Protein folding and quality control in the endoplasmic reticulum: Recent lessons from yeast and mammalian cell systems

Abstract

The evolution of eukaryotes was accompanied by an increased need for intracellular communication and cellular specialization. Thus, a more complex collection of secreted and membrane proteins had to be synthesized, modified, and folded. The endoplasmic reticulum (ER) thereby became equipped with devoted enzymes and associated factors that both catalyze the production of secreted proteins and remove damaged proteins. A means to modify ER function to accommodate and destroy misfolded proteins also evolved. Not surprisingly, a growing number of human diseases are linked to various facets of ER function. Each of these topics will be discussed in this article, with an emphasis on recent reports in the literature that employed diverse models.

Figure 1. Early events during protein folding and quality control in the mammalian ER

(1) Nascent polypeptides enter the ER through the Sec61 translocon complex, which contains ∼20 components (some of which are shown). Next, a core oligosaccharide, GlcNAC₂Man₉Glc₃, is cotranslationally transferred from a dolichol lipid precursor (orange oval) to an NXS/T (X≠P) consensus site by OST. Terminal glucoses (green triangles) are trimmed by glucosidase I and II (Glc I and Glc II) to a GlcNAC₂Man₉Glc₁ structure that binds the membrane protein, calnexin (CNX). Non-glycosylated substrates either bind BiP and undergo non-lectin mediated folding, and/or are posttranslationally glycosylated by the OST(B) isoform and thereby enter the CNX binding cycle. If translocation is aborted, substrates are delivered to cytosolic chaperones (e.g., Hsp70) and may be degraded. (2) Upon release from CNX, N-linked glycans are further deglucosylated by GlcII, and the substrate either folds spontaneously, is transferred to BiP-, PDI-, and/or Grp94-containing chaperone complexes, or is recognized as misfolded by UGT1 and reglucosylated. Reglucosylation enables rebinding to CNX and its lumenal homolog calreticulin (CRT), which stimulate isomerization and disulfide bond formation via the associated ERp57 PDI homolog. (3) When folded, the substrate is packaged into COPII-coated vesicles for transport to the Golgi. (4) During this process, certain terminal mannose residues (yellow circles) may be subjected to removal by ER Mannosidase I (ManI) and several EDEM homologs. Mannose trimming reduces the affinity for UGT1 and ultimately generates a GlcNAC₂Man₅ structure that binds additional lectins, including XTP3-B and OS-9 (not pictured), which bring the substrate to the retrotranslocation machinery (5) for retro-translocation to the cytosol, ubiquitination by E3 ligases, and degradation by the 26S proteasome. (6) In times of ER stress or excess protein load, misfolded substrates containing GlcNAC₂Man₉ glycans can also be delivered to the Golgi and sorted for lysosomal degradation, or modified by Golgi ManII and returned to the ER for degradation via ERAD.

Figure 2. The mammalian UPR is triggered by a multi-pronged signal transduction pathway

The accumulation of misfolded proteins in the ER and/or liberation of BiP from IRE1, PERK, and ATF6, activate a series of down-stream events. IRE1 induces the splicing of the XBP-1 message, which is then translated into an active transcription factor. There are actually two isoforms of IRE1 (α and β) but for simplicity only a single, generic protein is depicted. PERK phosphorylates eIF2α, which inhibits general protein synthesis but the ATF4 transcription factor can still be translated because of the presence of a short, upstream ORF that is skipped when eIF2α is phosphorylated. In contrast, ATF6, when activated, is released from the ER and migrates to the Golgi where it is clipped, thus releasing an active transcription factor, ATF6-p50. The immediate effects of UPR induction help the ER cope with an increase in the concentration of misfolded proteins. However, prolonged UPR induction triggers apoptosis through the ATF4-dependent transcription of the CHOP/Gadd153 transcription factor. CHOP both increases the expression of pro-apoptotic genes and inhibits the expression of Bcl2, an antiapoptotic protein.

Figure 3. ERAD substrates can be classified into those with misfolded protein lesions either in the ER lumen (ERAD-L), the cytoplasm (ERAD-C), or the membrane (ERAD-M)

The site of the misfolding lesion is depicted by a . Select yeast factors (in parentheses) required for the degradation of each class are listed, and mammalian homologs are indicated in cases in which these have also been shown to function during ERAD, although not necessarily in a class-specific manner. Note: Many mammalian E3s have been identified as contributing to the ERAD of select factors (e.g., CHIP, HRD1, RMA1, gp78, TEB4, and Parkin), but for simplicity these are not shown. In addition, there is evidence that Hsp70 and its constitutively expressed isoform, Hsc70, may play unique roles during protein folding and ERAD, respectively [153].