Cell-type-specific signaling networks in heterocellular organoids

Nat Methods. 2020 Mar;17(3):335-342. doi: 10.1038/s41592-020-0737-8. Epub 2020 Feb 17.


Despite the widespread adoption of organoids as biomimetic tissue models, methods to comprehensively analyze cell-type-specific post-translational modification (PTM) signaling networks in organoids are absent. Here, we report multivariate single-cell analysis of such networks in organoids and organoid cocultures. Simultaneous analysis by mass cytometry of 28 PTMs in >1 million single cells derived from small intestinal organoids reveals cell-type- and cell-state-specific signaling networks in stem, Paneth, enteroendocrine, tuft and goblet cells, as well as enterocytes. Integrating single-cell PTM analysis with thiol-reactive organoid barcoding in situ (TOBis) enables high-throughput comparison of signaling networks between organoid cultures. Cell-type-specific PTM analysis of colorectal cancer organoid cocultures reveals that shApc, KrasG12D and Trp53R172H cell-autonomously mimic signaling states normally induced by stromal fibroblasts and macrophages. These results demonstrate how standard mass cytometry workflows can be modified to perform high-throughput multivariate cell-type-specific signaling analysis of healthy and cancerous organoids.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Biomimetics*
  • Cell Differentiation
  • Coculture Techniques / methods
  • Colorectal Neoplasms / metabolism
  • Colorectal Neoplasms / pathology*
  • Cytophotometry / methods
  • Enterocytes / cytology
  • Enteroendocrine Cells / cytology
  • Female
  • Fibroblasts / cytology
  • Gene Expression Regulation*
  • Goblet Cells / cytology
  • Humans
  • Intestine, Small / cytology*
  • Macrophages / cytology
  • Mice
  • Mice, Inbred C57BL
  • Organ Culture Techniques
  • Organoids / metabolism*
  • Paneth Cells / cytology
  • Signal Transduction*
  • Single-Cell Analysis / methods
  • Sulfhydryl Compounds / chemistry
  • Tumor Suppressor Protein p53 / metabolism


  • Sulfhydryl Compounds
  • TP53 protein, human
  • Tumor Suppressor Protein p53