Cell Surface Receptor Identification Using Genome-Scale CRISPR/Cas9 Genetic Screens.

Sumana Sharma; Gavin J Wright

doi:10.3791/60803

Cell Surface Receptor Identification Using Genome-Scale CRISPR/Cas9 Genetic Screens.

Research output: Contribution to journal › Article › peer-review

Abstract

Intercellular communication mediated by direct interactions between membrane-embedded cell surface receptors is crucial for the normal development and functioning of multicellular organisms. Detecting these interactions remains technically challenging, however. This manuscript describes a systematic genome-scale CRISPR/Cas9 knockout genetic screening approach that reveals cellular pathways required for specific cell surface recognition events. This assay utilizes recombinant proteins produced in a mammalian protein expression system as avid binding probes to identify interaction partners in a cell-based genetic screen. This method can be used to identify the genes necessary for cell surface interactions detected by recombinant binding probes corresponding to the ectodomains of membrane-embedded receptors. Importantly, given the genome-scale nature of this approach, it also has the advantage of not only identifying the direct receptor but also the cellular components that are required for the presentation of the receptor at the cell surface, thereby providing valuable insights into the biology of the receptor.

Original language	English
Article number	e60803
Number of pages	15
Journal	Journal of Visualized Experiments
Issue number	160
DOIs	https://doi.org/10.3791/60803
Publication status	Published - 6 Jun 2020

Keywords

cellular genetics
staffpaper

Access to Document

10.3791/60803

documentFinal published version, 1.63 MBLicence: CC BY

Cite this

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title = "Cell Surface Receptor Identification Using Genome-Scale CRISPR/Cas9 Genetic Screens.",

abstract = "Intercellular communication mediated by direct interactions between membrane-embedded cell surface receptors is crucial for the normal development and functioning of multicellular organisms. Detecting these interactions remains technically challenging, however. This manuscript describes a systematic genome-scale CRISPR/Cas9 knockout genetic screening approach that reveals cellular pathways required for specific cell surface recognition events. This assay utilizes recombinant proteins produced in a mammalian protein expression system as avid binding probes to identify interaction partners in a cell-based genetic screen. This method can be used to identify the genes necessary for cell surface interactions detected by recombinant binding probes corresponding to the ectodomains of membrane-embedded receptors. Importantly, given the genome-scale nature of this approach, it also has the advantage of not only identifying the direct receptor but also the cellular components that are required for the presentation of the receptor at the cell surface, thereby providing valuable insights into the biology of the receptor.",

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