Mycobacterium tuberculosis universal stress protein Rv2623 regulates bacillary growth by ATP-Binding: requirement for establishing chronic persistent infection.
Mycobacterium tuberculosis universal stress protein Rv2623 regulates bacillary growth by ATP-Binding: requirement for establishing chronic persistent infection.
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结核分枝杆菌通用应激蛋白 Rv2623 通过 ATP 结合调节细菌生长：建立慢性持续感染的必要条件。
DOI：
10.1371/journal.ppat.1000460
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发表时间：
2009-05
期刊：
影响因子：
6.7
通讯作者：
Chan J
中科院分区：
文献类型：
--
作者：
Drumm JE;Mi K;Bilder P;Sun M;Lim J;Bielefeldt-Ohmann H;Basaraba R;So M;Zhu G;Tufariello JM;Izzo AA;Orme IM;Almo SC;Leyh TS;Chan J
结核潜伏期和再激活在结核病的发病机制中发挥着重要作用，但调节这些过程的机制仍不清楚。结核分枝杆菌通用应激蛋白 (USP) 同源物 rv2623 是当结核杆菌受到缺氧和亚硝化应激（被认为会促进潜伏期的条件）时诱导程度最高的基因之一。当结核分枝杆菌遇到与生长停滞相关的条件时，例如巨噬细胞的细胞内环境和慢性结核病小鼠的肺部，也会发生 rv2623 的诱导。因此，我们测试了 Rv2623 调节结核潜伏期的假设。我们观察到 Rv2623 缺陷突变体未能在豚鼠和小鼠中建立慢性结核感染，表现出与细菌负荷和死亡率增加相关的高毒力表型。与这种体内生长调节作用一致，rv2623 的组成型过度表达会减弱分枝杆菌的体外生长。对纯化的 Rv2623 的生化分析表明，这种分枝杆菌 USP 结合 ATP，并且 2.9 Å 分辨率的晶体结构表明，Rv2623 在一个新的核苷酸结合袋中与 ATP 结合。结构引导诱变产生了 ATP 结合能力降低的 Rv2623 突变体。对过表达这些突变体的分枝杆菌的分析表明，Rv2623 的体外生长抑制特性与其结合 ATP 的能力相关。总之，结果表明 i) 结核分枝杆菌 Rv2623 在体外和体内调节分枝杆菌生长，ii) Rv2623 是结核杆菌进入宿主慢性感染阶段所必需的；此外，iii) Rv2623 结合 ATP； iv) 该 USP 的生长调节属性取决于其 ATP 结合活性。我们认为 Rv2623 可能在促进持续感染的途径中充当 ATP 依赖性信号传导中间体。 结核分枝杆菌对全世界的公共卫生构成严重威胁。这种病原体能够在宿主体内建立临床上无症状的持续潜伏感染，随后重新激活导致疾病，这构成了控制结核病的重大挑战。我们的研究表明，缺乏通用应激蛋白（USP）（称为 Rv2623）的结核分枝杆菌突变体无法在动物宿主中建立慢性持续感染。根据受感染动物相对于亲本菌株的细菌生长、病理学和死亡率增加的评估，该突变菌株表现出高毒力表型。与这种体内生长调节特性一致，我们证明，当 Rv2623 在分枝杆菌中以高于野生型菌株的水平表达时，可在体外抑制细菌生长。通过生化和生物物理分析（包括 Rv2623 晶体结构），我们表明该 USP 在新型 ATP 结合袋中与 ATP 结合。通过定向诱变研究，我们进一步确定Rv2623调节细菌生长的能力取决于其ATP结合能力。我们的数据强烈表明，Rv2623是调节结核分枝杆菌进入慢性持续生长期的关键成分，因此为结核病休眠提供了宝贵的见解，并为开发新型抗结核疗法揭示了新的机会。
Tuberculous latency and reactivation play a significant role in the pathogenesis of tuberculosis, yet the mechanisms that regulate these processes remain unclear. The Mycobacterium tuberculosis universal stress protein (USP) homolog, rv2623, is among the most highly induced genes when the tubercle bacillus is subjected to hypoxia and nitrosative stress, conditions thought to promote latency. Induction of rv2623 also occurs when M. tuberculosis encounters conditions associated with growth arrest, such as the intracellular milieu of macrophages and in the lungs of mice with chronic tuberculosis. Therefore, we tested the hypothesis that Rv2623 regulates tuberculosis latency. We observed that an Rv2623-deficient mutant fails to establish chronic tuberculous infection in guinea pigs and mice, exhibiting a hypervirulence phenotype associated with increased bacterial burden and mortality. Consistent with this in vivo growth-regulatory role, constitutive overexpression of rv2623 attenuates mycobacterial growth in vitro. Biochemical analysis of purified Rv2623 suggested that this mycobacterial USP binds ATP, and the 2.9-Å-resolution crystal structure revealed that Rv2623 engages ATP in a novel nucleotide-binding pocket. Structure-guided mutagenesis yielded Rv2623 mutants with reduced ATP-binding capacity. Analysis of mycobacteria overexpressing these mutants revealed that the in vitro growth-inhibitory property of Rv2623 correlates with its ability to bind ATP. Together, the results indicate that i) M. tuberculosis Rv2623 regulates mycobacterial growth in vitro and in vivo, and ii) Rv2623 is required for the entry of the tubercle bacillus into the chronic phase of infection in the host; in addition, iii) Rv2623 binds ATP; and iv) the growth-regulatory attribute of this USP is dependent on its ATP-binding activity. We propose that Rv2623 may function as an ATP-dependent signaling intermediate in a pathway that promotes persistent infection. Mycobacterium tuberculosis poses serious threats to public health worldwide. The ability of this pathogen to establish in the host a clinically silent, persistent latent infection that can subsequently reactivate to cause diseases constitutes a major challenge in controlling tuberculosis. Our study showed that an M. tuberculosis mutant that is deficient in a universal stress protein (USP) designated Rv2623 fails to establish a chronic persistent infection in animal hosts. The mutant strain exhibits a hypervirulent phenotype as assessed by increased bacillary growth, pathology, and mortality in infected animals relative to the parental strain. Consistent with this in vivo growth-regulating attribute, we demonstrated that Rv2623, when expressed in mycobacteria at levels higher than that of the wild-type strain, retards bacterial growth in vitro. Using biochemical and biophysical analyses, including the Rv2623 crystal structure, we showed that this USP binds to ATP within a novel ATP-binding pocket. Through targeted mutagenesis studies, we further determined that the ability of Rv2623 to regulate bacillary growth is dependent on its ATP-binding capacity. Our data strongly suggest Rv2623 as a critical component that regulates the entry of M. tuberculosis into a chronic persistent growth phase, and therefore provide valuable insight into tuberculous dormancy and uncover new opportunities for the development of novel anti-tuberculous therapies.
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影响因子：
3.1
作者：
Chackerian, AA;Perera, TV;Behar, SM
通讯作者：
Behar, SM
影响因子：
3.8
作者：
Liu, Wen-Tssann;Karavolos, Michail H.;Khan, C. M. Anjam
通讯作者：
Khan, C. M. Anjam
DOI：
10.1046/j.1432-1033.2002.02978.x
发表时间：
2002-06-01
期刊：
EUROPEAN JOURNAL OF BIOCHEMISTRY
影响因子：
--
作者：
Bochkareva, ES;Girshovich, AS;Bibi, E
通讯作者：
Bibi, E
影响因子：
2.8
作者：
Bardarov, S;Bardarov, S;Jacobs, WR
通讯作者：
Jacobs, WR
影响因子：
3.6
作者：
NYSTROM, T;NEIDHARDT, FC
通讯作者：
NEIDHARDT, FC