Open AccessFunction recovery after chemobleaching (FRAC): Evidence for activity silent membrane receptors on cell surface
Author(s) -
Haiyan Sun,
Sojin Shikano,
Qiaojie Xiong,
Min Li
Publication year - 2004
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.0404178101
Subject(s) - biogenesis , endoplasmic reticulum , microbiology and biotechnology , receptor , proteome , cell surface receptor , membrane protein , golgi apparatus , cell membrane , cell , biology , membrane , organelle , function (biology) , transport protein , chemistry , biochemistry , gene
Membrane proteins represent approximately 30% of the proteome of both prokaryotes and eukaryotes. Unique to cell surface receptors is their biogenesis pathway, which involves vesicular trafficking from the endoplasmic reticulum through the Golgi apparatus and to the cell surface. Increasing evidence suggests specific regulation of biogenesis for different membrane receptors, hence affecting their surface expression. We report the development of a pulse-chase assay to monitor function recovery after chemobleaching (FRAC) to probe the transit time of the Kir2.1 K+ channel to reach the cell surface. Our results reveal that the channel activity is contributed by a small fraction of channel protein, providing evidence of activity-silent "sleeping" molecules on the cell surface. This method distinguishes molecular density from functional density, and the assay strategy is generally applicable to other membrane receptors. The ability of the reported method to access the biogenesis pathways in a high-throughput manner facilitates the identification and evaluation of molecules affecting receptor trafficking.