RRC ID 64133
Author Kinoshita N, Huang AJY, McHugh TJ, Suzuki SC, Masai I, Kim IH, Soderling SH, Miyawaki A, Shimogori T.
Title Genetically Encoded Fluorescent Indicator GRAPHIC Delineates Intercellular Connections.
Journal iScience
Abstract Intercellular contacts are essential for precise organ morphogenesis, function, and maintenance; however, spatiotemporal information of cell-cell contacts or adhesions remains elusive in many systems. We developed a genetically encoded fluorescent indicator for intercellular contacts with optimized intercellular GFP reconstitution using glycosylphosphatidylinositol (GPI) anchor, GRAPHIC (GPI anchored reconstitution-activated proteins highlight intercellular connections), which can be used for an expanded number of cell types. We observed a robust GFP signal specifically at the interface between cultured cells, without disrupting natural cell contact. Application of GRAPHIC to the fish retina specifically delineated cone-bipolar connection sites. Moreover, we showed that GRAPHIC can be used in the mouse central nervous system to delineate synaptic sites in the thalamocortical circuit. Finally, we generated GRAPHIC color variants, enabling detection of multiple convergent contacts simultaneously in cell culture system. We demonstrated that GRAPHIC has high sensitivity and versatility, which will facilitate the analysis of the complex multicellular connections without previous limitations.
Volume 15
Pages 28-38
Published 2019-5-31
DOI 10.1016/j.isci.2019.04.013
PII S2589-0042(19)30110-5
PMID 31026667
PMC PMC6482341
IF 4.447
Resource
DNA material pCSII-CMV-MCS _c-GRAPHIC T2A H2B-Azurite (RDB15873) pCSII-CMV-MCS _Y (YFP- type) c-GRAPHIC T2A H2B-Azurite (RDB15874) CSII-CMV-MCS (RDB04377) pCMV-VSV-G-RSV-Rev (RDB04393) pCAG-HIVgp (RDB04394) pCSII-CMV-MCS _n-GRAPHIC (RDB15869) pCSII-CMV-MCS_n-GRAPHIC T2A H2B-mCherry (RDB15870) pCSII-CMV-MCS_B (BFP type) n-GRAPHIC T2A H2B-mCherry (RDB15871) pCSII-CMV-MCS _c-GRAPHIC (RDB15872)