Structural Basis of Self-recognition in Hydroid Allorecognition Proteins

水螅同种异体识别蛋白自我识别的结构基础

• 批准号: 1557339
• 负责人: Matthew Nicotra
• 金额: $ 72.67万
• 依托单位: University of Pittsburgh
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-03-01 至 2020-02-29
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1557339&HistoricalAwards=false
• 关键词: Structural; Basis; Self; recognition; Hydroid

Many organisms across the tree of life can distinguish between themselves and other members of their species via cell-cell contact. In vertebrates, this process controls graft rejection and auto-immune responses. In colonial invertebrates -- animals like marine sponges, corals, and sea squirts -- this ability is called allorecognition. Allorecognition controls whether colonies aggressively compete for space or peacefully coexist when they encounter one another as they grow. Despite the biological significance of this phenomenon, the genes and proteins that underlie it remain poorly understood. In this project, the investigators will study the molecular basis of allorecognition in a cnidarian, Hydractinia symbiolongicarpus. Recently, two genes controlling allorecognition in Hydractinia have been identified. The investigators will determine how the proteins encoded by these genes allow Hydractinia to properly distinguish self from non-self. To do this, the investigators will combine biochemical experiments with computer modeling and x-ray crystallography to build a structural model of the proteins, which will reveal how they enable Hydractinia to discriminate between self and non-self. This knowledge will fill a significant gap in our understanding of invertebrate allorecognition and form the basis of future work to investigate how these phenomena evolve. As part of their work, the investigators will train two undergraduates, two graduate students, and one postdoc in protein biochemistry and structure determination. They will provide opportunities for three high school students to conduct independent research projects in their laboratories. They will also develop and implement an educational program about Hydractinia for elementary school students in southwestern Pennsylvania.In Hydractinia, two allorecognition proteins, Alr1 and Alr2, discriminate between self and non-self. Preliminary studies indicate these proteins engage in isoform-specific binding across cell membranes. This binding is hypothesized to be a general feature of both proteins and to be mediated by trans interactions between immunoglobulin superfamily-like (IgSF-like) domains in their extracellular regions and the formation of disulphide-linked cis multimers. Aim 1 will identify segments in the extracellular regions of each protein that control binding specificity and test whether the proteins form cis multimers via disulphide bonds. Aim 2 will identify amino acid positions within these segments that determine their binding profiles and use computational modeling to explore hypothesized binding mechanisms. Together, Aims 1 and 2 will be used to create a structural model of isoform-specific binding. Aim 3 will validate this model by determining the crystal structures of the extracellular domains of Alr1 and Alr2. Our team combines three PIs with expertise in invertebrate allorecognition, computational biology, and x-ray crystallography. The expected outcome of this project will be the first mechanistic understanding of how allorecognition proteins discriminate between self and non-self in any invertebrate. Comparison of this structural mechanism with those in other self-recognition systems will reveal which aspects of molecular recognition are unique to Hydractinia and which are shared with other species. The data generated in this project will guide future projects investigating how sequence evolution at allorecognition genes leads to novel allorecognition specificities and how signaling through the allorecognition proteins regulates allorecognition responses.

生命之树上的许多生物都可以通过细胞间的接触来区分自己和其他物种。在脊椎动物中，这一过程控制着移植排斥和自身免疫反应。 在群体无脊椎动物中--像海绵、珊瑚和海蜇这样的动物--这种能力被称为异源识别。同种异体识别控制着殖民地在成长过程中是积极争夺空间，还是和平共处。尽管这一现象具有生物学意义，但其背后的基因和蛋白质仍然知之甚少。在这个项目中，研究人员将研究刺胞动物Hydractinia symbiolongarpus中同种识别的分子基础。最近，已经确定了两个基因控制异源识别Hydractinia。研究人员将确定这些基因编码的蛋白质如何允许Hydractinia正确区分自我和非自我。为了做到这一点，研究人员将联合收割机生化实验与计算机建模和X射线晶体学相结合，以建立蛋白质的结构模型，这将揭示它们如何使Hydractinia区分自我和非自我。这些知识将填补我们对无脊椎动物同种异体识别的理解中的一个重要空白，并为未来研究这些现象如何演变奠定基础。作为他们工作的一部分，研究人员将在蛋白质生物化学和结构测定方面培训两名本科生，两名研究生和一名博士后。他们将为三名高中生提供在实验室进行独立研究项目的机会。他们还将为宾夕法尼亚州西南部的小学生开发和实施一项关于Hydractinia的教育计划。Hydractinia中有两种同种识别蛋白Alr 1和Alr 2，可以区分自我和非自我。初步研究表明，这些蛋白质参与跨细胞膜的同种型特异性结合。假设这种结合是这两种蛋白质的一般特征，并且通过其细胞外区域中的免疫球蛋白超家族样（IgSF样）结构域之间的反式相互作用和二硫键连接的顺式多聚体的形成来介导。目标1将确定在每种蛋白质的胞外区域中控制结合特异性的片段，并测试蛋白质是否通过二硫键形成顺式多聚体。目标2将确定这些片段内的氨基酸位置，决定其结合概况，并使用计算建模来探索假设的结合机制。目的1和2将一起用于创建异构体特异性结合的结构模型。目的3将通过确定Alr 1和Alr 2的胞外结构域的晶体结构来验证该模型。我们的团队结合了三个PI与无脊椎动物同种异体识别，计算生物学和X射线晶体学的专业知识。该项目的预期成果将是第一次从机制上了解同种识别蛋白如何在任何无脊椎动物中区分自我和非自我。这种结构机制与其他自我识别系统的比较将揭示哪些方面的分子识别是独特的Hydractinia和其他物种共享。在这个项目中产生的数据将指导未来的项目调查如何在异源识别基因序列进化导致新的异源识别特异性和信号如何通过异源识别蛋白调节异源识别反应。