The activity of a human endoplasmic reticulum-associated degradation E3, gp78, requires its Cue domain, RING finger, and an E2-binding site.
Efficient targeting of proteins for degradation from the secretory pathway is essential to homeostasis. This occurs through endoplasmic reticulum (ER)-associated degradation (ERAD). In this study, we establish that a human ubiquitin ligase (E3), gp78, and a specific E2, Ube2g2, are both critically important for ERAD of multiple substrates. gp78 exhibits a complex domain structure that, in addition to the RING finger, includes a ubiquitin-binding Cue domain and a specific binding site for Ube2g2. Disruption of either of these domains abolishes gp78-mediated ubiquitylation and protein degradation, resulting in accumulation of substrates in their fully glycosylated forms in the ER. This suggests that gp78-mediated ubiquitylation is an early step in ERAD that precedes dislocation of substrates from the ER. The in vivo requirement for both an E2-binding site distinct from the RING finger and a ubiquitin-binding domain intrinsic to an E3 suggests a previously unappreciated level of complexity in ubiquitin ligase function. These results also provide proof of principle that interrupting a specific E2-E3 interaction can selectively inhibit ERAD.
Chen
Bo
B
Laboratory of Protein Dynamics and Signaling, Center for Cancer Research, Building 560, Room 22-103, National Cancer Institute, Frederick, MD 21702, USA.
Mariano
Jennifer
J
Tsai
Yien Che
YC
Chan
Anna H
AH
Cohen
Mickael
M
Weissman
Allan M
AM
D020816:Amino Acid Motifs; D001665:Binding Sites; D002460:Cell Line; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006801:Humans; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D025801:Ubiquitin / Q000378:metabolism; D044763:Ubiquitin-Conjugating Enzymes / Q000235:genetics / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D025801:Ubiquitin; C471617:UBE2G2 protein, human; D044763:Ubiquitin-Conjugating Enzymes; D044767:Ubiquitin-Protein Ligases