The ARSA Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout population derived from the HAP1 near-haploid human cell line, with targeted disruption of the ARSA gene. This loss-of-function model enables detailed study of ARSA-mediated lysosomal sulfatide metabolism without requiring single-cell cloning.
HAP1 is a near-haploid, adherent cell line with fibroblast-like morphology, originally isolated from the KBM-7 chronic myeloid leukemia line. Most chromosomes exist in a haploid state, except for disomy 8, making it an ideal host for genetic screening. Single-allele disruption yields a functional knockout, simplifying gene function analysis and enabling clean genotype-phenotype correlation.
ARSA encodes arylsulfatase A, a lysosomal enzyme that catalyzes desulfation of sulfatide to galactosylceramide, a reaction requiring the activator protein saposin B. ARSA transcription is driven by TFEB and MITF, key regulators of lysosomal biogenesis. Loss of ARSA causes sulfatide accumulation, disrupting LAMP1-associated lysosomal integrity and activating cathepsins. This impairs galactosylceramide production, essential for oligodendrocyte maturation and myelin stability, leading to demyelination seen in metachromatic leukodystrophy.
In the HAP1 background, ARSA knockout faithfully recapitulates lysosomal sulfatide storage with complete penetrance due to haploidy. The lack of a second allele eliminates confounding variable expression, enabling robust phenotypic assays. This model is suited for high-content screening, using LysoTracker and anti-sulfatide immunofluorescence to monitor lysosomal dysfunction. The fibroblast-like nature also permits myelination co-culture assays with glial cells to assess functional rescue.
Applications include functional genomics of myelination, metachromatic leukodystrophy disease modeling, and high-throughput drug screening for lysosomal storage disorders. The knockout cells support gene therapy vector evaluation and substrate reduction therapy studies. Assays include ARSA activity measurement by p-nitrocatechol sulfate cleavage, Western blot, RT-qPCR, lipidomics, and flow cytometry for lysosomal mass. Co-culture systems enable myelin readouts. For custom solutions, contact Ascent Research.
