Abstract
Human cytomegalovirus US2 and US11 target newly synthesized class I major histocompatibility complex (MHC) heavy chains for rapid degradation by the proteasome through a process termed dislocation. The presence of US2 induces the formation of class I MHC heavy chain conjugates of increased molecular weight that are recognized by a conformation-specific monoclonal antibody, W6/32, suggesting that these class I MHC molecules retain their proper tertiary structure. These conjugates are properly folded glycosylated heavy chains modified by attachment of an estimated one, two, and three ubiquitin molecules. The folded ubiquitinated class I MHC heavy chains are not observed in control cells or in cells transfected with US11, suggesting that US2 targets class I MHC heavy chains for dislocation in a manner distinct from that used by US11. This is further supported by the fact that US2 and US11 show different requirements in terms of the conformation of the heavy chain molecule. Although ubiquitin conjugation may occur on the cytosolic tail of the class I MHC molecule, replacement of lysines in the cytosolic tail of heavy chains with arginine does not prevent their degradation by US2. In an in vitro system that recapitulates US2-mediated dislocation, heavy chains that lack these lysines still occur in an ubiquitin-modified form, but in the soluble (cytoplasmic) fraction. Such ubiquitin conjugation can only occur on the class I MHC lumenal domain and is likely to take place once class I MHC heavy chains have been discharged from the endoplasmic reticulum. We conclude that ubiquitinylation of class I MHC heavy chain is not required during the initial step of the US2-mediated dislocation reaction.
Highlights
That lack a partner subunit are retained in the endoplasmic reticulum (ER) and eventually transported to the cytosol for destruction by the proteasome [2, 3]
The folded ubiquitinated class I MHC heavy chains are not observed in control cells or in cells transfected with US11, suggesting that US2 targets class I MHC heavy chains for dislocation in a manner distinct from that used by US11
The HA-tagged class I MHC heavy chains continue to be targeted for proteasomal degradation in an US2- and US11-dependent manner [18]
Summary
That lack a partner subunit are retained in the ER and eventually transported to the cytosol for destruction by the proteasome [2, 3]. The human class I MHC heavy chain is a 43-kDa type I transmembrane glycoprotein with a large ER-lumenal/cytosolic domain and a short cytosolic tail. It is inserted into the ER membrane, glycosylated, and assembled with both the soluble class I MHC light chain or 2 microglobulin and the antigenic peptide. This properly folded and fully assembled trimeric complex is allowed to exit the ER and travel through the molecule, replacement of lysines in the cytosolic tail of secretory pathway to the cell surface. The data presented here show that US2 and US11 carry out their function differently, not-
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