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Multiplexed protein profiling reveals spatial subcellular signaling networks
oleh: Shuangyi Cai, Thomas Hu, Mythreye Venkatesan, Mayar Allam, Frank Schneider, Suresh S. Ramalingam, Shi-Yong Sun, Ahmet F. Coskun
Format: | Article |
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Diterbitkan: | Elsevier 2022-09-01 |
Deskripsi
Summary: Protein-protein interaction networks are altered in multi-gene dysregulations in many disorders. Image-based protein multiplexing sheds light on signaling pathways to dissect cell-to-cell heterogeneity, previously masked by the bulk assays. Herein, we present a rapid multiplexed immunofluorescence (RapMIF) method measuring up to 25-plex spatial protein maps from cultures and tissues at subcellular resolution, providing combinatorial 272 pairwise and 1,360 tri-protein signaling states across 33 multiplexed pixel-level clusters. The RapMIF pipeline automated staining, bleaching, and imaging of the biospecimens in a single platform. RapMIF showed that WNT/β-catenin signaling upregulated upon the inhibition of the AKT/mTOR pathway. Subcellular protein images demonstrated translocation patterns, spatial receptor discontinuity, and subcellular signaling clusters in single cells. Signaling networks exhibited spatial redistribution of signaling proteins in drug-responsive cultures. Machine learning analysis predicted the phosphorylated β-catenin expression from interconnected signaling protein images. RapMIF is an ideal signaling discovery approach for precision therapy design.