The DTX2 Knockout HAP1 Polyclonal Cells product offers a CRISPR/Cas9-edited polyclonal knockout cell population for disruption of the DTX2 gene in the HAP1 human near-haploid cell line. This polyclonal population provides a versatile loss-of-function model for investigating DTX2-dependent regulatory mechanisms in a clean genetic background.
The HAP1 cell line is derived from the KBM-7 chronic myeloid leukemia line and retains a near-haploid karyotype, making it exceptionally suited for genetic knockout studies. Its haploid state permits efficient gene disruption without the confounding influence of a second allele, thereby streamlining functional characterization of target genes. HAP1 cells are widely used in genetic screening and knockout validation, particularly for interrogating cancer-associated pathways and therapeutic targets.
DTX2 encodes an E3 ubiquitin ligase that negatively regulates Notch receptor trafficking and signal transduction. Activation of Notch receptors (NOTCH1-4) by ligands such as DLL1, DLL4, JAG1, and JAG2 triggers receptor proteolysis and release of the Notch intracellular domain (NICD), which complexes with CSL/RBPJ and MAML coactivators to drive transcription of HES1 and HEY1. DTX2 counteracts this pathway by ubiquitinating Notch receptors, targeting them for lysosomal degradation and thereby attenuating signaling. DTX2 interacts with Notch receptors, the paralogs DTX1 and DTX3, and E2 ubiquitin-conjugating enzymes, linking the ubiquitin-proteasome system to endocytic-lysosomal trafficking.
In the HAP1 background, loss of DTX2 disrupts this negative regulatory loop, permitting detailed study of Notch pathway dynamics. Because HAP1 cells are near-haploid, CRISPR/Cas9-mediated gene disruption yields an effective loss-of-function model that simplifies analysis of Notch receptor ubiquitination and turnover. This system can be used to examine changes in Notch receptor surface expression, NICD abundance, and downstream transcriptional responses, as well as potential compensatory activities of DTX1 and DTX3.
Typical applications include dissection of Notch signaling, identification of E3 ligase substrates, and analysis of ubiquitin-dependent receptor sorting. Researchers can employ western blotting to measure DTX2 and Notch target levels, RT-qPCR to quantify HES1/HEY1 transcription, immunofluorescence and flow cytometry to assess Notch receptor localization and surface expression, and luciferase-based Notch reporters. Co-immunoprecipitation can detect ubiquitinated Notch species, and functional assays for proliferation and apoptosis reveal consequences of DTX2 loss in cancer-relevant settings. These polyclonal knockout cells are also suited for haploid genetic screens and drug target validation. For more information, contact Ascent Research.
