Department of Microbiology and Molecular Genetics, Oklahoma State University, Stillwater, Oklahoma, USA.
Department of Microbiology and Molecular Genetics, Oklahoma State University, Stillwater, Oklahoma, USA
Appl Environ Microbiol. 2021 Apr 27;87(10). doi: 10.1128/AEM.00061-21.
is an opportunistic pathogen causing life-threatening infections. Previously, we showed that elevated calcium (Ca) levels increase the production of virulence factors in In an effort to characterize the Ca regulatory network, we identified a -egulated β-ropeller protein, CarP, and showed that expression of the encoding gene is controlled by the Ca-regulated two-component system CarSR. Here, by using a model, we showed that CarP plays a role in regulating virulence. By using transcriptome sequencing (RNA-Seq), reverse transcription (RT)-PCR, quantitative RT-PCR (RT-qPCR), and promoter fusions, we determined that is transcribed into at least two transcripts and regulated by several bacterial and host factors. The transcription of is elevated in response to Ca in cystic fibrosis isolates and PAO1 laboratory strain. Elevated Fe also induces The simultaneous addition of Ca and Fe increased the promoter activity synergistically, which requires the presence of CarR. analysis of the intergenic sequence upstream of predicted recognition sites of RhlR/LasR, OxyR, and LexA, suggesting regulation by quorum sensing (QS) and oxidative stress. In agreement, the promoter was activated in response to stationary-phase PAO1 supernatant and required the presence of elevated Ca and CarR but remained silent in the triple mutant lacking , and synthases. We also showed that transcription is regulated by oxidative stress and that CarP contributes to Ca-dependent HO tolerance. The multifactorial regulation of suggests that CarP plays an important role in adaptations to host environments. is a human pathogen causing life-threatening infections. It is particularly notorious for its ability to adapt to diverse environments within the host. Understanding the signals and the signaling pathways enabling adaptation is imperative for developing effective therapies to treat infections caused by this organism. One host signal of particular importance is calcium. Previously, we identified a component of the calcium-signaling network, CarP, whose expression is induced by elevated levels of calcium. Here, we show that plays an important role in virulence and is upregulated in strains isolated from sputa of patients with cystic fibrosis. We also identified several bacterial and host factors that regulate the transcription of Such multifactorial regulation highlights the interconnectedness between regulatory circuits and, together with the pleotropic effect of CarP on virulence, suggests the importance of this protein in adaptations to the host.
铜绿假单胞菌是一种机会性病原体，可导致危及生命的感染。此前，我们发现高水平的钙（Ca）会增加毒力因子的产生。为了描述 Ca 调控网络，我们鉴定了一个受 Ca 调控的二组分系统 CarSR 调控的 CarP。通过使用一个模型，我们发现 CarP 在调节铜绿假单胞菌毒力方面发挥作用。通过使用转录组测序（RNA-Seq）、反转录（RT）-PCR、定量 RT-PCR（RT-qPCR）和启动子融合，我们确定 至少转录成两种转录本，并受到多种细菌和宿主因子的调节。在囊性纤维化分离株和 PAO1 实验室株中，Ca 响应时转录水平升高。Fe 水平升高也会诱导 。Ca 和 Fe 同时添加会协同增加 启动子活性，这需要 CarR 的存在。对 上游基因间序列的 分析预测了 RhlR/LasR、OxyR 和 LexA 的识别位点，提示受群体感应（QS）和氧化应激调节。与预测一致，PAO1 静止期上清液响应激活 启动子，需要高水平 Ca 和 CarR 的存在，但在缺乏 、 和 合成酶的三突变体中保持沉默。我们还表明， 转录受氧化应激调节，CarP 有助于铜绿假单胞菌对 Ca 依赖性 HO 耐受。 受多种因素调节，表明 CarP 在铜绿假单胞菌适应宿主环境方面发挥重要作用。铜绿假单胞菌是一种引起危及生命感染的人类病原体。它特别臭名昭著的是它能够在宿主内的不同环境中适应。了解使 适应的信号和信号通路对于开发治疗该生物体引起的感染的有效疗法至关重要。一种特别重要的宿主信号是钙。此前，我们鉴定了 钙信号网络的一个组成部分 CarP，其表达受钙水平升高的诱导。在这里，我们表明 对毒力很重要，并且在从囊性纤维化患者痰液中分离的 菌株中上调。我们还鉴定了几个调节 的转录的细菌和宿主因子。这种多因素调节突出了调控回路之间的相互关联性，再加上 CarP 对毒力的多效性影响，表明该蛋白在 适应宿主方面的重要性。
