunction with the results of whole-genome expression profiling, this finding was indicative of their functional redundancy. In this study, we demonstrate that the single deletion mutants are phenotypically similar to wild-type M. tuberculosis H37Rv in vivo. The deletion of individual rpf-like genes had no discernible effect on the growth or long-term survival of M. tuberculosis in liquid culture, and the ability to resuscitate spontaneously from a nonculturable state in a most probable number assay was also unaffected for the three strains tested (the Delta rpfB, Delta rpfD, and Delta rpfE strains). In contrast, two multiple strains, KDT8 (Delta rpfA-mutation Delta rpfC Delta rpfB) and KDT9 (Delta rpfA Delta rpfC Delta rpfD), which lack three of the five rpf-like genes, were significantly yet differentially attenuated in a mouse infection model. These mutants were also unable to resuscitate spontaneously in vitro, demonstrating the importance of the Rpf-like proteins of M. tuberculosis in resuscitation from the nonculturable state. These results strongly suggest that the biological functions of the five rpf-like genes of M. tuberculosis are not wholly redundant and underscore the potential utility of these proteins as targets for therapeutic intervention.