The CD320 Knockout HEK293T Polyclonal Cells are a CRISPR/Cas9-mediated polyclonal knockout population of human embryonic kidney cells with targeted disruption of the CD320 gene. This loss-of-function system permits dissection of CD320-dependent receptor endocytosis and intracellular vitamin B12 (cobalamin) homeostasis. The polyclonal format provides a heterogeneous CD320-deficient pool, avoiding clonal artifacts inherent to monoclonal selection.
HEK293T cells, derived from HEK293, express the SV40 large T antigen for high transfection efficiency and robust plasmid replication. As adherent epithelial cells of human embryonic kidney origin, they exhibit metabolic and transport competence suitable for studying nutrient uptake pathways and endocytic trafficking. Their widespread use in biochemical and pharmacological assays makes them a reliable host for interrogating CD320 function.
CD320 encodes a transmembrane receptor that binds the transcobalamin II?Ccobalamin (TCII-Cbl) complex, triggering clathrin-mediated endocytosis and lysosomal delivery of vitamin B12. Released cobalamin serves as a cofactor for methionine synthase (MTR) in the cytoplasm and methylmalonyl-CoA mutase (MUT) in mitochondria, driving one-carbon metabolism and propionyl-CoA processing. CD320 interacts with the endocytic receptors cubilin and megalin, lysosomal transporter LMBD1, and clathrin adaptors. Disruption of CD320 reduces cobalamin supply, impairing MTR-dependent methionine synthesis and MUT-mediated succinyl-CoA production, with potential impacts on DNA methylation and cellular energetics.
This polyclonal CD320 knockout model in HEK293T cells recapitulates features of cobalamin deficiency disorders such as Imerslund-Gr?sbeck syndrome, pernicious anemia, methylmalonic aciduria, and homocystinuria. The epithelial background and metabolic adaptability of HEK293T cells enable investigation of how impaired vitamin B12 uptake alters metabolic flux, mitochondrial function, and one-carbon unit distribution??processes central to the pathophysiology of these conditions.
Researchers can employ radiolabeled vitamin B12 uptake assays, western blotting for CD320 and downstream targets, and RT-qPCR for metabolic gene expression to characterize the knockout. Immunofluorescence microscopy visualizes endocytic trafficking, while B12-deficient culture conditions and cell viability assays reveal functional consequences. Metabolomic profiling of methionine and succinyl-CoA levels further delineates metabolic disruptions. This tool supports advances in cobalamin biology and therapeutic development for B12-related diseases. For technical information, contact Ascent Research.
