The HERC2 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal population in which the human HERC2 gene has been disrupted. This genetically defined loss-of-function model abolishes HERC2 protein expression, enabling dissection of its cellular functions without the confounding influence of wild-type activity. As a polyclonal knockout, the cell pool maintains population heterogeneity while ensuring robust target-gene inactivation, avoiding clonal selection artifacts. This format is ideal for functional genomics, pathway analysis, and screening applications where consistent knockout across the population is required.
The host cell line, HAP1, is a near-haploid human cell line derived from the chronic myeloid leukemia cell line KBM-7. Its haploid karyotype allows complete gene knockout by disrupting a single allele, yielding unequivocal loss-of-function phenotypes. HAP1 cells retain functional DNA damage response, cell cycle checkpoint, and apoptotic pathways, making them a versatile platform for studying cancer-relevant signaling. The haploid nature also simplifies genetic manipulation and phenotypic interpretation, facilitating high-throughput and mechanistic studies.
HERC2 encodes an E3 ubiquitin-protein ligase that orchestrates DNA double-strand break repair and other processes. Activated by ATM and ATR kinases, HERC2 acts upstream of RNF8, promoting ubiquitination of histones H2A and H2AX to recruit BRCA1 to damage sites. Additionally, HERC2 binds MDM2 and p53, regulating p53 stability and activity to control cell cycle arrest and apoptosis. Interactions with USP20 and NEURL4 further modulate HERC2??s stability. Beyond the DDR, HERC2 is essential for centrosome duplication and melanogenesis, with mutations linked to oculocutaneous albinism type 2, autism spectrum disorder, and intellectual disability. Thus, HERC2 integrates DNA repair, cell cycle progression, and developmental signaling.
Disrupting HERC2 in the haploid HAP1 background allows unambiguous attribution of phenotypes to HERC2 loss. The absence of a second allele prevents compensatory upregulation, ensuring clear readouts of impaired BRCA1 loading, defective histone ubiquitination, and altered p53 dynamics. This model is therefore well-suited for interrogating the ATM/ATR?CHERC2?CRNF8?CBRCA1 signaling axis, as well as HERC2??s roles in centrosome integrity and genome maintenance. Moreover, the leukemic origin provides a context for studying HERC2-dependent drug resistance and checkpoint control in a cancer cell model.
Typical experimental applications include Western blotting for ??H2AX and p53, co-immunoprecipitation to assess ubiquitination, flow cytometry for cell cycle distribution, comet assays for DNA damage quantification, and immunofluorescence microscopy to visualize BRCA1 foci. Clonogenic survival assays can evaluate sensitivity to DNA-damaging agents. This polyclonal knockout cell population is suitable for large-scale screens, CRISPR modifier studies, and detailed mechanistic dissections of ubiquitin signaling. For technical support or custom applications, please contact Ascent Research.
