The ABCC2 Knockout HT29 Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout population derived from the HT29 human colorectal adenocarcinoma cell line, engineered to disrupt the ABCC2 (MRP2) gene. This product comprises a heterogeneous cell pool with diverse loss-of-function mutations, collectively abolishing MRP2 protein expression while avoiding the artifacts of clonal selection. It serves as a versatile tool for studying transporter function in an intestinal epithelial context.
HT29 is an epithelial cell line isolated from a primary colorectal adenocarcinoma, widely employed as an in vitro model of the intestinal epithelium. It forms polarized monolayers featuring tight junctions and apical microvilli, endogenously expressing an array of transporters and phase I/II enzymes that make it particularly suitable for drug absorption, metabolism, and efflux investigations.
ABCC2 encodes the multidrug resistance-associated protein 2 (MRP2), an apical membrane ATP-binding cassette transporter that actively effluxes a broad range of organic anions, including bilirubin diglucuronide, glutathione conjugates, and therapeutic agents such as methotrexate, statins, and cisplatin. ABCC2 transcription is tightly controlled by ligand-activated nuclear receptors: FXR, PXR, and CAR form heterodimers with RXR to bind promoter elements, integrating bile acid and xenobiotic signals; additionally, the oxidative stress-responsive transcription factor Nrf2 activates ABCC2 via antioxidant response elements as part of the glutathione conjugation detoxification system. Proper apical localization of MRP2 depends on interactions with PDZK1, a scaffolding protein with PDZ domains, and radixin, an ERM protein that links MRP2 to the actin cytoskeleton. These molecular interactions ensure efficient efflux, thereby preventing intracellular accumulation of toxic compounds and contributing to chemoresistance.
In the HT29 colorectal cancer background, ABCC2 knockout eliminates the primary apical conduit for organic anion excretion, sensitizing cells to bile acid toxicity, oxidative stress, and a variety of chemotherapeutics. This functional disruption mirrors the genetic defect in Dubin-Johnson syndrome, providing a cell-based model for hyperbilirubinemia and for interrogating MRP2-dependent drug resistance in intestinal tumors.
This knockout pool supports diverse research applications: quantitative efflux assays using fluorescent substrates like CDCF; drug sensitivity profiling against MRP2 substrates including methotrexate, cisplatin, and statins; confocal immunofluorescence to evaluate apical membrane localization and interactions with PDZK1 or radixin; and bile acid or glutathione conjugate transport assays to measure detoxification pathway flux. Upstream regulatory mechanisms can be dissected using nuclear receptor agonists or Nrf2 inducers. Knockout is typically verified by western blotting for MRP2 and RT-qPCR for ABCC2 mRNA. For additional details, please contact Ascent Research.
