The CCNO Knockout HAP1 Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout cell population targeting the CCNO gene in HAP1 cells. This polyclonal pool preserves genetic diversity while ensuring effective CCNO disruption, creating a reliable loss-of-function model. Designed for functional genomics, it supports high-throughput screening and pathway analysis without the constraints of single-cell isolates. The pooled editing strategy balances experimental reproducibility with biological complexity, making it suitable for diverse applications.
HAP1 is a near-haploid cell line derived from the KBM-7 chronic myeloid leukemia (CML) blast phase. It retains a near-haploid karyotype, with disomy restricted to chromosome 15, and harbors the BCR-ABL1 fusion oncogene typical of CML. The reduced genome copy number eliminates confounding allelic effects, making it ideal for CRISPR-based gene disruption studies. HAP1 cells are extensively used in functional genomics due to their facile editing and well-characterized signaling pathways. Although non-ciliated under standard conditions, HAP1 expresses core centriole duplication proteins, enabling investigation of deuterosome-mediated pathways when combined with differentiation protocols or ectopic expression of ciliogenic factors.
CCNO encodes Cyclin O, a cyclin specifically regulating deuterosome-mediated centriole amplification critical for multiciliogenesis. Cyclin O acts downstream of the transcriptional co-activator MCIDAS (multicilin) and E2F4, and is induced upon Notch signaling inhibition. It assembles into a complex with deuterosome proteins DEUP1 and CCDC78, and potentially associates with CDK2, to recruit centriole assembly factors. This promotes hierarchical loading of PLK4, STIL, and SAS-6 to initiate procentriole formation, with CPAP required for centriole elongation. Consequently, CCNO disruption blocks deuterosome-driven centriole amplification, impairing motile cilia production and mucociliary clearance in multiciliated cells.
In the HAP1 background, CCNO knockout provides a clean genetic canvas to dissect deuterosome-dependent centriole amplification independent of parallel pathways. While HAP1 cells are not ciliated, forced expression of MCIDAS or serum-starvation protocols can induce centriole amplification and ciliogenesis, enabling functional readouts. The absence of Cyclin O permits unambiguous interrogation of the MCIDAS?CE2F4?Cdeuterosome regulatory axis and its downstream effectors. This model supports comparative studies with canonical centriole duplication, and facilitates identification of novel components or chemical modulators of multiciliogenesis. The isogenic nature of HAP1 further ensures reproducibility across experimental replicates.
Key applications include functional dissection of multiciliogenesis, disease modeling for primary ciliary dyskinesia (PCD), and high-throughput screening for ciliopathy therapeutics. Endpoint assays encompass immunofluorescence for ciliary markers (acetylated tubulin, ARL13B), centriole counting via centrin or CP110 foci, western blot for deuterosome proteins (DEUP1, CCDC78), and RT-qPCR of ciliated cell markers. Under differentiation conditions, mucociliary clearance assays provide a functional readout. The polyclonal knockout format also supports pooled CRISPR screens and epistasis analyses to map centriole amplification pathways. For technical inquiries and ordering details, please contact Ascent Research.
