Novel Type 1 Pilus Receptors in Pyelonephritis and Recurrent UTI
肾盂肾炎和复发性尿路感染中的新型 1 型菌毛受体
基本信息
- 批准号:10180267
- 负责人:
- 金额:$ 39.38万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2021
- 资助国家:美国
- 起止时间:2021-04-01 至 2026-03-31
- 项目状态:未结题
- 来源:
- 关键词:AcuteAdhesivesAffinityApoptosisAttentionBacteriaBacterial AdhesinsBacterial AdhesionBehaviorBindingBiochemicalBiologicalBladderCRISPR screenCadherin DomainCadherinsCellsClinicalColonCommunitiesConfocal MicroscopyCryoelectron MicroscopyCrystallizationCystitisDuctal EpitheliumEpithelialEpithelial CellsEscherichia coliEscherichia coli AdhesinsEscherichia coli InfectionsExtracellular DomainFamilyFiberGastrointestinal tract structureGeneticGenomeHumanIn VitroInbred C3H MiceInfectionInfective cystitisIntercellular JunctionsInvestigationKidneyKnock-outKnockout MiceLectinLinkMannoseMannosidesMediatingMediator of activation proteinModelingMolecularMolecular ChaperonesMonoclonal AntibodiesMusMutationNephronsPathogenesisPathway interactionsPeptidesPharmacologyPilumPolysaccharidesProcessProteinsPublishingPyelonephritisRecurrenceRoleSeedsSiteSpecific qualifier valueStructureSurfaceSystemTestingTherapeuticTissuesTo specifyTubular formationUrinary tractUrinary tract infectionUrineUropathogenic E. coliUrsidae FamilyVaccinationVariantVirulenceX-Ray Crystallographybacterial communitybasecell typedesmoglein 2experimental studyextracellulargut colonizationhuman tissuein vitro Modelin vivoinhibitor/antagonistintestinal epitheliumkidney infectionmouse modelnovelpathogenreceptorrenal abscessrenal epitheliumresponsesmall moleculesmall molecule inhibitorsugartype 1 fimbriae receptor
项目摘要
PROJECT SUMMARY / ABSTRACT
Bacterial adhesion to the urinary tract epithelium is a critical step in establishing urinary tract infections. During
infection of the mammalian bladder (cystitis), uropathogenic Escherichia coli (UPEC) are well described to
employ type 1 pili, bearing the tip adhesin FimH, to bind oligomannose-decorated uroplakins that coat the
luminal surfaces of superficial bladder epithelial cells. However, less detail is known about host-pathogen
interactions in the kidney that enable initiation of upper-tract UTIs, including pyelonephritis and renal abscess.
We have found that type 1 pili, previously thought to be essential only in cystitis, also mediate establishment of
pyelonephritis and the initiation of renal abscesses in C3H mice. Furthermore, in an in vitro model of UPEC
binding to renal collecting duct epithelium, we identified a candidate renal epithelial receptor for FimH, namely
the mannosylated cell-junctional protein desmoglein-2 (Dsg2). This protein is expressed throughout the
nephron but most highly in collecting duct epithelium, and bears typical N-linked mannose-containing glycans
as well as cadherin family-specific O-linked mannosylation. In this project, we will test the central hypothesis
that desmoglein-2 is an epithelial receptor for FimH that mediates establishment of UPEC pyelonephritis and
can bind FimH in gut and exfoliated bladder. First, we will use multiple genetic and pharmacologic systems to
interrogate the importance of FimH binding to mannosylated Dsg2 in recently published, optimized mouse
models of UPEC pyelonephritis. Among these systems will be newly generated C3H mice carrying renal
epithelial-specific deletion of Dsg2. Next, we will quantify the binding affinity of the FimH lectin domain to the
purified extracellular domain of human DSG2 by SPR, and co-crystallize the relevant FimH and DSG2 domains
to reveal the structural basis for the DSG2-FimH interaction. Controls in these experiments will include
FimHQ133K, which carries a mutation that abrogates mannose binding; mannosides, high-affinity small-molecule
inhibitors of FimH binding; enzymatic pre-treatment of purified protein and kidney tissue sections to eliminate
N- or O-linked glycans; and monoclonal antibodies generated against the key DSG2 peptides mediating
interaction with FimH. Third, desmoglein-2 is also expressed widely on other epithelial cell types (in both
humans and mice), raising the added possibility that it binds FimH in other niches relevant to UTI
pathogenesis. These include the bladder after exfoliation (a rapid response to initial UPEC infection that
eliminates the primary FimH receptor) and the colon (which serves as a UPEC reservoir to seed recurrent
UTI). Therefore, we will use mouse and human tissue sections, an in vivo gut colonization model, and
additional new conditional Dsg2 knockout mice to investigate whether Dsg2 can serve as a FimH receptor in
these tissues. At the conclusion of these studies, we will have specified a novel receptor for type 1 pili,
elucidated the structural basis of FimH interaction with desmoglein-2, and defined the functional roles of this
pharmacologically targetable interaction in multiple UTI-relevant niches.
项目总结/摘要
细菌粘附到尿路上皮是建立尿路感染的关键步骤。期间
哺乳动物膀胱的感染(膀胱炎)、尿路致病性大肠杆菌(UPEC)被充分描述,
使用1型皮利,轴承尖端粘附素FimH,结合寡甘露糖修饰的uroplakins,包被
浅表膀胱上皮细胞的腔表面。然而,关于宿主-病原体的详细情况知之甚少
肾脏中的相互作用,使上尿路感染,包括肾盂肾炎和肾脓肿的启动。
我们发现,以前认为仅在膀胱炎中必不可少的1型皮利也介导了膀胱炎的建立。
肾盂肾炎和C3 H小鼠肾水肿的起始。此外,在UPEC的体外模型中,
结合肾集合管上皮,我们确定了FimH的候选肾上皮受体,即
甘露糖基化细胞连接蛋白桥粒芯糖蛋白2(Dsg 2)。这种蛋白质在整个
肾单位,但在集合管上皮细胞中最高,并具有典型的N-连接的含甘露糖聚糖
以及钙粘蛋白家族特异性O-连接的甘露糖基化。在这个项目中,我们将测试中心假设
桥粒芯糖蛋白-2是FimH的上皮受体,其介导UPEC肾盂肾炎的建立,
可结合肠道和脱落膀胱中的FimH。首先,我们将使用多种遗传和药理学系统,
在最近发表的优化的小鼠中,询问FimH与甘露糖基化Dsg 2结合的重要性
UPEC肾盂肾炎模型。在这些系统中,新产生的C3 H小鼠携带肾
上皮特异性Dsg 2缺失。接下来,我们将定量FimH凝集素结构域与
通过SPR纯化人DSG 2的胞外结构域,并共结晶相关的FimH和DSG 2结构域
揭示DSG 2-FimH相互作用的结构基础。这些实验中的对照包括
FimHQ 133 K,其携带废除甘露糖结合的突变;甘露糖苷,高亲和力小分子
FimH结合抑制剂;纯化蛋白和肾组织切片的酶预处理,以消除
N-或O-连接的聚糖;以及针对介导DSG 2的关键肽产生的单克隆抗体。
与FimH的相互作用第三,桥粒芯糖蛋白-2也在其他上皮细胞类型上广泛表达(在两种上皮细胞中均是如此)。
人和小鼠),增加了它在与UTI相关的其他小生境中结合FimH的可能性
发病机制这些包括脱落后的膀胱(对初始UPEC感染的快速反应,
消除主要的FimH受体)和结肠(作为UPEC储存库,
UTI)。因此,我们将使用小鼠和人体组织切片,体内肠道定植模型,
额外的新的条件性Dsg 2敲除小鼠,以研究Dsg 2是否可以作为FimH受体,
这些组织。在这些研究的结论中,我们将确定1型皮利的新受体,
阐明了FimH与桥粒芯糖蛋白-2相互作用的结构基础,并定义了这种相互作用的功能作用。
在多个UTI相关的小生境中进行可扩展的目标交互。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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DAVID ALAN HUNSTAD其他文献
DAVID ALAN HUNSTAD的其他文献
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{{ truncateString('DAVID ALAN HUNSTAD', 18)}}的其他基金
Novel Type 1 Pilus Receptors in Pyelonephritis and Recurrent UTI
肾盂肾炎和复发性尿路感染中的新型 1 型菌毛受体
- 批准号:
10378625 - 财政年份:2021
- 资助金额:
$ 39.38万 - 项目类别:
Novel Type 1 Pilus Receptors in Pyelonephritis and Recurrent UTI
肾盂肾炎和复发性尿路感染中的新型 1 型菌毛受体
- 批准号:
10594971 - 财政年份:2021
- 资助金额:
$ 39.38万 - 项目类别:
ANDROGEN INFLUENCE ON UTI SUSCEPTIBILITY AND SEVERITY
雄激素对尿路感染易感性和严重程度的影响
- 批准号:
9445746 - 财政年份:2018
- 资助金额:
$ 39.38万 - 项目类别:
ANDROGEN INFLUENCE ON UTI SUSCEPTIBILITY AND SEVERITY
雄激素对尿路感染易感性和严重程度的影响
- 批准号:
9754116 - 财政年份:2018
- 资助金额:
$ 39.38万 - 项目类别:
ANDROGEN INFLUENCE ON UTI SUSCEPTIBILITY AND SEVERITY
雄激素对尿路感染易感性和严重程度的影响
- 批准号:
9925646 - 财政年份:2018
- 资助金额:
$ 39.38万 - 项目类别:
TARGETING THE E COLI CHAPERONE SURA IN RECURRENT UTI
针对复发性尿路感染中的大肠杆菌伴侣 SURA
- 批准号:
8361365 - 财政年份:2011
- 资助金额:
$ 39.38万 - 项目类别:
TARGETING THE E COLI CHAPERONE SURA IN RECURRENT UTI
针对复发性尿路感染中的大肠杆菌伴侣 SURA
- 批准号:
8168719 - 财政年份:2010
- 资助金额:
$ 39.38万 - 项目类别:
ADHESIN-BASED NANOTHERAPEUTICS IN URINARY TRACT INFECTION
基于粘附素的纳米疗法治疗尿路感染
- 批准号:
7884834 - 财政年份:2010
- 资助金额:
$ 39.38万 - 项目类别:
ADHESIN-BASED NANOTHERAPEUTICS IN URINARY TRACT INFECTION
基于粘附素的纳米疗法治疗尿路感染
- 批准号:
8721936 - 财政年份:2010
- 资助金额:
$ 39.38万 - 项目类别:
ADHESIN-BASED NANOTHERAPEUTICS IN URINARY TRACT INFECTION
基于粘附素的纳米疗法治疗尿路感染
- 批准号:
8321543 - 财政年份:2010
- 资助金额:
$ 39.38万 - 项目类别:
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