Caimano Melissa J, Eggers Christian H, Hazlett Karsten R O, Radolf Justin D
Center for Microbial Pathogenesis, University of Connecticut Health Center, 263 Farmington Ave., Farmington 06030-3710, USA.
Infect Immun. 2004 Nov;72(11):6433-45. doi: 10.1128/IAI.72.11.6433-6445.2004.
Borrelia burgdorferi, the Lyme disease spirochete, undergoes dramatic changes in antigenic composition as it cycles between its arthropod and mammalian hosts. A growing body of evidence suggests that these changes reflect, at least in part, the need for spirochetes to adapt to the physiological stresses imposed by abrupt changes in environmental conditions and nutrient availability. In many microorganisms, global responses are mediated by master regulators such as alternative sigma factors, with Escherichia coli RpoS (sigmaS) serving as a prototype. The importance of this transcriptional activator in other bacteria, coupled with the report by Hubner et al. (A. Hubner, X. Yang, D. M. Nolen, T. G. Popova, F. C. Cabello, and M. V. Norgard, Proc. Natl. Acad. Sci. USA 98:12724-12729, 2001) demonstrating that the borrelial RpoS ortholog controls expression of OspC and decorin-binding protein A (DbpA), prompted us to examine more closely the roles of RpoS-dependent and -independent differential gene expression in physiological adaptation by the Lyme disease spirochete. We observed that B. burgdorferi rpoS (rpoSBb) was induced following temperature shift and transcript levels were further enhanced by reduced pH (pH 6.8). Using quantitative real-time reverse transcription-PCR (RT-PCR), we demonstrated that, in contrast to its ortholog (rpoSEc) in Escherichia coli, rpoSBb was expressed at significant levels in B. burgdorferi throughout all phases of growth following temperature shift. By comparing a B. burgdorferi strain 297 rpoSBb mutant to its wild-type counterpart, we determined that RpoSBb was not required for survival following exposure to a wide range of environmental stresses (i.e., temperature shift, serum starvation, increased osmolality, reactive oxygen intermediates, and increased or reduced oxygen tension), although the mutant was more sensitive to extremes of pH. While B. burgdorferi strains lacking RpoS were able to survive within intraperitoneal dialysis membrane chambers at a level equivalent to that of the wild type, they were avirulent in mice. Lastly, RT-PCR analysis of the ospE-ospF-elp paralogous lipoprotein families complements earlier findings that many temperature-inducible borrelial loci are controlled in an RpoSBb-independent manner. Together, these data point to fundamental differences between the role(s) of RpoS in B. burgdorferi and that in E. coli. Rather than functioning as a master regulator, RpoSBb appears to serve as a stress-responsive activator of a subset of virulence determinants that, together with the RpoS-independent, differentially expressed regulon, encompass the spirochete's genetic programs required for mammalian host adaptation.
莱姆病螺旋体伯氏疏螺旋体（Borrelia burgdorferi）在其节肢动物宿主和哺乳动物宿主之间循环时，抗原组成会发生显著变化。越来越多的证据表明，这些变化至少部分反映了螺旋体需要适应环境条件和营养可用性突然变化所带来的生理压力。在许多微生物中，全局反应是由诸如替代σ因子等主调节因子介导的，大肠杆菌的RpoS（σS）就是一个典型例子。这种转录激活因子在其他细菌中的重要性，再加上胡布纳等人（A. Hubner、X. Yang、D. M. Nolen、T. G. Popova、F. C. Cabello和M. V. Norgard，《美国国家科学院院刊》98:12724 - 12729，2001年）的报告表明伯氏疏螺旋体的RpoS直系同源物控制OspC和饰胶蛋白聚糖结合蛋白A（DbpA）的表达，促使我们更仔细地研究RpoS依赖性和非依赖性差异基因表达在莱姆病螺旋体生理适应中的作用。我们观察到，温度变化后伯氏疏螺旋体rpoS（rpoSBb）被诱导，且转录水平在pH降低（pH 6.8）时进一步增强。使用定量实时逆转录PCR（RT-PCR），我们证明，与其在大肠杆菌中的直系同源物（rpoSEc）不同，温度变化后，rpoSBb在伯氏疏螺旋体生长的所有阶段都有显著表达。通过将伯氏疏螺旋体菌株297的rpoSBb突变体与其野生型对应物进行比较，我们确定，暴露于多种环境压力（即温度变化、血清饥饿、渗透压增加、活性氧中间体以及氧气张力增加或降低）后，RpoSBb并非生存所必需，尽管该突变体对极端pH更为敏感。虽然缺乏RpoS的伯氏疏螺旋体菌株能够在腹膜透析膜腔室内以与野生型相当的水平存活，但它们在小鼠中无毒力。最后，对ospE - ospF - elp旁系脂蛋白家族的RT-PCR分析补充了早期的发现，即许多温度诱导的伯氏疏螺旋体基因座是以独立于RpoSBb的方式控制的。总之，这些数据表明RpoS在伯氏疏螺旋体和大肠杆菌中的作用存在根本差异。RpoSBb似乎并非作为主调节因子发挥作用，而是作为一组毒力决定因素的应激反应激活因子，与独立于RpoS的差异表达调节子一起，涵盖了螺旋体适应哺乳动物宿主所需的遗传程序。
