The DIO2 knockout HAP1 polyclonal cells are a CRISPR/Cas9-edited human cell population in which the DIO2 gene has been disrupted in a near-haploid chronic myeloid leukemia (CML) background. This polyclonal pool results from gene editing of the type II iodothyronine deiodinase, the enzyme responsible for converting the prohormone thyroxine (T4) to bioactive triiodothyronine (T3). The knockout thus enables loss-of-function studies of local thyroid hormone activation, allowing dissection of T3-dependent signaling pathways and providing a versatile tool for functional genomics and drug discovery.
HAP1 is a human near-haploid CML cell line derived from KBM-7, widely used for haploid genetic screening and CRISPR knockout generation. Its stable haploid karyotype ensures that targeting a single allele suffices for complete loss of function, reducing genetic noise. HAP1 cells support high-throughput techniques and express thyroid hormone receptors and deiodinases, making them an appropriate host for investigating DIO2-mediated signaling.
The DIO2 enzyme is transcriptionally regulated by TSH via the cAMP/PKA/CREB pathway and post-translationally controlled by WSB1-mediated ubiquitination and the VCP/p97 complex. DIO2 catalyzes T4-to-T3 conversion, which activates nuclear receptors THRA/THRB to drive T3-responsive genes like UCP1 and DIO3. Upstream inputs also include TNF-?? and NF-??B, while T3 exerts negative feedback. Knockout of DIO2 abrogates local T3 production, silencing these transcriptional programs and uncoupling cellular responses from TSH signaling.
In the HAP1 context, DIO2 knockout provides a clear genetic model for thyroid hormone biology in a human cancer cell line. Although HAP1 is a leukemia line, it retains functional thyroid hormone receptors, allowing studies of T3 signaling without thyroid-specific cells. The haploid genome ensures uniform loss of DIO2 activity across the polyclonal population, minimizing clonal bias. This model supports the identification of DIO2-dependent metabolic pathways and the testing of chemical modulators, relevant to metabolic disease and hormone resistance.
Applications include functional genomics of thyroid hormone metabolism, high-throughput screening for deiodinase modulators, and mechanistic dissection of T3-mediated gene regulation. Standard assays such as T3 ELISA, RT-qPCR for DIO2 and downstream targets (UCP1, DIO3), Western blotting, thyroid hormone receptor reporter assays, RNA-seq, and T3-dependent proliferation assays are readily applicable. The model also facilitates disease modeling for hypothyroidism, hyperthyroidism, metabolic syndrome, and developmental delay. For additional product details and ordering information, please contact Ascent Research.
