The JOSD1 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population enabling loss-of-function studies of the JOSD1 gene. JOSD1 encodes a Lys-63-specific deubiquitinase that regulates membrane protein trafficking and autophagy. This polyclonal format provides a heterogeneous pool of HAP1 cells with targeted disruption of JOSD1, reflecting genetic diversity and minimizing clonal bias. The model offers a robust tool for investigating deubiquitinase-dependent pathways in a near-haploid genetic background.
HAP1 is a human near-haploid fibroblast-like cell line derived from the KBM-7 chronic myeloid leukemia line. Its haploid karyotype simplifies genetic analyses by eliminating allelic variation, making it ideal for functional genomics and drug screening. The adherent cells retain key signaling networks, including those governing endocytosis and autophagy, and support high-content imaging. HAP1 provides a clean cellular chassis for precise interrogation of gene function.
JOSD1 removes Lys-63-linked polyubiquitin chains from substrates such as EGFR and BECN1, preventing their degradation and promoting receptor recycling and autophagy. The enzyme is regulated by mTOR signaling and cellular stress, and it interacts with the VPS34 complex components ATG14L and UVRAG to stabilize BECN1 and facilitate autophagosome formation. JOSD1 thus couples nutrient sensing to membrane trafficking and proteostatic control.
In the knockout model, loss of JOSD1 leads to enhanced degradation of EGFR and BECN1, causing reduced surface receptor expression and impaired autophagic flux. These cells display decreased LC3-II puncta and are susceptible to defects in endocytic recycling, recapitulating key aspects of JOSD1 dysfunction. The polyclonal nature ensures that phenotypic outcomes are robust and reproducible, supporting research into cancer and neurodegeneration.
This knockout model is suitable for functional characterization of deubiquitinases, cancer biology, autophagy studies, and DUB inhibitor drug discovery. Researchers can employ Western blotting for EGFR and LC3-II, immunofluorescence for LC3 puncta analysis, flow cytometry for surface receptors, and autophagic flux assays with bafilomycin A1. The cells also support proliferation assays and co-immunoprecipitation to assess ubiquitination status. For more information, contact Ascent Research.
