The ATP13A1 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the near-haploid HAP1 human cell line, in which the ATP13A1 gene has been disrupted to abolish functional protein expression. This polyclonal knockout pool provides a robust loss-of-function model for investigating endoplasmic reticulum (ER)-associated processes, particularly those governed by P5A-ATPase activity, without introducing clonal artifacts associated with single-cell isolation.
HAP1 cells originate from KBM-7, a chronic myeloid leukemia (CML) cell line isolated from a male patient, and are characterized by their near-haploid karyotype, which simplifies genetic manipulation and phenotypic analysis. As a neoplastic myeloid cell line, HAP1 retains key signaling pathways relevant to cancer biology, while its haploid state eliminates concerns about gene dosage effects in heterozygotes, making it an ideal background for functional genetics and screening applications.
The ATP13A1 gene encodes an ER-resident P5A-type ATPase that is critically involved in protein quality control, where it facilitates the retro-translocation of misfolded substrates from the ER lumen to the cytosol for proteasomal degradation. ATP13A1 physically and functionally interacts with the Sec61 translocon, the chaperone BiP/GRP78, and components of the HRD1-SEL1L E3 ubiquitin ligase complex, as well as lectins OS9 and XTP3B, to coordinate ER-associated degradation (ERAD). Loss of ATP13A1 impairs ER-to-cytosol dislocation of aberrant proteins, triggering accumulation of unfolded polypeptides and subsequent activation of the unfolded protein response (UPR) through IRE1??, PERK, and ATF6 pathways. This leads to upregulation of downstream effectors such as the transcription factors ATF4 and XBP1s, and the pro-apoptotic factor CHOP/DDIT3, while also perturbing calcium homeostasis due to ATP13A1??s role in regulating ER calcium flux.
In the HAP1 context, ATP13A1 knockout produces a genetically tractable model for dissecting the molecular interplay between ER stress and cellular fate decisions. The near-haploid background simplifies interpretation of UPR signaling dynamics and allows efficient coupling of the knockout phenotype with chemical or genetic screens aimed at identifying modulators of ER stress survival. This model is particularly relevant for studying ATP13A1-related neurodevelopmental disorders, including epilepsy, intellectual disability, and ataxia, as well as broader ER proteostasis perturbations associated with malignancies that rely on adaptive UPR signaling.
The ATP13A1 Knockout HAP1 Polyclonal Cells are well-suited for a variety of experimental approaches, including Western blotting to monitor UPR markers such as BiP and CHOP, RT-qPCR to quantify XBP1 splicing and ATF4/CHOP mRNA levels, immunofluorescence to assess ER morphology changes, and flow cytometry-based apoptosis or viability assays. Additionally, these cells can be employed in calcium imaging studies, proteostasis reporter assays, and sensitivity testing to ER stress-inducing agents like thapsigargin or tunicamycin. Researchers can use this knockout model for functional dissection of ERAD and UPR pathways, CRISPR-based genetic interaction screens, or drug discovery campaigns targeting ER stress resilience mechanisms. For further details, please contact Ascent Research.
