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.

项目摘要 与相关的高发病率和死亡率相关的细菌病原体仍然是全球主要的健康问题。 在宿主病原体相互作用中，病毒性受病原体产生的生物分子的控制 在感染过程中靶向不同的组织。鉴定具有细胞类型特异性的效应蛋白将是 了解与感染相关的病理生理学的重要性。但是，识别 相关的蛋白质作用，尤其是在蛋白质水平上，已被证明在技术上具有挑战性。我们寻求 克服与直接识别蛋白质效应相关的障碍，通过接口多路复用 具有细胞类型特异性亲和力捕获的定量蛋白质组学以产生富集工作流程，称为 仿生毒素学（BV）[Lapek等，2017; ACS纳米]。我们发布的新初步数据 证明仿生学和定量蛋白质组学的配对是捕获密钥的强大途径 病毒涉及的蛋白质作用。基于我们当前的数据，我们假设：1）BV是一个可以 定义细菌蛋白的宿主细胞类型特异性； 2）BV平台适合体内鼠标模型 感染； 3）细胞类型特异性可以用作靶蛋白进行研究的排名度量。这里，我们 将应用BV研究重要人类病原体金黄色葡萄球菌的蛋白质组。在AIM 1中，我们 将定义金黄色葡萄球菌蛋白质组的段，对人皮细胞的特异性。微生物学 然后，技术将用于在体外研究BV捕获蛋白的子集。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。在AIM 2中，我们 将BV播种到体内系统中，以捕获S.金黄色葡萄球菌蛋白效应子，特异性 活鼠中的巨噬细胞。传统上，在体内，在宿主背景中鉴定细菌蛋白 事实证明很困难。那就是使用BV捕获和鉴定体内细菌衍生的蛋白质作用 具有高度创新性。值得注意的是，该建议将集中于表征未知功能的蛋白质。怎么可能 如果几乎一半在其基因组中编码的蛋白质是 功能未知？该提案将是填补知识的关键空白的一步。科学 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器的强大工具 在蛋白质水平上，在BV平台上独立写的透视图中突出了一个概念 [Distler等，2017； ACS纳米]。这项变革性的工作将成为一种强大的资源， 从尚未获得的宿主细胞特异性的角度来看，用于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