The ACTR3 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal population of HAP1 cells carrying a targeted disruption of the ACTR3 gene. This loss-of-function model ablates expression of the Arp3 protein, a core subunit of the Arp2/3 actin nucleation complex, and is delivered as an unfractionated polyclonal pool to enable robust population-level phenotypic analyses.
The host HAP1 cell line is a near-haploid, adherent, fibroblast-like derivative of the KBM-7 chronic myeloid leukemia line. Its haploid karyotype facilitates unambiguous genotype-phenotype correlation in CRISPR-generated knockouts, and its p53 deficiency prevents induction of senescence or apoptosis that might otherwise confound functional studies. HAP1 cells have been widely adopted for pooled and arrayed loss-of-function screens, offering a genetically tractable model for interrogating genes involved in fundamental cellular processes.
ACTR3 encodes the Arp3 subunit of the Arp2/3 complex, which nucleates branched actin filaments driving lamellipodia formation, endocytosis, and podosome assembly. The complex is activated by WAVE and WASP family nucleation-promoting factors, themselves regulated by Rac1 and Cdc42 Rho GTPases. Key regulatory inputs include PIP2, CK2 kinase, and the Nck adaptor, while cortactin modulates complex stability. The Arp2/3 complex interacts with actin, capping protein, and the five ARPC subunits (ARPC1?C5) to promote membrane protrusion and vesicle scission downstream of Rho GTPase signaling.
In HAP1 cells, disruption of ACTR3 impairs Arp2/3-mediated actin nucleation, leading to defects in cell spreading, migration, and invasion. The near-haploid background sensitizes the model for identifying genetic interactions and synthetic lethal partners within the actin cytoskeleton network, while the hematopoietic origin makes it relevant for studies of phagocytosis and immune cell function. Combined with p53 deficiency, this system facilitates clean loss-of-function phenotypes, offering insights into cancer metastasis and Wiskott-Aldrich syndrome mechanisms.
Applications include live-cell actin imaging, scratch wound and transwell migration assays, and immunofluorescence with phalloidin to visualize F-actin architecture. Western blotting and co-immunoprecipitation enable assessment of Arp2/3 complex integrity, and RT-qPCR confirms transcript absence. The polyclonal pool is well-suited for functional genomics screens to identify modulators of cell migration, endocytosis, and pathogen entry, and for validating anti-metastatic drug targets. For further details, please contact Ascent Research.
