Tryptophan derivatives in EHEC pathogenesis

EHEC 发病机制中的色氨酸衍生物

• 批准号: 10219567
• 负责人: VANESSA SPERANDIO
• 金额: $ 62.7万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-01 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10219567
• 关键词: Anabolism; Animal Model; Bacteria; Behavioral; Binding; Blood flow; Brain; Cells; Chemicals; Chlorides; Citrobacter rodentium; Colitis; Colon; Communication; Cues; Disease; Engineering; Enterochromaffin Cells; Enterocytes; Environment; Enzymes; Epithelial Cells; Escherichia coli EHEC; Excision; Fluoxetine; Gastrointestinal Diseases; Gastrointestinal tract structure; Gene Expression; Genes; Genetic Transcription; Grant; Human; Hyperplasia; Immune response; Indoles; Infection; Intestinal Mucosa; Intestines; Knockout Mice; Lamina Propria; Lesion; Mediating; Membrane; Modeling; Mus; Muscle Contraction; Names; Neurotransmitters; Nucleic Acid Regulatory Sequences; Orphan; Pathogenesis; Pathogenicity Island; Pharmacology; Phosphoric Monoester Hydrolases; Phosphotransferases; Physiology; Potassium; Process; Prognosis; Receptor Signaling; Regulation; Role; Serotonin; Serotonin Agonists; Shiga Toxin; Signal Pathway; Signal Transduction; Signaling Molecule; Smooth Muscle; Source; Study models; Tissues; Tryptophan; Tryptophan 5-monooxygenase; Tryptophanase; Virulence; arm; enteric pathogen; gastrointestinal; gut-brain axis; host microbiota; host-microbe interactions; in vivo; in vivo Model; inflammatory disease of the intestine; inhibitor/antagonist; intestinal barrier; intestinal epithelium; microbiota; mouse model; novel; pathogen; protein-histidine kinase; receptor; renal damage; response; reuptake; serotonin receptor; transcriptome

PROJECT SUMMARY The colon contains tryptophan derivatives such as indole, which is a microbiota-derived signaling molecule, and the host-derived serotonin neurotransmitter that is primarily synthesized in the GI tract. Indole is also known to be absorbed by host cells and helps strengthen the integrity of the intestinal barrier, being regarded as a beneficial chemical cue within microbial/host interactions. Indole is synthesized by tryptophanase, which is encoded by the tnaA gene. We have shown that the concentration of indole is significantly higher in the lumen of the colon (the compartment where the microbiota resides) compared to colonic tissues (where indole is absorbed by intestinal epithelial cells). Serotonin is synthesized in enterochromaffin cells by the enzyme tryptophan hydroxylase (TpH1). Upon its synthesis, serotonin is released into the lamina propria and is secreted into the lumen. Serotonin signaling in the intestinal mucosa is terminated by removal of serotonin by the serotonin selective reuptake transporter (SERT), which is expressed by epithelial cells. We showed that both serotonin and indole converge to decrease virulence gene expression from enterohemorrhagic E. coli (EHEC) and Citrobacter rodentium, a murine pathogen employed as a surrogate animal model for EHEC. We also identified the bacterial receptor for these signals as CpxA. Upon sensing serotonin and/or indole, CpxA functions primarily as a phosphatase, dephosphorylating itself and CpxR, that activates virulence in its phosphorylated state. Through transcriptome studies we also identified the Indole Sequestering Receptor (Isr), which in the absence of indole directly activates virulence expression. However, in the presence of indole, Isr is no longer able to activate transcription of virulence genes. Using TpH1 pharmacological inhibitors (decrease the levels of serotonin in the gut) and SERT knockout mice (have increased levels of luminal serotonin), we showed that the presence of higher levels of serotonin in the intestine of mice decreased virulence in C. rodentium, while decreased levels of serotonin are conducive to increased pathogenesis. Moreover, we synthetically altered the concentration of indole in the GI tract of mice. This allowed us to assess the role of self-produced versus microbiota-produced indole, and show that decreased indole concentrations promote bacterial pathogenesis, while increased levels of indole decreases bacterial virulence gene expression during murine infection. Altogether, both serotonin and indole decrease virulence of C. rodentium during murine infection. Our studies show that fluctuations in the levels of indole and the serotonin neurotransmitter significantly impact disease prognosis. However several questions regarding this exquisite signaling regulation of bacterial virulence remain unanswered. Consequently the specific aims of this grant are: Aim 1. Define the CpxA/CpxR and Isr serotonin/indole signaling cascade. Aim 2. Investigate the intersection of serotonin with endogenous and exogenous indole signaling in bacterial pathogenesis during mammalian infection.

项目总结 结肠含有色氨酸衍生物，如吲哚，这是一种微生物区系衍生的信号分子，以及 宿主衍生的5-羟色胺神经递质，主要在胃肠道合成。吲哚也是已知的 被宿主细胞吸收，有助于加强肠道屏障的完整性，被视为 微生物/宿主相互作用中有益的化学线索。吲哚是由色氨酸酶合成的，它是 由TNAA基因编码。我们已经证明了吲哚在管腔中的浓度要高得多。 结肠(微生物区系所在的隔间)与结肠组织(吲哚 被肠道上皮细胞吸收)。5-羟色胺是在肠嗜铬细胞中通过该酶合成的 色氨酸羟化酶(TPH1)。合成后，5-羟色胺被释放到固有层，并被分泌。 进入管腔。肠粘膜中的5-羟色胺信号通过清除5-羟色胺而终止。 5-羟色胺选择性再摄取转运体(SERT)，由上皮细胞表达。我们证明了这两个 5-羟色胺和吲哚共同作用降低肠出血性大肠杆菌毒力基因表达 和轮状柠檬酸杆菌，一种小鼠病原体，被用作EHEC的替代动物模型。我们也 确定这些信号的细菌受体是CpxA。在感觉到5-羟色胺和/或吲哚时，CpxA 主要作为一种磷酸酶发挥作用，使自身和CpxR去磷酸化，激活其毒力 磷酸化状态。通过转录组研究，我们还鉴定了吲哚隔离受体(ISR)， 在没有吲哚的情况下，它直接激活毒力表达。然而，在吲哚存在的情况下，ISR是 不再能够激活毒力基因的转录。使用TPH1药物抑制剂(减少 肠道中的5-羟色胺水平)和SERT基因敲除小鼠(腔内5-羟色胺水平增加)，我们显示 小鼠肠道中较高水平的5-羟色胺降低了轮齿芽孢杆菌的毒力， 而血清5-羟色胺水平降低则有利于增加发病机制。而且，我们综合起来， 改变小鼠胃肠道中吲哚的浓度。这使我们能够评估自我创造的作用 与微生物群产生的吲哚相比，并表明降低吲哚浓度促进细菌 发病机制，而增加吲哚水平降低细菌毒力基因在小鼠中的表达 感染。总而言之，5-羟色胺和吲哚都可以降低小鼠感染轮齿芽孢杆菌的毒力。我们的 研究表明，吲哚和5-羟色胺神经递质水平的波动对 疾病预后。然而，关于细菌的这种精巧的信号调控的几个问题 致命性仍未得到回答。因此，这笔赠款的具体目标是：目标1.确定CpxA/CpxR ISR 5-羟色胺/吲哚信号转导通路。目的2.研究5-羟色胺与内源性的交集 外源吲哚信号在哺乳动物感染过程中的细菌发病机制中的作用。