HEXB Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HAP1 cell line. This product provides a functional loss-of-function model for the HEXB gene, which encodes the beta subunit of the lysosomal enzyme beta-hexosaminidase. The polyclonal format ensures a heterogeneous knockout population, suitable for downstream pooled assays and functional screenings. By disrupting HEXB, researchers can investigate the enzyme’s role in glycosphingolipid catabolism without introducing clonal variation.
The HAP1 cell line is a human near-haploid adherent cell line derived from the KBM-7 chronic myeloid leukemia (CML) cell line. HAP1 cells maintain a stable haploid karyotype, making them an optimal host for genetic perturbation studies, particularly haploid genetic screens. The male origin and near-haploid state allow unambiguous loss-of-function phenotypes due to a single allelic copy of most genes. This background is well-suited for studying lysosomal biology, as HAP1 cells express a functional lysosomal compartment and key autophagy and trafficking machinery.
HEXB encodes the beta subunit of the dimeric enzyme beta-hexosaminidase, which forms homodimers (HEXB) or heterodimerizes with the alpha subunit (HEXA). This enzyme localizes to lysosomes via the mannose-6-phosphate receptor pathway and catalyzes the cleavage of terminal N-acetyl-D-hexosamine residues from GM2 gangliosides and other glycoconjugates. The reaction requires the GM2 activator protein (GM2A) and is essential for proper degradation of sphingolipids within lysosomes. Upstream, HEXB expression is regulated by the MiT/TFE family of transcription factors, including TFEB, TFE3, and MITF, which coordinate lysosomal biogenesis and autophagy in response to lysosomal stress. Downstream, loss of HEXB function leads to accumulation of GM2 gangliosides and other substrates, impaired lysosomal degradation, and cellular dysfunction.
In the HAP1 polyclonal knockout context, disruption of HEXB creates a physiologically relevant model for Sandhoff disease, a GM2 gangliosidosis caused by mutations in HEXB. The near-haploid background simplifies genetic interaction studies, enabling efficient mapping of modifiers or suppressors of lysosomal storage pathology. Because HAP1 cells retain key lysosomal functions and signaling pathways, this knockout model allows investigation of substrate accumulation, enzyme trafficking, and compensatory mechanisms without the complexity of diploid systems. The polyclonal nature further supports population-based assays that require diverse genetic backgrounds.
This knockout cell product is suitable for a wide range of experimental applications, including fluorogenic hexosaminidase activity assays to quantify residual enzymatic function, western blotting to confirm HEXB protein loss, and immunofluorescence microscopy to monitor lysosomal expansion via LAMP1/2 staining. Mass spectrometry can be employed to profile ganglioside accumulation, while RNA-seq enables transcriptomic analysis of lysosomal gene expression changes. Haploid genetic screens leveraging the HAP1 background can identify novel regulators of lysosomal function or therapeutic targets for lysosomal storage disorders. For additional details or technical support, please contact Ascent Research.
