EDEM2 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population designed to disrupt the EDEM2 gene in the human near-haploid HAP1 cell line. The mixed pool of edited cells carries a range of gene disruptions, providing a robust loss-of-function model without requiring single-cell cloning.
The HAP1 parental line is a male adherent near-haploid cell line derived from chronic myeloid leukemia (CML) and expresses the BCR-ABL oncogene. Its near-haploid karyotype simplifies genetic studies, as a single allele knockout often yields functional null phenotypes. HAP1 cells maintain essential ER quality-control machinery, making them suitable for investigating glycoprotein processing and ER stress pathways.
EDEM2 encodes an ER-resident ??-mannosidase that trims terminal ??1,2-linked mannose residues from N-linked glycans on misfolded glycoproteins, generating a degradation signal recognized by the lectins OS-9 and XTP3-B. This trim-and-expose mechanism tags substrates for dislocation into the cytosol through the HRD1-SEL1L retrotranslocation complex, followed by VCP/p97-mediated extraction and ubiquitin-dependent proteasomal degradation. EDEM2 expression is transcriptionally upregulated by the UPR branches regulated by spliced XBP1, ATF6, and IRE1?? in response to ER stress. EDEM2 physically interacts with ERAD components including SEL1L, HRD1, OS-9, XTP3-B, and PDIA6, and processes clinically relevant substrates such as mutant tyrosinase and the ??1-antitrypsin Z variant.
In the near-haploid HAP1 context, disruption of EDEM2 eliminates the mannose trimming step that commits misfolded glycoproteins to ERAD, leading to their accumulation and constitutive ER stress. This amplifies UPR signaling and sensitizes cells to proteotoxic stress, providing a clear platform for examining ERAD dependency. The BCR-ABL-positive leukemia background further enables investigation of how ERAD dysfunction impacts oncogenic signaling and therapeutic responses.
This knockout model supports diverse applications in glycoprotein quality control, UPR signaling, and protein aggregation research. Key experimental approaches include cycloheximide chase assays to measure glycoprotein half-life, proteasome inhibition with MG-132 or bortezomib to confirm degradation dependence, tunicamycin-induced ER stress to examine adaptive responses, and immunofluorescence for subcellular distribution of substrates. Western blotting, RT-qPCR, and flow cytometry provide complementary quantitative readouts. The model is well-suited for drug discovery screens targeting ER proteostasis and for dissecting the role of EDEM2 in cancer cell survival. For technical assistance, please contact Ascent Research.
