The ADGRA3 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited population of near-haploid HAP1 cells with targeted disruption of the ADGRA3 gene, providing a loss-of-function model. This polyclonal pool contains a heterogeneous set of edited alleles, offering a robust system for studying ADGRA3-dependent processes without clonal selection artifacts. Permanent gene disruption is achieved via CRISPR/Cas9, enabling sustained investigation of the receptor??s biological roles.
HAP1 cells originate from the male chronic myeloid leukemia-derived KBM-7 line and maintain a near-haploid karyotype, which ensures that disruption of a single allele results in complete loss of target gene function. This characteristic eliminates compensatory effects from a second allele and simplifies genotype?Cphenotype analyses. HAP1??s stable haploidy and robust proliferation make it a standard platform for genetic screens, and its hematopoietic background is relevant for adhesion and signaling research.
ADGRA3 (GPR125) is an adhesion GPCR that couples cell adhesion to intracellular signaling via heterotrimeric G proteins. Its large extracellular domain binds matrix proteins and undergoes autoproteolytic cleavage to expose a tethered agonist. Activated ADGRA3 signals through G??12/13 and G??q to regulate RhoA, ROCK, adenylyl cyclase, and phospholipase C, modulating actin dynamics and second messengers (cAMP, Ca2+). The receptor interacts with PDZ scaffolds such as MAGI-2 and integrates with Wnt/PCP components including Frizzled and Dishevelled, influencing TCF/LEF transcription factor activity. Thus, ADGRA3 acts at the nexus of adhesion, mechanotransduction, and gene regulation, with implications for neurodevelopment.
In the HAP1 context, ADGRA3 knockout provides a clean genetic background to dissect its specific contributions to adhesion-dependent signaling and cytoskeletal remodeling. The haploid nature eliminates potential masking effects from a wild-type allele, allowing unambiguous study of receptor function. This model is highly relevant to neuropsychiatric research, as ADGRA3 variants have been linked to schizophrenia and bipolar disorder, and its signaling pathways influence neuronal morphogenesis. The knockout cells thus enable systematic investigation of pathogenic mechanisms.
These polyclonal knockout cells support a wide range of functional assays: Western blot and RT-qPCR confirm gene disruption, immunofluorescence and flow cytometry assess protein expression and localization, and cell adhesion assays quantify altered matrix interactions. Downstream signaling can be evaluated via RhoA activation pull-downs, calcium flux measurements, and cAMP assays. Additionally, the cells serve as a tool for Wnt/PCP pathway analysis and drug target validation for ADGRA3-related disorders. For technical inquiries and further support, contact Ascent Research.
