Host cell factors controlling type III secretion effector translocation
控制III型分泌效应器易位的宿主细胞因子
基本信息
- 批准号:10586145
- 负责人:
- 金额:$ 20.13万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2022
- 资助国家:美国
- 起止时间:2022-03-07 至 2024-02-29
- 项目状态:已结题
- 来源:
- 关键词:5&apos Splice SiteAffectAttenuatedBacteriaBacterial Attachment SiteBiochemical PathwayCRISPR/Cas technologyCell LineCell membraneCell physiologyCell secretionCellsDiseaseDockingDown-RegulationEbola virusExperimental DesignsFeedbackGene ActivationGene ExpressionGenesGenomicsGoalsHaploid CellsHaploidyHigh-Throughput Nucleotide SequencingHospitalsHuman Cell LineImmuneImmune systemInfectionInjectionsInsertion MutationIntegration Host FactorsLife StyleMediatingMolecularMolecular ConformationMolecular MachinesMutagenesisMutagensMutationNeedlesPathogenicityPathway interactionsPerforationPhospholipasePlasma CellsPlayProcessProductionProteinsPseudomonas aeruginosaResearchResistanceRoleSiteStructureSyringesSystemToxinType III Secretion System PathwayWorkbasecandidate identificationcytotoxicdrug developmentexperimental studygain of functiongain of function mutationgenome-wideloss of function mutationmortalitymutantnoveloverexpressionpathogenpromotersecretion processsensorsmall molecule inhibitor
项目摘要
Abstract
Type III secretion systems (T3SS) are intricate molecular machines that allow Gram-negative pathogens
to directly inject effector proteins into host cells. While the arsenal of injected effector proteins varies among
pathogens and supports a wide variety of pathogenic lifestyles, the core apparatus is conserved. A key feature
of T3SSs is that effector translocation is triggered by host-cell contact. In order to inject proteins, the bacterium
attaches to the host cell and uses the T3SS to insert pore-forming translocator proteins into the host cell plasma
membrane. The tip of the needle is docked to this pore, forming the translocon. Host cell contact is sensed by a
conformational change in the pore that is propagated to the base of the apparatus. While there is ample evidence
that translocon assembly and function are modulated by the host cell, which cellular functions are involved, and
what step in the translocation process they influence is poorly understood.
We are proposing to perform a genome wide saturation mutagenesis selection in a haploid human cell
line to identify cellular pathways that impact effector translocation by the Pseudomonas aeruginosa type III
secretion system. We will employ a transposon that also harbors a constitutive promoter and splice donor site,
which will allow us to not only identify loss of function mutations, but also gain of function mutations resulting
from artificial production of a downstream gene. Moreover, we will take advantage of the P. aeruginosa system
and perform the selection with either wild type bacteria or a mutant strain that does not require a specific host
cell trigger to commence effector secretion. This will allow us to isolate the host cell process that triggers effector
secretion. To achieve these goals, the application is divided into two specific aims. Aim 1 encompasses the
selection for transposon insertion mutations that confer resistance to the P. aeruginosa T3SS, as well as the
high throughput sequencing and analysis needed to identify candidate hits. In Aim 2 we will validate mutations
we identify.
Taken together this work will shed light on an understudied aspect of the type III secretion process: the
contribution of the host cell to translocon function. By identifying both loss- and gain of function mutants, and by
performing the selection in a haploid cell line, this project will be more comprehensive than any study performed
to date. The design of the experiments also allows us to specifically identify factors contributing to triggering of
effector secretion on host cell contact, a hallmark feature of type III secretion systems that remains a mystery to
this day.
摘要
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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Arne Rietsch其他文献
Arne Rietsch的其他文献
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{{ truncateString('Arne Rietsch', 18)}}的其他基金
Host cell factors controlling type III secretion effector translocation
控制III型分泌效应器易位的宿主细胞因子
- 批准号:
10416972 - 财政年份:2022
- 资助金额:
$ 20.13万 - 项目类别:
Development of recombinase-based tools to study established infections
开发基于重组酶的工具来研究已确定的感染
- 批准号:
10201472 - 财政年份:2020
- 资助金额:
$ 20.13万 - 项目类别:
Development of recombinase-based tools to study established infections
开发基于重组酶的工具来研究已确定的感染
- 批准号:
10040615 - 财政年份:2020
- 资助金额:
$ 20.13万 - 项目类别:
Type III Secretion Translocon Structure and Function
III型分泌易位子的结构和功能
- 批准号:
8701601 - 财政年份:2014
- 资助金额:
$ 20.13万 - 项目类别:
P. aeruginosa type III secreted effectors in corneal disease
角膜疾病中铜绿假单胞菌 III 型分泌效应子
- 批准号:
8404007 - 财政年份:2012
- 资助金额:
$ 20.13万 - 项目类别:
P. aeruginosa type III secreted effectors in corneal disease
角膜疾病中铜绿假单胞菌 III 型分泌效应子
- 批准号:
8962457 - 财政年份:2012
- 资助金额:
$ 20.13万 - 项目类别:
P. aeruginosa type III secreted effectors in corneal disease
角膜疾病中铜绿假单胞菌 III 型分泌效应子
- 批准号:
8217524 - 财政年份:2012
- 资助金额:
$ 20.13万 - 项目类别:
P. aeruginosa type III secreted effectors in corneal disease
角膜疾病中铜绿假单胞菌 III 型分泌效应子
- 批准号:
8597435 - 财政年份:2012
- 资助金额:
$ 20.13万 - 项目类别:
P. aeruginosa type III secreted effectors in corneal disease
角膜疾病中铜绿假单胞菌 III 型分泌效应子
- 批准号:
9115162 - 财政年份:2012
- 资助金额:
$ 20.13万 - 项目类别:
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