Balasubramanian Divya, Ohneck Elizabeth A, Chapman Jessica, Weiss Andy, Kim Min Kyung, Reyes-Robles Tamara, Zhong Judy, Shaw Lindsey N, Lun Desmond S, Ueberheide Beatrix, Shopsin Bo, Torres Victor J
Department of Microbiology, New York University School of Medicine, New York, New York, USA.
Proteomics Resource Center, Office of Collaborative Science, New York University School of Medicine, New York, New York, USA.
mBio. 2016 Jun 21;7(3):e00818-16. doi: 10.1128/mBio.00818-16.
Staphylococcus aureus is a formidable human pathogen that uses secreted cytolytic factors to injure immune cells and promote infection of its host. Of these proteins, the bicomponent family of pore-forming leukocidins play critical roles in S. aureus pathogenesis. The regulatory mechanisms governing the expression of these toxins are incompletely defined. In this work, we performed a screen to identify transcriptional regulators involved in leukocidin expression in S. aureus strain USA300. We discovered that a metabolic sensor-regulator, RpiRc, is a potent and selective repressor of two leukocidins, LukED and LukSF-PV. Whole-genome transcriptomics, S. aureus exoprotein proteomics, and metabolomic analyses revealed that RpiRc influences the expression and production of disparate virulence factors. Additionally, RpiRc altered metabolic fluxes in the trichloroacetic acid cycle, glycolysis, and amino acid metabolism. Using mutational analyses, we confirmed and extended the observation that RpiRc signals through the accessory gene regulatory (Agr) quorum-sensing system in USA300. Specifically, RpiRc represses the rnaIII promoter, resulting in increased repressor of toxins (Rot) levels, which in turn negatively affect leukocidin expression. Inactivation of rpiRc phenocopied rot deletion and increased S. aureus killing of primary human polymorphonuclear leukocytes and the pathogenesis of bloodstream infection in vivo. Collectively, our results suggest that S. aureus senses metabolic shifts by RpiRc to differentially regulate the expression of leukocidins and to promote invasive disease.
The bicomponent pore-forming leukocidins play pivotal roles in the ability of S. aureus to kill multiple host immune cells, thus enabling this pathogen to have diverse tissue- and species-tropic effects. While the mechanisms of leukocidin-host receptor interactions have been studied in detail, the regulatory aspects of leukocidin expression are less well characterized. Moreover, the expression of the leukocidins is highly modular in vitro, suggesting the presence of regulators other than the known Agr, Rot, and S. aureus exoprotein pathways. Here, we describe how RpiRc, a metabolite-sensing transcription factor, mediates the repression of two specific leukocidin genes, lukED and pvl, which in turn has complex effects on the pathogenesis of S. aureus Our findings highlight the intricacies of leukocidin regulation by S. aureus and demonstrate the involvement of factors beyond traditional virulence factor regulators.
金黄色葡萄球菌是一种强大的人类病原体，它利用分泌的溶细胞因子损伤免疫细胞并促进其在宿主中的感染。在这些蛋白质中，双组分成孔白细胞毒素家族在金黄色葡萄球菌的发病机制中起关键作用。这些毒素表达的调控机制尚未完全明确。在这项研究中，我们进行了一项筛选，以鉴定参与金黄色葡萄球菌USA300菌株白细胞毒素表达的转录调节因子。我们发现，一种代谢传感调节因子RpiRc是两种白细胞毒素LukED和LukSF-PV的有效且选择性的阻遏物。全基因组转录组学、金黄色葡萄球菌外蛋白组学和代谢组学分析表明，RpiRc影响不同毒力因子的表达和产生。此外，RpiRc改变了三羧酸循环、糖酵解和氨基酸代谢中的代谢通量。通过突变分析，我们证实并扩展了RpiRc通过USA300中的辅助基因调节（Agr）群体感应系统发出信号的观察结果。具体而言，RpiRc抑制rnaIII启动子，导致毒素阻遏物（Rot）水平升高，进而对白细胞毒素表达产生负面影响。rpiRc的失活模拟了rot缺失，并增加了金黄色葡萄球菌对原代人多形核白细胞的杀伤作用以及体内血流感染的发病机制。总体而言，我们的结果表明，金黄色葡萄球菌通过RpiRc感知代谢变化，以差异调节白细胞毒素的表达并促进侵袭性疾病。
双组分成孔白细胞毒素在金黄色葡萄球菌杀死多种宿主免疫细胞的能力中起关键作用，从而使这种病原体具有多种组织和物种嗜性效应。虽然白细胞毒素与宿主受体相互作用的机制已得到详细研究，但白细胞毒素表达的调控方面的特征尚不明确。此外，白细胞毒素的表达在体外具有高度模块化，这表明除了已知的Agr、Rot和金黄色葡萄球菌外蛋白途径外，还存在其他调节因子。在这里，我们描述了代谢物传感转录因子RpiRc如何介导对两个特定白细胞毒素基因lukED和pvl的抑制，这反过来又对金黄色葡萄球菌的发病机制产生复杂影响。我们的发现突出了金黄色葡萄球菌对白细胞毒素调控的复杂性，并证明了除传统毒力因子调节因子之外的其他因子的参与。
