Image-based high-throughput mapping of TGF-beta-induced phosphocomplexes at a single-cell level
| العنوان: | Image-based high-throughput mapping of TGF-beta-induced phosphocomplexes at a single-cell level |
|---|---|
| المؤلفون: | Masood Kamali-Moghaddam, Rasel A. Al-Amin, Ehsan Manouchehri Doulabi, Johan Björkesten, Peter Lönn, Johan Heldin, Radiosa Gallini, Ulf Landegren, Johan Oelrich |
| المصدر: | Communications Biology, Vol 4, Iss 1, Pp 1-9 (2021) Communications Biology |
| بيانات النشر: | Uppsala universitet, Molekylära verktyg, 2021. |
| سنة النشر: | 2021 |
| مصطلحات موضوعية: | In situ, QH301-705.5, Cell, Phosphatase, Medicine (miscellaneous), Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy), Article, General Biochemistry, Genetics and Molecular Biology, Protein–protein interaction, Transforming Growth Factor beta1, chemistry.chemical_compound, Protein Interaction Mapping, medicine, Image Processing, Computer-Assisted, HaCaT Cells, Humans, Biology (General), Phosphorylation, Throughput (business), Medicinsk bioteknologi (med inriktning mot cellbiologi (inklusive stamcellsbiologi), molekylärbiologi, mikrobiologi, biokemi eller biofarmaci), Smad4 Protein, Chemistry, High-throughput screening, Growth factor signalling, Cell biology, medicine.anatomical_structure, Single-Cell Analysis, General Agricultural and Biological Sciences, Linker, DNA, Transforming growth factor |
| الوصف: | Protein interactions and posttranslational modifications orchestrate cellular responses to e.g. cytokines and drugs, but it has been difficult to monitor these dynamic events in high-throughput. Here, we describe a semi-automated system for large-scale in situ proximity ligation assays (isPLA), combining isPLA in microtiter wells with automated microscopy and computer-based image analysis. Phosphorylations and interactions are digitally recorded along with subcellular morphological features. We investigated TGF-β-responsive Smad2 linker phosphorylations and complex formations over time and across millions of individual cells, and we relate these events to cell cycle progression and local cell crowding via measurements of DNA content and nuclear size of individual cells, and of their relative positions. We illustrate the suitability of this protocol to screen for drug effects using phosphatase inhibitors. Our approach expands the scope for image-based single cell analyses by combining observations of protein interactions and modifications with morphological details of individual cells at high throughput. To improve our ability to monitor cellular responses to e.g. cytokines or drugs, Lönn et al have developed a semi-automated system for large-scale in situ proximity ligation assays (isPLA) in HaCAT keratinocyte cells. Their approach expands the scope for image-based single cell analyses by combining observations of protein interactions and modifications with morphological details of individual cells at high throughput. |
| وصف الملف: | application/pdf |
| اللغة: | English |
| URL الوصول: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::8d8fa2e1a506a5e0a72143f491041a88 http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-459770 |
| حقوق: | OPEN |
| رقم الأكسشن: | edsair.doi.dedup.....8d8fa2e1a506a5e0a72143f491041a88 |
| قاعدة البيانات: | OpenAIRE |
| الوصف غير متاح. |