The ABCD4 Knockout Huh-7 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the Huh-7 hepatocellular carcinoma line, featuring targeted disruption of the ABCD4 gene. This polyclonal model enables loss-of-function studies of the lysosomal cobalamin transporter ABCD4 in a hepatic background.
The Huh-7 cell line was established from a hepatocellular carcinoma of a Japanese male and retains expression of liver-specific proteins such as albumin and transferrin. Widely used for modeling liver metabolism, hepatocarcinogenesis, and viral hepatitis, these cells provide a physiologically relevant system for investigating hepatic cobalamin trafficking and one-carbon metabolism.
ABCD4 encodes a lysosomal membrane protein that functions as a cobalamin transporter, mediating vitamin B12 efflux from lysosomes into the cytoplasm. It forms a complex with LMBRD1 and is transcriptionally regulated by TFEB. Downstream, ABCD4 is essential for the activity of methionine synthase (MTR) and methylmalonyl-CoA mutase (MMUT), which depend on cobalamin as a cofactor. The transporter also interacts with MMACHC, a cytoplasmic cobalamin processing enzyme. CRISPR-mediated ABCD4 disruption leads to lysosomal cobalamin sequestration, causing functional cobalamin deficiency that impairs the methionine cycle and methylmalonyl-CoA metabolism, and results in accumulation of homocysteine and methylmalonic acid??hallmarks of the cblJ type of inherited cobalamin disorders.
In Huh-7 hepatocarcinoma cells, ABCD4 knockout disrupts cobalamin-dependent pathways integral to hepatic function, including S-adenosylmethionine synthesis and propionate catabolism. This model recapitulates the biochemical defects of combined methylmalonic acidemia and homocystinuria, cblJ type, providing a platform to study liver-specific metabolic derangements and to evaluate therapeutic strategies targeting lysosomal cobalamin transport.
Applications include modeling cobalamin metabolism disorders, lysosomal transport studies, vitamin B12 trafficking, metabolic engineering, and drug metabolism in liver cells. Representative assays encompass Western blotting, RT-qPCR, intracellular cobalamin quantification, LC-MS/MS-based homocysteine and methylmalonic acid profiling, immunofluorescence for lysosomal markers, and cell viability assays under cobalamin-depleted conditions. For additional product information, please contact Ascent Research.
