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Mechanisms of tether function in endolysosomal trafficking - Renewal - Resubmission 01

内溶酶体运输中系链功能的机制 - 更新 - 重新提交 01

基本信息

批准号：
10379460
负责人：
AARON P TURKEWITZ
金额：
$ 34.57万
依托单位：
UNIVERSITY OF CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-05-01 至 2024-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10379460
关键词：
Address Animal Model Animals Binding Biochemical Capsid Proteins Carrier Proteins Cell fusion Cell surface Cells Chimeric Proteins Complex Disease Electron Microscopy Endosomes Epitopes Eukaryota Exocytosis Fostering Gel Chromatography Gene Silencing Genes Genetic Screening Guanosine Triphosphate Phosphohydrolases Human Hybrids Image In Vitro Individual Link Lipids Lysosomes Malignant Neoplasms Mass Spectrum Analysis Measures Membrane Membrane Fusion Modeling Molecular Mucopolysaccharidoses Mutation Nerve Degeneration Organelles Organism Pathway interactions Periodicity Photobleaching Physiology Play Protein Dynamics Protein Overexpression Protein Sortings Proteins Publishing Recording of previous events Role SNAP receptor Signal Transduction Specificity System Testing Tetrahymena Tetrahymena thermophila Vacuole Variant Vesicle Work base experimental study fungus in vitro activity in vivo in vivo evaluation insight kidney dysfunction light microscopy paralogous gene protein complex protein protein interaction protein transport reconstitution tool trafficking uptake

项目摘要

Project Summary The accurate trafficking of proteins through membrane compartments depends on the ability of specific vesicles to recognize one another and undergo efficient fusion. Accuracy is conferred in part by molecular tethers, which are multivalent complexes that bind to vesicle determinants and foster specific vesicle-vesicle contacts. Endosome-lysosome trafficking is required for the processing of cargo taken up from the cell surface, and also paradoxically for the formation of some secretory organelles. Two key tethers involved in endolysosomal trafficking are CORVET (class C core vacuole/endosome tethering) and HOPS (homotypic fusion and protein sorting). Both are hetero-hexameric complexes, whose importance in cell and organismal physiology is shown by the fact that human variants in the genes encoding CORVET/HOPS subunits are linked with mucopolysaccharidosis, renal dysfunction, neurodegeneration and cancer, among other diseases. At the cellular level, CORVET and HOPS both function in homotypic fusion, i.e., allow two vesicles with the same Rab determinants to recognize one another. The mechanisms of action of CORVET/HOPS have been deduced primarily via in vitro reconstitution experiments. However, key aspects of current models have yet to be tested in vivo, including important mechanistic details such as whether the complexes are stably associated with membranes, whether individual complexes undergo cyclical assembly/disassembly, and whether different subunits have distinct cycling dynamics. In addition, the assembly state of tethers on vesicles is unknown, a salient question because the tether subunits are structurally similar to coat proteins that form large assemblies on membranes. In the lineage of ciliates including the model organism Tetrahymena thermophila, the HOPS complex was lost. Concurrently, CORVET complexes multiplied in these cells, with individual complexes becoming specialized for distinct endolysosomal pathways. Due to its specific evolutionary history combined with experimental strengths, Tetrahymena offers a unique new system to analyze these universal tethers. Issues to be addressed include the copy number of CORVET complexes associated with vesicles, the dynamics of both the holo-complexes and individual subunits, and the role of specific protein-protein interactions. Experiments will be based on a combination of biochemical analysis, correlated light and electron microscopy to follow tagged proteins expressed at endogenous levels, and cell fusion to detect protein dynamics. Correlative light and electron microscopy will be used to visualize the ultrastructure of membrane compartments decorated by tagged proteins, allowing for detailed in vivo analysis of tether function.

项目总结

项目成果

期刊论文数量（15）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

An Alveolata secretory machinery adapted to parasite host cell invasion.

DOI：
10.1038/s41564-020-00854-z
发表时间：
2021-04
期刊：
Nature microbiology
影响因子：
28.3
作者：
Aquilini E;Cova MM;Mageswaran SK;Dos Santos Pacheco N;Sparvoli D;Penarete-Vargas DM;Najm R;Graindorge A;Suarez C;Maynadier M;Berry-Sterkers L;Urbach S;Fahy PR;Guérin AN;Striepen B;Dubremetz JF;Chang YW;Turkewitz AP;Lebrun M
通讯作者：
Lebrun M

Genome plasticity in response to stress in Tetrahymena thermophila: selective and reversible chromosome amplification and paralogous expansion of metallothionein genes.