Abstract
Native cargo proteins exit the endoplasmic reticulum (ER) in COPII-coated vesicles, whereas resident and misfolded proteins are substantially excluded from vesicles by a retention mechanism that remains unresolved. We probed the ER retention process using the proteostasis regulator 4-phenylbutyrate (4-PBA), which we show targets COPII protein to reduce the stringency of retention. 4-PBA competes with p24 proteins to bind COPII. When p24 protein uptake is blocked, COPII vesicles package resident proteins and an ER-trapped mutant LDL receptor. We further show that 4-PBA triggers the secretion of a KDEL-tagged luminal resident, implying that a compromised retention mechanism causes saturation of the KDEL retrieval system. The results indicate that stringent ER retention requires the COPII coat machinery to actively sort biosynthetic cargo from diffusible misfolded and resident ER proteins.
Highlights
Resident chaperones and bound folding intermediates make up the majority of endoplasmic reticulum (ER) content and must be retained at that site (Barlowe and Helenius, 2016; Gomez-Navarro and Miller, 2016)
Deletion of p24 genes in yeast causes misfolded proteins and chaperones to escape the ER, with the latter being secreted into the extracellular medium; these effects may be attributable in part to a chronic unfolded protein response (UPR) in the knockout strains (Belden and Barlowe, 2001; Copic et al, 2009; Elrod-Erickson and Kaiser, 1996)
We reasoned that the shortness of the cytoplasmic tail (10–15 cytoplasmic residues) of p24 and ERGIC-53 proteins contributes significantly to tight binding of the FC signal to membrane-associated COPII protein and that this affinity enhancement would be lost in a solution-based biochemical search for the interaction
Summary
Resident chaperones and bound folding intermediates make up the majority of ER content and must be retained at that site (Barlowe and Helenius, 2016; Gomez-Navarro and Miller, 2016). We sought to test the hypothesis that COPII-associated machinery restricts the passive uptake into vesicles of mobile resident complexes and associated misfolded proteins. Candidate machinery for retention includes the p24-family proteins. These highly conserved transmembrane proteins cycle continuously between ER and Golgi membranes as abundant constituents of COPI and COPII vesicles (Kaiser, 2000; Otte et al, 2001; Stamnes et al, 1995). Deletion of p24 genes in yeast causes misfolded proteins and chaperones to escape the ER, with the latter being secreted into the extracellular medium; these effects may be attributable in part to a chronic unfolded protein response (UPR) in the knockout strains (Belden and Barlowe, 2001; Copic et al, 2009; Elrod-Erickson and Kaiser, 1996).
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