Cell Surface Proteomics Reveals Hypoxia-Regulated Pathways in Cervical and Bladder Cancer.

IF 3.6 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY

Proteomes Pub Date : 2025-08-05 DOI:10.3390/proteomes13030036

Faris Alanazi, Ammar Sharif, Melissa Kidd, Emma-Jayne Keevill, Vanesa Biolatti, Richard D Unwin, Peter Hoskin, Ananya Choudhury, Tim A D Smith, Conrado G Quiles

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引用次数: 0

Abstract

Background Plasma membrane proteins (PMPs) play key roles in cell signalling, adhesion, and trafficking, and are attractive therapeutic targets in cancer due to their surface accessibility. However, their typically low abundance limits detection by conventional proteomic approaches.

Methods: To improve PMP detection, we employed a surface proteomics workflow combining cell surface biotinylation and affinity purification prior to LC-MS/MS analysis in cervical (SiHa) and bladder (UMUC3) cancer cell lines cultured under normoxic (21% O₂) or hypoxic (0.1% O₂) conditions.

Results: In SiHa cells, 43 hypoxia-upregulated proteins were identified exclusively in the biotin-enriched fraction, including ITGB2, ITGA7, AXL, MET, JAG2, and CAV1/CAV2. In UMUC3 cells, 32 unique upregulated PMPs were detected, including CD55, ADGRB1, SLC9A1, NECTIN3, and ACTG1. These proteins were not observed in corresponding whole-cell lysates and are associated with extracellular matrix remodelling, immune modulation, and ion transport. Biotinylation enhanced the detection of membrane-associated pathways such as ECM organisation, integrin signalling, and PI3K-Akt activation. Protein-protein interaction analysis revealed links between membrane receptors and intracellular stress regulators, including mitochondrial proteins.

Conclusions: These findings demonstrate that surface biotinylation improves the sensitivity and selectivity of plasma membrane proteomics under hypoxia, revealing hypoxia-responsive proteins and pathways not captured by standard whole-cell analysis.

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细胞表面蛋白质组学揭示宫颈癌和膀胱癌的缺氧调节途径。

质膜蛋白（pmp）在细胞信号传导、粘附和运输中起着关键作用，由于其表面可及性而成为癌症治疗的重要靶点。然而，它们典型的低丰度限制了传统蛋白质组学方法的检测。方法：为了提高PMP的检测，我们在LC-MS/MS分析前，对在常氧（21% O2）或低氧（0.1% O2）条件下培养的宫颈癌（SiHa）和膀胱癌（UMUC3）细胞株进行了结合细胞表面生物素化和亲和纯化的表面蛋白质组学工作流程。结果：在SiHa细胞中，在生物素富集部位特异性鉴定出43种低氧上调蛋白，包括ITGB2、ITGA7、AXL、MET、JAG2和CAV1/CAV2。在UMUC3细胞中，检测到32种独特的上调pmp，包括CD55、ADGRB1、SLC9A1、NECTIN3和ACTG1。在相应的全细胞裂解物中未观察到这些蛋白，它们与细胞外基质重塑、免疫调节和离子运输有关。生物素化增强了膜相关通路的检测，如ECM组织、整合素信号传导和PI3K-Akt激活。蛋白质相互作用分析揭示了膜受体和细胞内应激调节因子（包括线粒体蛋白）之间的联系。结论：这些发现表明，表面生物素化提高了缺氧条件下质膜蛋白质组学的敏感性和选择性，揭示了标准全细胞分析无法捕获的缺氧反应蛋白和途径。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Proteomes Biochemistry, Genetics and Molecular Biology-Clinical Biochemistry

CiteScore

6.50

自引率

3.00%

发文量

审稿时长

11 weeks

期刊介绍： Proteomes (ISSN 2227-7382) is an open access, peer reviewed journal on all aspects of proteome science. Proteomes covers the multi-disciplinary topics of structural and functional biology, protein chemistry, cell biology, methodology used for protein analysis, including mass spectrometry, protein arrays, bioinformatics, HTS assays, etc. Our aim is to encourage scientists to publish their experimental and theoretical results in as much detail as possible. Therefore, there is no restriction on the length of papers. Scope: -whole proteome analysis of any organism -disease/pharmaceutical studies -comparative proteomics -protein-ligand/protein interactions -structure/functional proteomics -gene expression -methodology -bioinformatics -applications of proteomics