This product consists of a CRISPR/Cas9-edited polyclonal HeLa cell population with targeted disruption of the KNTC1 gene, generating a heterogeneous loss-of-function model for studying kinetochore function and mitotic regulation. The polyclonal format preserves genetic diversity among edited cells, enabling robust assessment of KNTC1-dependent processes without clonal artifacts. These cells provide a powerful tool for dissecting the spindle assembly checkpoint and chromosome segregation mechanisms.
HeLa cells, derived from a human cervical adenocarcinoma, are characterized by HPV-18 integration, which inactivates the tumor suppressors p53 and Rb. This immortalized line is a cornerstone of mitosis research due to its rapid proliferation and well-defined karyotypic abnormalities, including aneuploidy. The HPV-driven transformation sensitizes these cells to perturbations in mitotic control, making them an ideal background for interrogating KNTC1 function.
KNTC1 (also known as Rod) is a core subunit of the RZZ (Rod-ZW10-Zwilch) complex, which assembles at kinetochores during prometaphase. Its kinetochore localization is regulated by mitotic kinases including PLK1, Aurora B, and CDK1. Once at the kinetochore, the RZZ complex recruits dynein-dynactin through direct interactions with ZW10 and Zwilch, facilitating microtubule capture and the stripping of spindle checkpoint proteins such as MAD1, MAD2, and BUB1, ultimately silencing the checkpoint and enabling anaphase transition.
In the HeLa context, where baseline mitotic fidelity is already compromised, KNTC1 disruption exacerbates chromosome misalignment, spindle checkpoint activation, and aneuploidy. This knockout model therefore provides a sensitized system to study the interplay between RZZ-mediated kinetochore-microtubule attachment and the SAC. It is particularly valuable for dissecting how cancer cells cope with mitotic stress and for identifying synthetic lethal interactions with checkpoint inhibitors or microtubule poisons.
Researchers can employ this polyclonal knockout population in a variety of assays, including high-content immunofluorescence microscopy to quantify kinetochore localization and chromosome congression defects, flow cytometry to assess changes in ploidy and cell cycle distribution, and live-cell time-lapse imaging to track mitotic progression in real time. Co-immunoprecipitation and immunoblotting experiments allow characterization of RZZ complex integrity and dynein-dynactin interaction dynamics. Moreover, these cells are suitable for drug sensitivity profiling with anti-mitotic agents like paclitaxel or with small-molecule inhibitors targeting the SAC kinase MPS1. For additional information or custom applications, please contact Ascent Research.
