Cellular and structural studies of the role of CD98:integrin complexes in fibrosis.

CD98：整合素复合物在纤维化中作用的细胞和结构研究。

• 批准号: 2436361
• 金额: --
• 依托单位: University of Leicester
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2020
• 资助国家: 英国
• 起止时间: 2020 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2436361
• 关键词: Cellular; structural; studies; role; CD98

(i) We shall investigate by immuno-histochemistry the formation of profibrotic cell surface complexes caused by misfolded mutants of A1AT (liver cells) or misfolded mutants of SP-C (lung cells). To probe the dependency of the cell's response to glycoprotein misfolding, we shall use WT cells and cells in which the gene encoding the ER glycoprotein folding checkpoint enzyme UGGT has been deleted;(ii) Mass spectrometry of co-immunoprecipitated complexes will be used to identify other interacting membrane proteins (Figure 1C) including the identity of the -integrin pairing with 1-integrin here;(iii) profibrotic cell surface complexes will be purified, in the absence and presence of LPS (modelling profibrotic 'exacerbation' stimulus) and their composition and stoichiometry characterised by mass-spectrometry and biochemical methods. The complexes will be reconstituted from recombinantly expressed components and purified; their structures will be studied by X-ray crystallography (with Dr Andrew Quigley, Harwell) and Cryo-EM (at the Midlands Cryo-EM Facility in Leicester).Expected outcomes: the molecular detail of the profibrotic complexes structure/function will be elucidated and their dependency on the ER glycoprotein folding machinery assessed.

（i）我们将通过免疫 - 归化学研究由A1AT（肝细胞）（肝细胞）或SP-C（肺细胞）错误折叠突变体引起的纤维化细胞表面复合物的形成。 To probe the dependency of the cell's response to glycoprotein misfolding, we shall use WT cells and cells in which the gene encoding the ER glycoprotein folding checkpoint enzyme UGGT has been deleted;(ii) Mass spectrometry of co-immunoprecipitated complexes will be used to identify other interacting membrane proteins (Figure 1C) including the identity of the -integrin pairing with 1-整合素；（iii）在不存在和存在LPS（建模纤维化的“加剧”刺激）的情况下，将纯化纤维化细胞表面复合物及其组成和化学计量学以质量 - 光谱法和生物化学方法为特征。这些复合物将从重组表达的组件中重新组成并纯化；他们的结构将通过X射线晶体学（与Andrew Quigley博士，Harwell）和Cryo-EM（在莱斯特的Midlands Cryo-Em设施中）进行研究：预期的结果：阐明纤维化复合物结构/功能的分子细节，并将阐明对ER糖蛋白折叠式折叠机器的依赖。

项目成果

High-Throughput Production and Optimization of Membrane Proteins After Expression in Mammalian Cells.

哺乳动物细胞表达后膜蛋白的高通量生产和优化。

• DOI: 10.1007/978-1-0716-3147-8_5
• 发表时间: 2023
• 期刊: Methods in molecular biology (Clifton, N.J.)
• 作者: Gamage N

Familial hypereosinophilia associated with eosinophilic gastrointestinal symptoms in individuals with a missense mutation in CKLF-like MARVEL transmembrane domain containing 3.

在含有 3 的 CKLF 样 MARVEL 跨膜结构域中存在错义突变的个体中，家族性嗜酸性粒细胞增多与嗜酸性胃肠道症状相关。

• DOI: 10.1111/cea.13957
• 发表时间: 2021
• 期刊: journal of the British Society for Allergy and Clinical Immunology
• 作者: Wardlaw AJ