Aim
To elucidate the effects of SEL1L-mediated ERAD on Huh7 and explore the underlying mechanisms in vivo and in vitro.
Background
Proteins play a central role in regulating biological functions, and various pathways regulate their synthesis and secretion. Endoplasmic reticulum-associated protein degradation (ERAD) is crucial for monitoring protein synthesis and processing unfolded or misfolded proteins in actively growing tumor cells. However, the role of the multiple ERAD complexes in liver cancer remains unclear.
Conclusion
ERAD inhibition suppressed the proliferation and migration of Huh7 and promoted its apoptosis. EXT2 plays an important role and ERAD might be a potential treatment for Huh7 hepatocellular carcinoma.
Methods
Huh7 cells were treated with ERAD inhibitor to identify ERAD's role. Cell counting kit-8, 5-ethynyl-2'-deoxyuridine and colony formation experiments were performed. Apoptosis level and migration ability were assessed using fluorescence activated cell sorting and Transwell assay, respectively. Huh7 SEL1L knockout cell line was established via clustered regularly interspaced short palindromic repeats, proliferation, apoptosis, and migration were assessed through previous experiments. The role of SEL1L in vivo and the downstream target of SEL1L were identified using Xenograft and mass spectrometry, respectively.
Results
The ERAD inhibitor suppressed cell proliferation and migration and promoted apoptosis. SEL1L-HRD1 significantly influenced Huh7 cell growth. SEL1L knockout suppressed tumor cell proliferation and migration and enhanced apoptosis. Mass spectrometry revealed EXT2 is a primary substrate of ERAD. SEL1L knockout significantly increased the protein expression of EXT2. Furthermore, EXT2 knockdown partially restored the effect of SEL1L knockout.