Unveiling cellular communications through rapid pan-membrane-protein labeling
Abstract Dynamic protein distribution within and across the plasma membrane is pivotal in regulating cell communication. However, rapid, high-density labeling methods for multiplexed live imaging across diverse cell types remain scarce. Here, we demonstrate N-hydroxysuccinimide (NHS)-ester-based ami...
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| Format: | Article |
| Language: | English |
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Nature Portfolio
2025-04-01
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| Series: | Nature Communications |
| Online Access: | https://doi.org/10.1038/s41467-025-58779-2 |
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| author | Hirushi Gunasekara Yu-Shiuan Cheng Vanessa Perez-Silos Alejandro Zevallos-Morales Daniel Abegg Alyssa Burgess Liang-Wei Gong Richard D. Minshall Alexander Adibekian Carlos Murga-Zamalloa Alison E. Ondrus Ying S. Hu |
| author_facet | Hirushi Gunasekara Yu-Shiuan Cheng Vanessa Perez-Silos Alejandro Zevallos-Morales Daniel Abegg Alyssa Burgess Liang-Wei Gong Richard D. Minshall Alexander Adibekian Carlos Murga-Zamalloa Alison E. Ondrus Ying S. Hu |
| author_sort | Hirushi Gunasekara |
| collection | DOAJ |
| description | Abstract Dynamic protein distribution within and across the plasma membrane is pivotal in regulating cell communication. However, rapid, high-density labeling methods for multiplexed live imaging across diverse cell types remain scarce. Here, we demonstrate N-hydroxysuccinimide (NHS)-ester-based amine crosslinking of fluorescent dyes to uniformly label live mammalian cell surface proteins. Using model cell systems, we capture previously elusive membrane topology and cell-cell interactions. Live imaging shows transient membrane protein accumulation at cell-cell contacts and bidirectional migration patterns guided by membrane fibers in DC2.4 dendritic cells. Multiplexed superresolution imaging reveals the biogenesis of membrane tunneling nanotubes that facilitate intercellular transfer in DC2.4 cells, and caveolin 1-dependent endocytosis of insulin receptors in HEK293T cells. 3D superresolution imaging reveals membrane topology remodeling in response to stimulation, generation of microvesicles, and phagocytic activities in Jurkat T cells. Furthermore, NHS-labeling remains stable in vivo, enabling visualization of intercellular transfer among splenocytes using a T cell lymphoma mouse model. |
| format | Article |
| id | doaj-art-7727d39d210c4fcdb4bb9d8973b0b84f |
| institution | OA Journals |
| issn | 2041-1723 |
| language | English |
| publishDate | 2025-04-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Nature Communications |
| spelling | doaj-art-7727d39d210c4fcdb4bb9d8973b0b84f2025-08-20T02:17:48ZengNature PortfolioNature Communications2041-17232025-04-0116111510.1038/s41467-025-58779-2Unveiling cellular communications through rapid pan-membrane-protein labelingHirushi Gunasekara0Yu-Shiuan Cheng1Vanessa Perez-Silos2Alejandro Zevallos-Morales3Daniel Abegg4Alyssa Burgess5Liang-Wei Gong6Richard D. Minshall7Alexander Adibekian8Carlos Murga-Zamalloa9Alison E. Ondrus10Ying S. Hu11Department of Chemistry, College of Liberal Arts and Sciences, University of Illinois ChicagoDepartment of Chemistry, College of Liberal Arts and Sciences, University of Illinois ChicagoDepartment of Pathology, College of Medicine, University of Illinois ChicagoDepartment of Pathology, College of Medicine, University of Illinois ChicagoDepartment of Chemistry, College of Liberal Arts and Sciences, University of Illinois ChicagoDepartment of Chemistry, College of Liberal Arts and Sciences, University of Illinois ChicagoDepartment of Biological Sciences, College of Liberal Arts and Sciences, University of Illinois ChicagoDepartments of Anesthesiology & Pharmacology and Regenerative Medicine, College of Medicine, University of Illinois ChicagoDepartment of Chemistry, College of Liberal Arts and Sciences, University of Illinois ChicagoDepartment of Pathology, College of Medicine, University of Illinois ChicagoDepartment of Chemistry, College of Liberal Arts and Sciences, University of Illinois ChicagoDepartment of Chemistry, College of Liberal Arts and Sciences, University of Illinois ChicagoAbstract Dynamic protein distribution within and across the plasma membrane is pivotal in regulating cell communication. However, rapid, high-density labeling methods for multiplexed live imaging across diverse cell types remain scarce. Here, we demonstrate N-hydroxysuccinimide (NHS)-ester-based amine crosslinking of fluorescent dyes to uniformly label live mammalian cell surface proteins. Using model cell systems, we capture previously elusive membrane topology and cell-cell interactions. Live imaging shows transient membrane protein accumulation at cell-cell contacts and bidirectional migration patterns guided by membrane fibers in DC2.4 dendritic cells. Multiplexed superresolution imaging reveals the biogenesis of membrane tunneling nanotubes that facilitate intercellular transfer in DC2.4 cells, and caveolin 1-dependent endocytosis of insulin receptors in HEK293T cells. 3D superresolution imaging reveals membrane topology remodeling in response to stimulation, generation of microvesicles, and phagocytic activities in Jurkat T cells. Furthermore, NHS-labeling remains stable in vivo, enabling visualization of intercellular transfer among splenocytes using a T cell lymphoma mouse model.https://doi.org/10.1038/s41467-025-58779-2 |
| spellingShingle | Hirushi Gunasekara Yu-Shiuan Cheng Vanessa Perez-Silos Alejandro Zevallos-Morales Daniel Abegg Alyssa Burgess Liang-Wei Gong Richard D. Minshall Alexander Adibekian Carlos Murga-Zamalloa Alison E. Ondrus Ying S. Hu Unveiling cellular communications through rapid pan-membrane-protein labeling Nature Communications |
| title | Unveiling cellular communications through rapid pan-membrane-protein labeling |
| title_full | Unveiling cellular communications through rapid pan-membrane-protein labeling |
| title_fullStr | Unveiling cellular communications through rapid pan-membrane-protein labeling |
| title_full_unstemmed | Unveiling cellular communications through rapid pan-membrane-protein labeling |
| title_short | Unveiling cellular communications through rapid pan-membrane-protein labeling |
| title_sort | unveiling cellular communications through rapid pan membrane protein labeling |
| url | https://doi.org/10.1038/s41467-025-58779-2 |
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