{"title":"用光-E-粘连蛋白对细胞-细胞粘连进行可逆光调节","authors":"Christopher Raab, Seraphine Wegner","doi":"10.21769/BioProtoc.4995","DOIUrl":null,"url":null,"abstract":"The cell–cell adhesion molecule E-cadherin has been intensively studied due to its prevalence in tissue function and its spatiotemporal regulation during epithelial-to-mesenchymal cell transition. Nonetheless, regulating and studying the dynamics of it has proven challenging. We developed a photoswitchable version of E-cadherin, named opto-E-cadherin, which can be toggled OFF with blue light illumination and back ON in the dark. Herein, we describe easy-to-use methods to test and characterise opto-E- cadherin cell clones for downstream experiments. Key features • This protocol describes how to implement optogenetic cell–cell adhesion molecules effectively (described here on the basis of opto-E-cadherin), while highlighting possible pitfalls. • Utilises equipment commonly found in most laboratories with high ease of use. • Phenotype screening is easy and done within a few hours (comparison of cell clusters in the dark vs. blue light in an aggregation assay). • Three different functionality assay systems are described. • After the cell line is established, all experiments can be performed within three days.","PeriodicalId":93907,"journal":{"name":"Bio-protocol","volume":null,"pages":null},"PeriodicalIF":1.0000,"publicationDate":"2024-05-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Reversible Photoregulation of Cell–Cell Adhesions With Opto-E-cadherin\",\"authors\":\"Christopher Raab, Seraphine Wegner\",\"doi\":\"10.21769/BioProtoc.4995\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"The cell–cell adhesion molecule E-cadherin has been intensively studied due to its prevalence in tissue function and its spatiotemporal regulation during epithelial-to-mesenchymal cell transition. Nonetheless, regulating and studying the dynamics of it has proven challenging. We developed a photoswitchable version of E-cadherin, named opto-E-cadherin, which can be toggled OFF with blue light illumination and back ON in the dark. Herein, we describe easy-to-use methods to test and characterise opto-E- cadherin cell clones for downstream experiments. Key features • This protocol describes how to implement optogenetic cell–cell adhesion molecules effectively (described here on the basis of opto-E-cadherin), while highlighting possible pitfalls. • Utilises equipment commonly found in most laboratories with high ease of use. • Phenotype screening is easy and done within a few hours (comparison of cell clusters in the dark vs. blue light in an aggregation assay). • Three different functionality assay systems are described. • After the cell line is established, all experiments can be performed within three days.\",\"PeriodicalId\":93907,\"journal\":{\"name\":\"Bio-protocol\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":1.0000,\"publicationDate\":\"2024-05-20\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Bio-protocol\",\"FirstCategoryId\":\"0\",\"ListUrlMain\":\"https://doi.org/10.21769/BioProtoc.4995\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q3\",\"JCRName\":\"BIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Bio-protocol","FirstCategoryId":"0","ListUrlMain":"https://doi.org/10.21769/BioProtoc.4995","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"BIOLOGY","Score":null,"Total":0}
Reversible Photoregulation of Cell–Cell Adhesions With Opto-E-cadherin
The cell–cell adhesion molecule E-cadherin has been intensively studied due to its prevalence in tissue function and its spatiotemporal regulation during epithelial-to-mesenchymal cell transition. Nonetheless, regulating and studying the dynamics of it has proven challenging. We developed a photoswitchable version of E-cadherin, named opto-E-cadherin, which can be toggled OFF with blue light illumination and back ON in the dark. Herein, we describe easy-to-use methods to test and characterise opto-E- cadherin cell clones for downstream experiments. Key features • This protocol describes how to implement optogenetic cell–cell adhesion molecules effectively (described here on the basis of opto-E-cadherin), while highlighting possible pitfalls. • Utilises equipment commonly found in most laboratories with high ease of use. • Phenotype screening is easy and done within a few hours (comparison of cell clusters in the dark vs. blue light in an aggregation assay). • Three different functionality assay systems are described. • After the cell line is established, all experiments can be performed within three days.