Biomimetic Virulomics for Capture and Identification of Cell-Type Specific EffectorProteins

用于捕获和鉴定细胞类型特异性效应蛋白的仿生病毒组学

• 批准号: 10092942
• 负责人: David J Gonzalez
• 金额: $ 19.72万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-01 至 2023-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10092942
• 关键词: Affinity; Bacterial Infections; Bacterial Proteins; Binding; Biological Models; Biology; Biomimetics; Catalogs; Cells; Communities; Data; Detection; Development; Disease; Erythrocytes; Functional disorder; Genes; Genome; Genus staphylococcus; Goals; Hospitals; Human; Immunity; In Vitro; Individual; Infection; Knowledge; Mass Spectrum Analysis; Measures; Microbiological Techniques; Molecular; Morbidity - disease rate; Mus; Nature; Paper; Pathogenesis; Pathogenicity; Property; Proteins; Proteome; Proteomics; Publishing; Reporting; Resources; Seeds; Skin; Skin Tissue; Soft Tissue Infections; Specificity; Staphylococcus aureus; System; Tissues; Virulence; Virulence Factors; Work; base; cell type; global health; human pathogen; in vivo; innovation; insight; keratinocyte; macrophage; mortality; mouse model; nano; novel; pathogen; pathogenic bacteria; predictive tools; skin lesion; tool

PROJECT SUMMARY Bacterial pathogens remain a major global health concern with associated high morbidity and mortality rates. Within host−pathogen interactions, virulence is governed by biomolecules produced by the pathogen that target different tissues during infection. Identification of effector proteins with cell-type specificity would be paramount in understanding the pathophysiology associated with infection. However, the identification of relevant protein effectors, particularly at the protein-level, has proven to be technically challenging. We sought to overcome the hurdles associated with direct identification of protein effectors by interfacing multiplexed quantitative proteomics with cell-type specific affinity capture to yield an enrichment workflow, termed Biomimetic Virulomics (BV) [Lapek et al., 2017; ACS Nano]. Our published and new preliminary data demonstrate that the pairing of biomimetics and quantitative proteomics is a powerful avenue to capture key protein effectors involved in virulence. Based on our current data we hypothesize that: 1) BV is a tool that can define host cell-type specificity of bacterial proteins; 2) the BV platform is amenable to an in vivo mouse model of infection; 3) cell-type specificity can be used as a ranking measure to target proteins for study. Herein, we will apply BV to study the proteome of the important human pathogen, Staphylococcus aureus. In Aim 1, we will define the segments of the S. aureus proteome with specificity towards human skin cells. Microbiology techniques will then be used to functionally study a subset of the BV-captured proteins in vitro. This aim is significant given S. aureus remains the leading cause of skin and soft tissue infections in the US. In Aim 2, we will seed BV to an in vivo system in order to capture S. aureus protein effector with specificity towards macrophages in a live mouse. Traditionally, in vivo, identification of bacterial proteomes in the host background has proven to be difficult. Thus, the use of BV to capture and identify bacterial-derived proteins effectors in vivo is highly innovative. Notably, this proposal will focus on characterizing proteins of unknown function. How can we expect to fully understand S aureus infection biology if nearly half of the proteins encoded in its genome are of unknown function? This proposal will be a step forward in filling this critical gap in knowledge. The scientific premise of this study is based on a powerful tool for capturing protein effector with host cell specificity directly at the protein level, a notion that was highlighted in a perspective independently written on the BV platform [Distler et al., 2017; ACS Nano]. Together, this transformative work will serve as a powerful resource and hypothesis-generating tool for host-pathogen studies from an as-yet unattained host cell-specific perspective.

项目概要 细菌病原体仍然是全球主要的健康问题，具有较高的发病率和死亡率。 在宿主与病原体的相互作用中，毒力由病原体产生的生物分子控制， 感染期间针对不同的组织。具有细胞类型特异性的效应蛋白的鉴定将是 对于理解与感染相关的病理生理学至关重要。然而，鉴定 相关的蛋白质效应器，特别是在蛋白质水平上，已被证明在技术上具有挑战性。我们寻求 通过连接多重连接来克服与直接鉴定蛋白质效应子相关的障碍 定量蛋白质组学与细胞类型特异性亲和力捕获产生富集工作流程，称为 仿生病毒组学 (BV) [Lapek 等，2017； ACS纳米]。我们发布的和新的初步数据 证明仿生学和定量蛋白质组学的配对是捕获关键的有力途径 参与毒力的蛋白质效应子。根据我们当前的数据，我们假设：1）BV 是一种工具，可以 定义细菌蛋白质的宿主细胞类型特异性； 2) BV平台适用于体内小鼠模型 感染； 3) 细胞类型特异性可用作研究目标蛋白质的排名指标。在此，我们 将应用 BV 研究人类重要病原体金黄色葡萄球菌的蛋白质组。在目标 1 中，我们 将定义对人类皮肤细胞具有特异性的金黄色葡萄球菌蛋白质组片段。微生物学 然后，技术将用于在体外对 BV 捕获的蛋白质的子集进行功能研究。这个目标是 鉴于金黄色葡萄球菌仍然是美国皮肤和软组织感染的主要原因，这一点意义重大。在目标 2 中，我们 将 BV 接种到体内系统，以捕获具有特异性的金黄色葡萄球菌蛋白效应子 活小鼠的巨噬细胞。传统上，体内鉴定宿主背景中的细菌蛋白质组 事实证明这很困难。因此，使用 BV 捕获和鉴定体内细菌衍生的蛋白质效应子 具有高度的创新性。值得注意的是，该提案将侧重于表征未知功能的蛋白质。怎么可以 如果金黄色葡萄球菌基因组中编码的近一半蛋白质是 未知功能？该提案将在填补这一关键知识空白方面向前迈出一步。科学的 这项研究的前提是基于直接捕获具有宿主细胞特异性的蛋白质效应物的强大工具 在蛋白质水平上，BV 平台上独立撰写的观点中强调了这一概念 [迪斯特勒等人，2017； ACS纳米]。总之，这项变革性的工作将成为强大的资源和 从尚未实现的宿主细胞特异性角度进行宿主病原体研究的假设生成工具。

项目成果

Group A Streptococcal S Protein Utilizes Red Blood Cells as Immune Camouflage and Is a Critical Determinant for Immune Evasion.

A 组链球菌 S 蛋白利用红细胞作为免疫伪装，是免疫逃避的关键决定因素。

• DOI: 10.1016/j.celrep.2019.11.001
• 发表时间: 2019
• 期刊: Cell reports
• 影响因子: 8.8
• 作者: Wierzbicki,IgorH;Campeau,Anaamika;Dehaini,Diana;Holay,Maya;Wei,Xiaoli;Greene,Trever;Ying,Man;Sands,JennaS;Lamsa,Anne;Zuniga,Elina;Pogliano,Kit;Fang,RonnieH;LaRock,ChristopherN;Zhang,Liangfang;Gonzalez,DavidJ
• 通讯作者: Gonzalez,DavidJ

Unique virulence role of post-translocational chaperone PrsA in shaping Streptococcus pyogenes secretome.

• DOI: 10.1080/21505594.2021.1982501
• 发表时间: 2021-12
• 期刊: Virulence
• 影响因子: 5.2
• 作者: Wu ZY;Campeau A;Liu CH;Gonzalez DJ;Yamaguchi M;Kawabata S;Lu CH;Lai CY;Chiu HC;Chang YC
• 通讯作者: Chang YC