The DPH5 knockout SK-HEP-1 polyclonal cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the human SK-HEP-1 liver adenocarcinoma cell line. This loss-of-function model disrupts DPH5, which encodes the enzyme catalyzing trimethylation in diphthamide biosynthesis. The polyclonal format provides a heterogeneous pool of gene-edited alleles, enabling population-level studies without single-cell cloning. These cells are suited for investigating DPH5 loss in a hepatic context, including effects on protein translation and toxin sensitivity.
The SK-HEP-1 cell line originated from ascitic fluid of a male patient with liver adenocarcinoma and displays adherent epithelial morphology. It serves as a hepatocellular carcinoma model and also exhibits liver sinusoidal endothelial characteristics, making it valuable for liver cancer and endothelial biology research. The tumorigenic nature of SK-HEP-1 cells enhances their relevance in oncological studies. DPH5 knockout in this background allows examination of diphthamide-related processes specifically within a malignant liver environment.
DPH5 catalyzes the trimethylation of a histidine residue on elongation factor 2 (EEF2) to produce diphthine, a key intermediate in diphthamide biosynthesis. This modification is critical for translational fidelity and forms the target site for ADP-ribosylation by diphtheria and Pseudomonas exotoxins. DPH5 functions within a multi-enzyme complex that includes DPH1, DPH2, DPH3, DPH4, DPH6, and DPH7, which sequentially modify EEF2. Disruption of DPH5 leads to loss of diphthamide modification, altering translational control and toxin sensitivity.
In the SK-HEP-1 liver adenocarcinoma background, DPH5 knockout cells provide a clinically relevant system to explore diphthamide modification in hepatocellular carcinoma. Given the liver??s role in protein synthesis, DPH5 deficiency may reveal vulnerabilities in translation exploited by cancer cells. This model also enables assessment of how diphthamide loss impacts sensitivity to bacterial toxins, informing targeted toxin-based therapies. The polyclonal population retains the genetic heterogeneity typical of tumor-derived lines, facilitating translational research.
Typical applications include studying translational control via polysome profiling and RNA-seq, assessing diphthamide status by western blotting for modified EEF2, and measuring ADP-ribosylation of EEF2 following toxin exposure. Cell viability assays under diphtheria toxin or Pseudomonas exotoxin challenge quantify functional DPH5 loss. RT-qPCR verifies DPH5 transcript levels. This model is valuable for investigating diphthamide deficiency disorders, neurological impairments linked to translational dysregulation, and bacterial toxin susceptibility in a liver context. For further technical details, please contact Ascent Research.
