Protein unfolding is not a prerequisite for endoplasmic reticulum-to-cytosol dislocation.

We examined the effects of protein folding on endoplasmic reticulum (ER)-to-cytosol transport (dislocation) by exploiting the well-characterized dihydrofolate reductase (DHFR) domain. DHFR retains the capacity to bind folate analogues in the lumen of microsomes and in the ER of intact cells, upon which it acquires a conformation resistant to proteinase K digestion. Here we show that a Class I major histocompatibility complex heavy chain fused to DHFR is still recognized by the human cytomegalovirus-encoded glycoproteins US2 and US11, resulting in dislocation of the fusion protein from the ER in vitro and in vivo. A folded state of the DHFR domain does not impair dislocation of Class I MHC heavy chains in vitro or in living cells. In fact, a slight acceleration of the dislocation of DHFR heavy chain fusion was observed in vitro in the presence of a folate analogue. These results suggest that one or more of the channels used for dislocation can accommodate polypeptides that contain a tightly folded domain of considerable size. Our data raise the possibility that the Sec61 channel can be modified to accommodate a folded DHFR domain for dislocation, but not for translocation into the ER, or that a channel altogether distinct from Sec61 is used for dislocation.