Anterior gradient 2 (AGR2) is an endoplasmic reticulum (ER)-resident protein disulfide isomerase (PDI) known to be overexpressed in many human epithelial cancers and is involved in cell migration, cellular transformation, angiogenesis, and metastasis. This protein inhibits the activity of the tumor suppressor p53, and its expression levels can be used to predict cancer patient outcome. However, the precise network of AGR2-interacting partners and clients remains to be fully characterized. Herein, we used label-free quantification and also stable isotope labeling with amino acids in cell culture-based LC-MS/MS analyses to identify proteins interacting with AGR2. Functional annotation confirmed that AGR2 and its interaction partners are associated with processes in the ER that maintain intracellular metabolic homeostasis and participate in the unfolded protein response, including those associated with changes in cellular metabolism, energy, and redox states in response to ER stress. As a proof of concept, the interaction between AGR2 and PDIA3, another ER-resident PDI, was studied in more detail. Pathway analysis revealed that AGR2 and PDIA3 play roles in protein folding in ER, including post-translational modification and in cellular response to stress. We confirmed the AGR2-PDIA3 complex formation in cancer cells, which was enhanced in response to ER stress. Accordingly, molecular docking characterized potential quaternary structure of this complex; however, it remains to be elucidated whether AGR2 rather contributes to PDIA3 maturation in ER, the complex directly acts in cellular signaling, or mediates AGR2 secretion. Our study provides a comprehensive insight into the protein-protein interaction network of AGR2 by identifying functionally relevant proteins and related cellular and biochemical pathways associated with the role of AGR2 in cancer cells.
- Klíčová slova
- anterior gradient protein 2, mass spectrometry, protein disulfide isomerase, protein–protein interactions, secretory pathway,
- MeSH
- chromatografie kapalinová MeSH
- lidé MeSH
- mapy interakcí proteinů MeSH
- mukoproteiny * metabolismus MeSH
- nádory * MeSH
- onkogenní proteiny * metabolismus MeSH
- proteindisulfidisomerasy * MeSH
- simulace molekulového dockingu MeSH
- tandemová hmotnostní spektrometrie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- AGR2 protein, human MeSH Prohlížeč
- mukoproteiny * MeSH
- onkogenní proteiny * MeSH
- proteindisulfidisomerasy * MeSH
Accurate classification of breast tumors is vital for patient management decisions and enables more precise cancer treatment. Here, we present a quantitative proteotyping approach based on sequential windowed acquisition of all theoretical fragment ion spectra (SWATH) mass spectrometry and establish key proteins for breast tumor classification. The study is based on 96 tissue samples representing five conventional breast cancer subtypes. SWATH proteotype patterns largely recapitulate these subtypes; however, they also reveal varying heterogeneity within the conventional subtypes, with triple negative tumors being the most heterogeneous. Proteins that contribute most strongly to the proteotype-based classification include INPP4B, CDK1, and ERBB2 and are associated with estrogen receptor (ER) status, tumor grade status, and HER2 status. Although these three key proteins exhibit high levels of correlation with transcript levels (R > 0.67), general correlation did not exceed R = 0.29, indicating the value of protein-level measurements of disease-regulated genes. Overall, this study highlights how cancer tissue proteotyping can lead to more accurate patient stratification.
- Klíčová slova
- SWATH-MS, breast cancer, data independent acquisition, proteomics, tissue, transcriptomics, tumor classification,
- MeSH
- fosfatasy genetika metabolismus MeSH
- lidé MeSH
- nádory prsu klasifikace metabolismus patologie MeSH
- proteinkinasa CDC2 genetika metabolismus MeSH
- proteom analýza metabolismus MeSH
- proteomika metody MeSH
- receptor erbB-2 genetika metabolismus MeSH
- receptory pro estrogeny metabolismus MeSH
- rychlé screeningové testy MeSH
- tandemová hmotnostní spektrometrie metody MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- CDK1 protein, human MeSH Prohlížeč
- ERBB2 protein, human MeSH Prohlížeč
- fosfatasy MeSH
- phosphatidylinositol-3,4-bisphosphate 4-phosphatase MeSH Prohlížeč
- proteinkinasa CDC2 MeSH
- proteom MeSH
- receptor erbB-2 MeSH
- receptory pro estrogeny MeSH
Estrogen receptors [ERs (subtypes α and β)], classified as a nuclear receptor super family, are intracellular proteins with an important biological role as the transcription factors for estrogen target genes. For ER-induced transcription, an interaction must exist between ligand and coregulators. Coregulators may stimulate (coactivators) or inhibit (corepressors) transcription, following binding with a specific region of the gene, called the estrogen response element. Misbalanced activity of coregulators or higher ligand concentrations may cause increased cell proliferation, resulting in specific types of cancer. These are exhibited as overexpression of ER proteins. Breast cancer currently ranks first in the incidence and second in the mortality of cancer in females worldwide. In addition, 70% of breast tumors are ERα positive and the importance of these proteins for diagnostic use is indisputable. Early diagnosis of the tumor and its classification has a large influence on the selection of appropriate therapy, as ER-positive tumors demonstrate a positive response to hormonal therapy. Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI TOF MS) has been hypothesized to have great potential, as it offers reliable, robust and efficient analysis methods for biomarker monitoring and identification. The present review discusses ER protein analysis by MALDI TOF MS, including the crucial step of protein separation.
- Klíčová slova
- cancer, estrogen receptor, estrogen response element, matrix-assisted laser desorption/ionization time of flight,
- Publikační typ
- časopisecké články MeSH
