N‐glycan processing selects ERAD‐resistant misfolded proteins for ER‐to‐lysosome‐associated degradation

Efficient degradation of by‐products of protein biogenesis maintains cellular fitness. Strikingly, the major biosynthetic compartment in eukaryotic cells, the endoplasmic reticulum (ER), lacks degradative machineries. Misfolded proteins in the ER are translocated to the cytosol for proteasomal degradation via ER‐associated degradation (ERAD). Alternatively, they are segregated in ER subdomains that are shed from the biosynthetic compartment and are delivered to endolysosomes under control of ER‐phagy receptors for ER‐to‐lysosome‐associated degradation (ERLAD). Demannosylation of N‐linked oligosaccharides targets terminally misfolded proteins for ERAD. How misfolded proteins are eventually marked for ERLAD is not known. Here, we show for ATZ and mutant Pro‐collagen that cycles of de‐/re‐glucosylation of selected N‐glycans and persistent association with Calnexin (CNX) are required and sufficient to mark ERAD‐resistant misfolded proteins for FAM134B‐driven lysosomal delivery. In summary, we show that mannose and glucose processing of N‐glycans are triggering events that target misfolded proteins in the ER to proteasomal (ERAD) and lysosomal (ERLAD) clearance, respectively, regulating protein quality control in eukaryotic cells.