Identification of Cell Signaling Molecules by Expression Cloning

See allHide authors and affiliations

Science's STKE  09 Oct 2001:
Vol. 2001, Issue 103, pp. pl9
DOI: 10.1126/stke.2001.103.pl9

You are currently viewing the abstract.

View Full Text

Log in to view the full text

Log in through your institution

Log in through your institution


Using expression cloning one can isolate proteins with specific biological functions. This methodology can be adapted for the identification of novel players in the regulation of cell signaling. Here, we describe an expression cloning strategy to identify suppressors of Ras signaling. This screen is based on the observation that the activation of the small guanosine triphosphate (GTP)-binding protein H-Ras initiates a mitogen-activated protein kinase (MAPK)-dependent signaling pathway that inactivates integrin ligand binding. Our strategy depends on flow cytometry and a monoclonal antibody that recognizes integrin activation states. Flow cytometry enhances the screen's sensitivity thereby allowing us to examine function quantitatively at the level of a single cell millions of times in one screen. The following protocol provides a detailed method for the isolation of proteins that regulate cell signaling.

View Full Text

Stay Connected to Science Signaling

Editor's Blog