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Visualization of the ER‐to‐cytosol dislocation reaction of a type I membrane protein
Author(s) -
Fiebiger Edda,
Story Craig,
Ploegh Hidde L.,
Tortorella Domenico
Publication year - 2002
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/21.5.1041
Subject(s) - biology , cytosol , membrane protein , membrane , transport protein , microbiology and biotechnology , biophysics , biochemistry , enzyme
The human cytomegalovirus gene products US2 and US11 induce proteasomal degradation of MHC class I heavy chains. We have generated an enhanced green fluorescent protein–class I heavy chain (EGFP–HC) chimeric molecule to study its dislocation and degradation in US2‐ and US11‐expressing cells. The EGFP–HC fusion is stable in control cells, but is degraded rapidly in US2‐ or US11‐expressing cells. Proteasome inhibitors induce in a time‐dependent manner the accumulation of EGFP–HC molecules in US2‐ and US11‐expressing cells, as assessed biochemically and by cytofluorimetry of intact cells. Pulse–chase analysis and subcellular fractionation show that EGFP–HC proteins are dislocated from the endoplasmic reticulum and can be recovered as deglycosylated fluorescent intermediates in the cytosol. These results raise the possibility that dislocation of glycoproteins from the ER may not require their full unfolding.