The HERC4 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout population designed to disrupt the HERC4 gene in the HAP1 cell line. This product provides a heterogeneous pool of cells harboring diverse loss-of-function mutations at the target locus, generated by CRISPR/Cas9-mediated gene disruption. Polyclonal knockout pools enable immediate phenotypic analysis without the need for single-cell cloning, offering a rapid and cost-effective approach for functional studies. The cells retain the near-haploid genomic architecture of the parental HAP1 line, ensuring that knockout effects are observed without interference from a second allele.
HAP1 is a human near-haploid cell line derived from the KBM-7 chronic myeloid leukemia line. With the exception of a disomic chromosome 15, HAP1 cells possess a single copy of each chromosome, presenting a simplified genetic landscape that facilitates gene editing and phenotype interpretation. This haploid state minimizes genetic buffering and allows unambiguous assignment of gene function. HAP1 cells are widely adopted in biomedical research for CRISPR-based knockout studies, pathway analysis, and drug target validation due to their stable growth, ease of manipulation, and relevance to hematopoietic malignancies.
HERC4 is a HECT domain-containing E3 ubiquitin ligase central to the ubiquitin-proteasome system. It accepts activated ubiquitin from E2 ubiquitin-conjugating enzymes through a conserved cysteine residue, forming a thioester intermediate before catalyzing ubiquitin transfer to substrate proteins. Polyubiquitinated substrates are then targeted for degradation by the 26S proteasome. Key pathway components include ubiquitin, E1 ubiquitin-activating enzyme, E2 ubiquitin-conjugating enzymes, and proteasome subunits. HERC4 interacts directly with E2 enzymes and may be regulated by phosphorylation. Although downstream substrates remain largely uncharacterized, HERC4 is implicated in protein quality control and has been associated with cancer susceptibility and neurological disorders.
The combination of HERC4 knockout and the HAP1 near-haploid background creates a powerful system to study E3 ligase biology. Because HAP1 cells are functionally haploid, CRISPR-mediated disruption of HERC4 produces a complete loss of protein function without confounding heterozygosity. This enables clear dissection of ubiquitination events and substrate turnover. The CML origin of HAP1 makes this model relevant for studying ubiquitin-proteasome dysregulation in leukemia. Knockout cells can be used to assess proteasome activity, apoptosis, and cell cycle progression, illuminating HERC4??s role in cellular homeostasis and oncogenesis.
Researchers can employ this polyclonal knockout pool for diverse experimental approaches. Western blotting confirms HERC4 loss, while co-immunoprecipitation coupled with mass spectrometry identifies interacting partners and substrates. Ubiquitination assays measure global or target-specific ubiquitination changes. Proteasome activity assays and drug sensitivity screens provide functional readouts. Additional applications include cell proliferation studies and genetic interaction screens to map HERC4 regulatory networks. For further product information and technical support, please contact Ascent Research.
