Manganese transport and virulence in Brucella

描述（由申请人提供）：质子同调子MntH是布鲁氏菌菌株中唯一的高亲和力锰转运体(2)。在实验感染的小鼠中，abortus B. mntH突变体显示出的极度衰减表明，与已检测的其他细菌病原体相比，Mn在布鲁氏菌菌株的毒力中起着异常重要的作用。葡萄糖分解代谢最近被证明对布鲁氏菌菌株在交替激活的巨噬细胞中持续存在的能力至关重要，而这反过来又需要在实验感染的小鼠中维持慢性感染（51）。布鲁氏菌只通过戊糖磷酸途径分解葡萄糖（15），而维持该途径碳流所需的两种酶——丙酮酸激酶（PykM）（20）和核酮糖-5-磷酸3- epimase (Rpe)（46）的活性被预测特别容易受到细胞Mn水平降低的影响。因此，本申请的具体目标1中概述的研究将验证这样一个假设，即PykM和Rpe活性降低导致的葡萄糖分解代谢缺陷对B. abortus mntH突变体表现出的衰减有重要贡献。实验证据还表明，布鲁氏菌菌株有能力增加mnth介导的Mn摄取，并以这种金属代替细胞蛋白中的铁，作为抗氧化应激的保护机制，其方式与最近在大肠杆菌中证明的类似(3)。本提案Specific Aim 2中描述的研究将确定负责H2O2响应性诱导B. abortus 2308中mntH表达的转录调节因子，并确定这种mntH调节模式是否在体外保护该菌株免受氧化应激，以及其在小鼠中的毒力是否必需。从拟议的研究中获得的结果将开始解释锰在布鲁氏菌菌株的基本生理和毒力中起着如此关键的作用。失去MntH对毒力有如此不利影响的事实也使这种转运体成为开发改进疫苗和抗生素以预防和治疗人类布鲁氏菌病（一种具有世界重要性的主要人畜共患疾病）的有吸引力的靶标。

DESCRIPTION (provided by applicant): The proton symporter MntH serves as the sole high affinity manganese transporter in Brucella strains (2). The extreme attenuation displayed by a B. abortus mntH mutant in experimentally infected mice indicates that Mn plays an exceptionally important role in the virulence of Brucella strains in comparison with other bacterial pathogens that have been examined. Glucose catabolism has recently been shown to be critical for the ability of Brucella strains to persist in alternatively activated macrophages, which in turn is required for the maintenance of chronic infections in experimentally infected mice (51). Brucella strains catabolize glucose exclusively via the pentose phosphate pathway (15), and the activities of two enzymes required for maintaining carbon flow through this pathway, pyruvate kinase (PykM) (20) and ribulose-5-phosphate 3-epimerase (Rpe) (46), are predicted to the particularly susceptible to reduced cellular Mn levels. Consequently, the studies outlined in Specific Aim 1 of this application will test the hypothesis that defective glucose catabolism resulting from reduced PykM and Rpe activities makes a significant contribution to the attenuation exhibited by the B. abortus mntH mutant. Experimental evidence also suggests that Brucella strains have the capacity to increase their MntH-mediated Mn uptake and substitute this metal for iron in cellular proteins as a protective mechanism against oxidative stress, in a similar manner to that recently demonstrated in Escherichia coli (3). The studies described in Specific Aim 2 of this proposal will identify the transcriptional regulator responsible for the H2O2- responsive induction of mntH expression in B. abortus 2308, and determine if this mode of mntH regulation protects this strain from oxidative stress in vitro and is required for its virulence in mice. Results obtained from the proposed studies will begin to explain Mn plays such a critical role in the basic physiology and virulence of Brucella strains. The fact that loss f MntH has such an adverse effect on virulence also makes this transporter an attractive target for the development of improved vaccines and antibiotics to prevent and treat human brucellosis, a major zoonotic disease of worldwide importance.