The CCS Knockout Huh-7 Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal population derived from the Huh-7 hepatocellular carcinoma cell line, featuring targeted disruption of the CCS gene encoding the copper chaperone for superoxide dismutase 1 (SOD1). This polyclonal knockout model was generated using guide RNAs directed against CCS, resulting in a heterogeneous pool of cells with loss-of-function mutations, thereby eliminating functional CCS protein expression. The product provides a powerful tool for investigating copper-dependent antioxidant defense and oxidative stress signaling in a hepatic cellular context without the confounding effects of clonal selection.
The host Huh-7 cell line was originally established from a well-differentiated hepatocellular carcinoma resected from a 57-year-old Japanese male. These cells retain key properties of liver parenchymal cells, including active metabolism and detoxification pathways, and are permissive to hepatitis C virus infection. Huh-7 cells endogenously express vital copper homeostasis machinery and antioxidant systems, making them an ideal platform for studying the interplay between copper trafficking and oxidative stress management in a disease-relevant hepatocyte-derived model. Their robust proliferation and well-characterized signaling networks facilitate high-throughput and mechanistic investigations.
CCS functions as a dedicated copper chaperone that specifically loads Cu(I) ions onto SOD1, a step essential for SOD1 enzymatic activation and its subsequent dismutation of superoxide radicals into hydrogen peroxide and oxygen. Transcriptional regulation of CCS is controlled by metal-responsive transcription factor-1 (MTF-1) and SP1, which respond to intracellular copper levels and oxidative stress via NRF2-mediated pathways. Following CCS-mediated copper insertion, SOD1 acts downstream to mitigate reactive oxygen species (ROS), with hydrogen peroxide further detoxified by catalase and glutathione peroxidase. The CCS?CSOD1 axis is therefore a critical node in cellular antioxidant defense, and its disruption leads to elevated superoxide accumulation that can trigger oxidative damage and apoptosis.
In Huh-7 hepatocarcinoma cells, CCS knockout directly impairs copper delivery to SOD1, resulting in diminished superoxide dismutase activity and increased susceptibility to oxidative insult. This model recapitulates key aspects of human copper metabolism disorders and neurodegenerative conditions such as amyotrophic lateral sclerosis (ALS), where SOD1 dysfunction is implicated. Given the liver??s central role in copper distribution and detoxification, the CCS knockout in Huh-7 cells enables dissection of hepatic mechanisms governing copper homeostasis and ROS detoxification. The model also provides a tractable system for exploring how aberrant antioxidant signaling contributes to hepatocellular carcinoma progression and drug response.
Researchers can employ this polyclonal knockout population for a wide range of functional studies, including assessment of SOD1 activity via in-gel assays, quantification of intracellular copper content, and detection of ROS using fluorescent probes such as DCFDA. Western blotting and RT-qPCR can verify CCS and SOD1 expression changes, while immunofluorescence allows examination of SOD1 localization. Cell viability and apoptosis assays under oxidative stress conditions, coupled with drug screening for candidate SOD1 activators or copper modulators, are readily performed. This product thus supports investigations into oxidative stress biology, copper trafficking, and ALS-relevant pathology. For technical specifications and ordering details, please contact Ascent Research.
