Signal perception and transduction regulating Giardia cyst formation

信号感知和转导调节贾第鞭毛虫包囊形成

• 批准号: 10707172
• 负责人: Alexander Richard Paredez
• 金额: $ 50.74万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-19 至 2027-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10707172
• 关键词: Amino Acid Transporter; Amino Acids; Awareness; Biology; Biosensor; Cell Cycle Arrest; Cell membrane; Cells; Cellular Assay; Cellular biology; Cessation of life; Cholesterol; Citrulline; Clinical Treatment; Complement; Complex; Cues; Cyclic AMP; Cyst; Cytoskeleton; Data; Defect; Detection; Development; Developmental Biology; Diarrhea; Effectiveness; Energy-Generating Resources; Environment; Exposure to; Family; Foundations; Free Radicals; G-Protein-Coupled Receptors; Gastrointestinal Diseases; Generations; Genetic Transcription; Giardia; Giardia lamblia; Giardiasis; Goals; Growth; Guanosine Triphosphate Phosphohydrolases; Homologous Gene; Image; Incidence; Infection; Intervention; Intestines; Knowledge; Life Cycle Stages; Malabsorption Syndromes; Mastigophora; Membrane; Membrane Proteins; Metabolic; Molecular; Molecular Conformation; Mutate; Names; Nutrient; Opsin; Oral; Parasites; Pathway interactions; Perception; Persons; Pharmaceutical Preparations; Phosphotransferases; Process; Production; Proliferating; Proteins; Regulation; Reporting; Resistance; Risk; Role; Shapes; Signal Transduction; Starvation; Stimulus; Testing; Translating; Work; clinically relevant; design; detection of nutrient; drug action; gain of function; gastrointestinal; genetic analysis; gut colonization; improved; insight; interest; knock-down; loss of function; nanoluciferase; novel; overexpression; phosphoproteomics; prevent; programs; protein protein interaction; public health relevance; receptor; response; rho; rho GTP-Binding Proteins; sensor; side effect; spatiotemporal; therapeutic development; trafficking; transmission process; treatment strategy; uptake

PROJECT SUMMARY/ABSTRACT Giardia lamblia, the causative agent of giardiasis, leads to gastrointestinal disorders, long-term growth defects and even death. Estimates place worldwide incidence at over 200 million symptomatic cases per year. Of concern, up to 20% of giardiasis cases are resistant to front-line clinical treatments, and resistance to all major antigiardial drugs has been reported. In addition to their increasing lack of effectiveness, front-line and second- line nitro drugs act intracellularly via non-specific free radical generation, and therefore have a high incidence of negative side effects. There is a critical need to better understand the basic biology of the parasite in order to ultimately design improved intervention and treatment strategies. Flagellated trophozoites proliferate to colonize the intestine where cues including cholesterol starvation and increased pH at the end of the intestinal tract promote terminal differentiation into cysts that detach for transmission. While current treatments target the flagellated trophozoites, encystation could be exploited to clear Giardia infections. The regulation of encystation is poorly understood across the diversity of encysting parasites; thus, studies focused on the encystation pathway are of fundamental cell and developmental biology interest, as well as profound clinical relevance. The aim of this proposal is to understand how encystation cues are perceived and transduced to initiate the encystation program. Our preliminary studies indicate that elevated cAMP is necessary to activate encystation. Our preliminary studies identified a role for Giardia’s sole Rho family GTPase, GlRac, in the regulation of cAMP. Protein-protein interaction studies with GlRac identified a previously uncharacterized seven- transmembrane PQ-loop protein that belongs to the TOG (transporter/opsin/GPCR) superfamily. Knockdown of this protein results in increased encystation indicating that it negatively regulates encystation. We named this protein EncystR for its role in encystation. EncystR localizes to the plasma membrane in actively growing cells, but exposure to encystation stimuli (high pH and cholesterol depletion), causes rapid internalization. EncystR internalization requires GlRac, supporting a functional relationship between these proteins. EncystR negatively regulates encystation through control of cAMP levels, but the mechanism remains to be determined. Related PQ loop proteins are multi-function amino acid transporter-receptors that are known to be pH responsive and can regulate development. EncystR is an exciting new entry point into uncovering the regulation of Giardia’s developmental program. The overall goal of this project is to delineate the roles of EncystR and GlRac in regulating cAMP and the role of cAMP in eliciting the encystation program. This work will establish a framework for understanding the regulation of encystation from signal detection to encystation. Ultimately, we aim to identify the means to short circuit the normal encystation program such that cell cycle arrest and cytoskeletal disassembly can be activated without producing a protective cyst wall. This would clear infections without the risk of transmission.

项目摘要/摘要 贾第鞭毛虫病的致病药物贾第鞭毛虫（Giardia Lamblia）导致胃肠道疾病，长期生长 缺陷甚至死亡。估计全球事件每年有超过2亿个症状案件。 令人担忧的是，多达20％ 已经报道了抗生素药物。除了越来越缺乏效力，前线和第二 线氮药通过非特异性自由基产生细胞内作用，因此 负面影响。迫切需要更好地了解寄生虫的基本生物学 最终设计改进的干预和治疗策略。鞭毛的滋养体扩散到定殖 提示包括胆固醇饥饿和肠道结束时pH增加的肠道 促进末端分化为脱离传播的囊肿。而目前的治疗目标是 可以利用旗帜的滋养体，百科全书清除贾第鞭毛虫感染。百货公司的调节 在寄生虫的多样性中，人们对众所周知；因此，研究重点是百货公司 途径具有基本细胞和发育生物学的兴趣，以及深远的临床相关性。 该提案的目的是了解如何感知和翻译百科全书来启动 encystation程序。我们的初步研究表明，高架cAMP是激活百科全书所必需的。 我们的初步研究确定了Giardia唯一的Rho家族GTPase Glrac的作用 营。蛋白质 - 蛋白质相互作用研究与GLRAC确定了先前未表征的七个 属于TOG（转运蛋白/OPSIN/GPCR）超家族的跨膜PQ-LOOP蛋白。击倒 该蛋白质导致广告增加，表明它对综合性进行负调节。我们命名了 蛋白质encystr在百科全中的作用。 encystr在积极生长的细胞中定位于质膜， 但是暴露于百科全刺激（高pH和胆固醇消耗）会导致快速内在化。 encystr 内部化需要GLRAC，支持这些蛋白质之间的功能关系。元素负面 通过控制营地的水平来调节百货公司，但该机制仍有待确定。有关的 PQ环蛋白是多功能氨基酸转运蛋白受体，已知是pH响应性的，并且 可以规范发展。 EncoSTR是一个令人兴奋的新入口处，可以揭示贾第鞭毛虫的调节 发展计划。 该项目的总体目标是描述Encostr和Glrac在调节营地和 CAMP在引发百科全计划中的作用。这项工作将建立理解的框架 从信号检测到百货公司的百货公司调节。最终，我们旨在确定 短路正常的百货程序，使细胞周期停滞和细胞骨架拆卸可以是 激活而无需产生受保护的囊肿壁。这将清除感染而不会出现传播风险。