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Diffusible GRAPHIC to visualize morphology of cells after specific cell–cell contact
- Source :
- Scientific Reports, Vol 10, Iss 1, Pp 1-11 (2020), Scientific Reports
- Publication Year :
- 2020
- Publisher :
- Nature Portfolio, 2020.
-
Abstract
- The ability to identify specific cell–cell contact in the highly heterogeneous mammalian body is crucial to revealing precise control of the body plan and correct function. To visualize local connections, we previously developed a genetically encoded fluorescent indicator, GRAPHIC, which labels cell–cell contacts by restricting the reconstituted green fluorescent protein (GFP) signal to the contact site. Here, we modify GRAPHIC to give the reconstituted GFP motility within the membrane, to detect cells that make contact with other specific cells. Removal of leucine zipper domains, located between the split GFP fragment and glycophosphatidylinositol anchor domain, allowed GFP reconstituted at the contact site to diffuse throughout the entire plasma membrane, revealing cell morphology. Further, depending on the structural spacers employed, the reconstituted GFP could be selectively targeted to N terminal (NT)- or C terminal (CT)-probe-expressing cells. Using these novel constructs, we demonstrated that we can specifically label NT-probe-expressing cells that made contact with CT-probe-expressing cells in an epithelial cell culture and in Xenopus 8-cell-stage blastomeres. Moreover, we showed that diffusible GRAPHIC (dGRAPHIC) can be used in neuronal circuits to trace neurons that make contact to reveal a connection map. Finally, application in the developing brain demonstrated that the dGRAPHIC signal remained on neurons that had transient contacts during circuit development to reveal the contact history. Altogether, dGRAPHIC is a unique probe that can visualize cells that made specific cell–cell contact.
- Subjects :
- 0301 basic medicine
Blastomeres
Leucine zipper
Swine
Xenopus
Science
Green Fluorescent Proteins
Motility
GPI-Linked Proteins
Protein Engineering
Cell morphology
Article
Green fluorescent protein
Rats, Sprague-Dawley
Mice
03 medical and health sciences
0302 clinical medicine
medicine
Animals
Humans
Cells, Cultured
Neurons
Leucine Zippers
Mice, Inbred ICR
Multidisciplinary
biology
Chemistry
Biological techniques
biology.organism_classification
Recombinant Proteins
Epithelium
Rats
Cell biology
HEK293 Cells
Intercellular Junctions
030104 developmental biology
medicine.anatomical_structure
Membrane
Microscopy, Fluorescence
Medicine
030217 neurology & neurosurgery
Function (biology)
Neuroscience
Subjects
Details
- Language :
- English
- ISSN :
- 20452322
- Volume :
- 10
- Issue :
- 1
- Database :
- OpenAIRE
- Journal :
- Scientific Reports
- Accession number :
- edsair.doi.dedup.....d8220e71962bb8fef35f081111e1f88a