Manganese transport and virulence in Brucella

描述(申请人提供)：质子转运蛋白mnth是布鲁氏菌菌株中唯一的高亲和力锰转运蛋白(2)。一个流产杆菌突变株在实验感染的小鼠中表现出的极端衰减表明，与已检测的其他细菌病原体相比，锰在布鲁氏菌菌株的毒力中起着特别重要的作用。最近证明，葡萄糖分解代谢对于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，而巨噬细胞又是维持实验感染小鼠慢性感染所必需的(51)。布鲁氏菌只通过磷酸戊糖途径分解葡萄糖(15)，而维持通过这一途径的碳流所需的两种酶--丙酮酸激酶(PYKM)(20)和核酮糖-5-磷酸3-异构酶(RPE)(46)的活性被预测为特别容易受到细胞内锰水平降低的影响。因此，在本申请的特定目标1中概述的研究将检验这样的假设，即由于PYKM和RPE活性降低而导致的葡萄糖分解代谢缺陷对B.bortus mnth突变体表现出的衰减有显著贡献。实验证据还表明，布鲁氏菌菌株有能力增加其mnth介导的锰摄取，并在细胞蛋白质中用这种金属取代铁作为对抗氧化应激的保护机制，与最近在大肠杆菌中展示的方式相似[3]。在本提案的特定目标2中描述的研究将确定负责过氧化氢响应诱导流产杆菌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所必需。拟议的研究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nth对毒力有如此不利的影响这一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的有吸引力的目标，布鲁氏菌病是一种具有全球重要性的主要人畜共患病。

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.