Article
- The EMBO Journal (2002) 21, 1041 - 1053
- doi:10.1093/emboj/21.5.1041
Subject Categories:
Visualization of the ER-to-cytosol dislocation reaction of a type I membrane protein
Edda Fiebiger1, Craig Story1, Hidde L. Ploegh1 and Domenico Tortorella1
- Department of Pathology, Harvard Medical School, 200 Longwood Avenue, Armenise Building, Boston, MA 02115, USA
Correspondence to:
Hidde L. Ploegh, E-mail: ploegh@hms.harvard.edu
Received 31 August 2001; Accepted 14 January 2002; Revised 14 January 2002
Abstract
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.
Keywords:
- dislocation,
- endoplasmic reticulum,
- HCMV US2,
- HCMV US11,
- proteasomal degradation
