This product offers a CRISPR/Cas9-edited polyclonal knockout cell population of the SK-HEP-1 human hepatic adenocarcinoma cell line, featuring targeted disruption of the DNAJB14 gene. DNAJB14 encodes a co-chaperone for Hsp70 and serves as a crucial subunit of the gp78 E3 ubiquitin ligase complex, which is essential for endoplasmic reticulum-associated degradation (ERAD). The polyclonal format provides a genetically diverse knockout pool, facilitating functional studies without the constraints of clonal selection. This model is intended for advanced research into protein quality control, the ubiquitin-proteasome system, and the unfolded protein response (UPR) within a malignant hepatic context.
SK-HEP-1 cells were originally established from the ascites of a liver adenocarcinoma patient and are a widely employed model for hepatocellular carcinoma research. These adherent epithelial cells recapitulate key aspects of hepatic malignancy, including activated stress response pathways, making them an appropriate host for examining ERAD disruption in cancer. Their robust growth characteristics support a range of biochemical and imaging-based assays.
At the molecular level, DNAJB14 functions as a J-domain co-chaperone that facilitates Hsp70 substrate recognition and delivery to the gp78-HRD1 ligase complex, which also includes p97/VCP, derlin, and ubiquitin-conjugating enzymes. This complex coordinates the retrotranslocation and polyubiquitination of misfolded proteins from the ER membrane for subsequent proteasomal degradation. DNAJB14 expression is induced by ER stress through transcription factors such as ATF6 and the IRE1-XBP1 and PERK-eIF2?? axes. Consequently, its knockout disrupts ERAD efficiency, leading to accumulation of misfolded proteins, persistent UPR activation, and downstream effects on targets like CHOP and BiP.
In the SK-HEP-1 background, DNAJB14 knockout provides a platform to study how compromised ERAD influences liver adenocarcinoma cell pathophysiology. Since cancer cells often depend on robust protein quality control to survive oncogenic and metabolic stress, this model allows dissection of adaptive responses, including UPR-mediated survival signals and apoptotic thresholds. It enables investigation of synthetic lethal interactions with proteasome inhibitors or other ER stress-inducing agents.
This polyclonal knockout population supports diverse experimental workflows. Common assays include western blotting for UPR markers (BiP, CHOP), ubiquitination assays to assess substrate clearance, proteasome activity measurements, qPCR for UPR target genes, immunofluorescence for ER morphology, and co-immunoprecipitation to probe interactions among gp78, Hsp70, and p97/VCP. Cell viability assays under ER stress provide functional readouts. The heterogeneous knockout pool offers a comprehensive view of gene disruption effects. For additional information or custom project inquiries, please contact Ascent Research.
