Dynamics and retention of misfolded proteins in native ER membranes

When co-translationally inserted into endoplasmic reticulum (ER) membranes, newly synthesized proteins encounter the lumenal environment of the ER, wh ich contains chaperone proteins that facilitate the folding reactions neces sary for protein oligomerization, maturation and export from the ER. Here w e show, using a temperature-sensitive variant of vesicular stomatitis virus G protein tagged with green fluorescent protein (VSVG-GFP), and fluorescen ce recovery after photobleaching (FRAP), the dynamics of association of fol ded and misfolded VSVG complexes with ER chaperones. We also investigate th e potential mechanisms underlying protein retention in the ER. Misfolded VS VG-GFP complexes at 40 degrees C are highly mobile in ER membranes and do n ot reside in post-ER compartments, indicating that they are not retained in the ER by immobilization or retrieval mechanisms. These complexes are immo bilized in ATP-depleted or tunicamycin-treated cells, in which VSVG-chapero ne interactions are no longer dynamic, These results provide insight into t he mechanisms of protein retention in the ER and the dynamics of protein-fo lding complexes in native ER membranes.