The CBS Knockout AGS Polyclonal Cells represent a CRISPR/Cas9-edited polyclonal cell population derived from the human gastric adenocarcinoma cell line AGS, engineered to disrupt the cystathionine beta-synthase (CBS) gene. This polyclonal knockout model contains a heterogeneous mix of mutations within the CBS locus, introduced by non-homologous end joining following Cas9-mediated double-strand breaks, resulting in loss-of-function across the cell population. This product provides a robust system for investigating CBS-dependent transsulfuration and hydrogen sulfide (H2S) biosynthesis in a gastric epithelial background, without the need for single-cell cloning.
The parental AGS cell line was originally established from a gastric adenocarcinoma and serves as a widely accepted in vitro model of gastric epithelial biology and carcinogenesis. These adherent cells exhibit epithelial morphology and retain key characteristics of transformed gastric cells, making them suitable for studying tumor cell proliferation, migration, drug response, and redox regulation. Their utility in gastric cancer research is well-documented, and the knockout version extends this platform to targeted pathway analysis.
CBS encodes the enzyme cystathionine beta-synthase, which catalyzes the pyridoxal phosphate-dependent condensation of homocysteine and serine to generate cystathionine, the first committed step of the transsulfuration pathway. This reaction is critical for the subsequent synthesis of cysteine, glutathione, and the gasotransmitter H2S. CBS activity is allosterically stimulated by S-adenosylmethionine and requires a heme cofactor for proper function. The enzyme is transcriptionally activated by SP1, NF-Y, and HIF1??, and is upregulated by all-trans retinoic acid (ATRA) and elevated homocysteine. Downstream, cystathionine is cleaved by cystathionine gamma-lyase (CTH) to yield cysteine, which supports glutathione production and further generation of taurine and sulfate. H2S produced by CBS and CTH acts as a signaling molecule with roles in vasorelaxation, cytoprotection, and redox balance. The knockout of CBS disrupts this entire cascade, leading to decreased cystathionine, cysteine, glutathione, and H2S levels, while potentially causing homocysteine accumulation and sensitizing cells to oxidative damage.
In the AGS gastric cancer context, CBS loss profoundly alters the cellular redox landscape and homocysteine metabolism. The transsulfuration pathway is a major source of cysteine for glutathione synthesis, and its impairment can deplete antioxidant defenses, making cells more vulnerable to reactive oxygen species (ROS) and chemotherapeutic agents. Additionally, reduced H2S production may impact signaling pathways that regulate cell proliferation, apoptosis, and angiogenesis??processes often dysregulated in gastric adenocarcinoma. This model thus enables researchers to dissect the role of CBS in gastric tumor biology, including the interplay between homocysteine, oxidative stress, and cancer cell survival. It is particularly relevant for studying the metabolic vulnerabilities of gastric cancers that may rely on the transsulfuration pathway for redox homeostasis.
Researchers can employ this CBS knockout polyclonal cell pool in a wide array of functional and phenotypic assays. Standard validation techniques include western blotting and RT-qPCR to confirm CBS protein and mRNA depletion. Quantification of homocysteine, cystathionine, cysteine, and glutathione levels via HPLC or LC-MS provides direct metabolic readouts, while methylene blue-based or fluorescent probes measure H2S production. Functional studies may involve exposing cells to hydrogen peroxide to assess oxidative stress sensitivity, using DCFDA for intracellular ROS detection, and performing Annexin V apoptosis assays. Proliferation (MTS/BrdU) and migration/invasion (Transwell) assays evaluate tumorigenic potential, and drug sensitivity profiling can identify synthetic lethal interactions or resistance mechanisms. This model is a valuable tool for advancing transsulfuration-targeted therapeutic strategies in gastric and other cancers. For additional technical information, please contact Ascent Research.
