Critical Reviews in Biochemistry and Molecular Biology, 40:75–91, 2005CopyrightcTaylor & Francis Inc.ISSN: 1040-9238 print / 1549-7798 onlineDOI: 10.1080/10409230590918685Search and Destroy: ER Quality Controland ER-Associated Protein DegradationAyaz Sayeedand Davis T. W. NgDepartment of Biochemistryand Molecular Biology,Pennsylvania State University,University Park, PA, USAABSTRACT Proteins synthesized in the endoplasmic reticulum (ER) encounterquality control checkpoints that verify their fitness to proceed in the secretorypathway. Molecules undergoing folding and assembly are kept out of the ex-ocytic pathway until maturation is complete. Misfolded side products that in-evitably form are removed from the mixture of conformers and returned to thecytosol for degradation. How unfolded proteins are recognized and how irre-versibly misfolded proteins are sorted to ER-associated degradation pathwayswas poorly understood. Recent developments from a combination of geneticand biochemical analyses has revealed new insights into these mechanisms.The emerging view shows distinct pathways working in collaboration to filterthe diverse range of unfolded proteins from the transport flow and to divertmisfolded molecules for destruction.KEYWORDS ER quality control, ERAD, protein folding, ubiquitin, proteasome, chaperone,glycosylation, protein degradationINTRODUCTIONNearly all proteins secreted from the cell or resident along the secretory path-way begin their journey within the membranes of the endoplasmic reticulum(ER). They cross or integrate into the membrane through a proteinaceous porecalled the translocon (Johnson & van Waes, 1999; Meacock et al., 2000; Romisch,1999). Physically separated from the cytosol, the ER maintains its own distinctlumenal environment for the specific needs of secretory protein biogenesis. Anextensive array of factors are on hand to carry out essential maturation stepsthat can include signal sequence cleavage, N- and O-linked glycosylation, gly-cosylphosphatidylinositol (GPI)-anchor addition, folding and oligomerization,disulphide bond formation, and isomerization. Since these activities are foundnowhere else along the secretory pathway, the uncontrolled flux of immatureproteins out of the ER would be disastrous. This scenario is averted by a mech-anism termed “ER quality control (ERQC),” which monitors protein foldingand assembly and prevents the transport of immature molecules.TheERmust also contend with proteins that become irreversibly misfolded.Errors in transcription and translation, environmental stress (including oxida-tive damage and unfavorable temperatures), and subunit stoichiometric imbal-ances are some of the leading causes for erroneous protein biogenesis (Ellgaard&Helenius, 2003; Wickner et al., 1999). Up to 30% of all newly synthesizedEditor: Elizabeth CraigAddress correspondence to D. T. W.Ng, Department of Biochemistry andMolecular Biology, Pennsylvania StateUniversity, University Park, PA 16802.E-mail: [email protected] are estimated to be defective (Schubertet al., 2000). Left unchecked, aberrant proteins canbe dire to an organism, as they are inherently toxic(Bucciantini et al., 2002). To neutralize them, cells de-ploy another pathway coupled to ERQC. Termed ER-associated degradation (ERAD), offending moleculesare taken out of folding pathways and targeted fordegradation.The flux through the ER includes soluble proteins,single and multi-spanning integral membrane proteins,and lipid anchored proteins. Misfolded proteins thatemerge from any of these populations can be elimi-nated by ERAD. Regardless of species, they are recog-nized, targeted, and translocated to the ubiquitylationmachinery located on the cytosolic face of the ER formodification (most substrates). The molecules are nextextracted from the membrane and finally degraded bythe 26S proteasome. The process seems simple enough,but the wide variety of substrates made it difficult toimagine that a single pathway could sufficiently moni-tor all molecules. It is now clear that the task is accom-plished through a collaboration of multiple pathways.Investigators are currently undertaking the challenge todelineate these pathways, to understand how the cellarrives at the decision that a protein is misfolded, andhow substrates are targeted to the degradative machin-ery. This review will emphasize these and other emerg-ing concepts and is not intended to be comprehensive.For additional reading, the reader is directed to sev-eral excellent reviews recently published (Ellgaard &Helenius, 2003; Kostova & Wolf, 2003; McCracken& Brodsky, 2003; Sitia & Braakman, 2003; Trombetta& Parodi, 2003).ER QUALITY CONTROL: SORTINGAND RETENTIONProteins synthesized in the ER pose special problemsfor quality assurance. Soluble molecules can diffuse inthe lumenal milieu, whereas more spatially constrainedmembrane proteins can have domains exposed to thelumen, lipid bilayer, and the cytosol. A quality controlsystem must be able to detect and retain the full rangeof unfolded proteins while allowing the progress of ma-ture molecules. Since most nascent secretory proteinseventually fold and traffic out of the ER, the mech-anisms that comprise ERQC appear to be indepen-dent from the decisions to degrade molecules deemedmisfolded.Chaperones: Screeners at the FirstLine of DefensePrecisely how cells sort and retain unfolded pro-teins remains unclear. The best-understood mechanismfound in higher eukaryotes, the calnexin/calreticulincycle, integrates activities of both folding and qualitycontrol to serve a subset of glycoproteins. Calnexin andcalreticulin are homologous lectin chaperones residentin the ER, each with their own substrate preferences.Both bind newly synthesized glycoproteins that con-tain N-linked glycan(s) trimmed to a single terminalglucose residue by glucosidase I and II. While bound,substrates undergo conformational maturation assistedby a variety of folding enzymes (Frenkel et al., 2004;Molinari & Helenius, 1999; Oliver et al., 1997; Pollocket al., 2004). The process continues until the remainingglucose residue is cleaved by glucosidase II and the sub-strate is released. Next, a dedicated folding sensor, UDP-glucose:glycoprotein glucosyl transfrease (GT), sampleseach substrate. Folding intermediates are reglucosylatedby GT to allow another round of lectin binding andfolding. The substrate specificity of GT is notable, asit recognizes partially structured non-native conforma-tions over native