The ANKRD44 Knockout HT29 Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout cell population in the HT29 human colorectal adenocarcinoma background. This product delivers a genetically mixed pool of cells harboring targeted disruption of the ANKRD44 gene, bypassing the selection of a single clone and instead reflecting a heterogeneous knockout profile. The polyclonal format is particularly useful for studies requiring a representative loss-of-function model while avoiding clonal artifacts, and it permits the evaluation of gene function in a more physiologically diverse cellular context.
The HT29 parental cell line is an epithelial model derived from a human colorectal adenocarcinoma, extensively employed in cancer biology and signal transduction research. HT29 cells exhibit robust growth characteristics and are well-characterized for studies of DNA damage response, cell cycle regulation, and tumor biology. Their colorectal origin makes them a relevant system for investigating pathways implicated in colorectal cancer pathogenesis, including those governing genomic stability and therapeutic resistance.
ANKRD44 (also designated PPP6R3) operates as a substrate-targeting regulatory subunit of protein phosphatase 6 (PP6), directing the catalytic subunit PPP6C to dephosphorylate specific substrates in the DNA damage response. It facilitates PP6-mediated dephosphorylation of ATM, ATR, CHK1, CHK2, and ??H2AX, thereby modulating checkpoint activation and repair pathway selection. Upstream activators include DNA double-strand breaks, replication stress, ionizing radiation, and chemotherapeutic agents such as cisplatin and doxorubicin. Consequently, ANKRD44 knockout impairs dephosphorylation of these damage signaling proteins, resulting in persistent checkpoint signaling and genomic instability. The PP6 holoenzyme also incorporates the structural subunits PPP6R1 and PPP6R2 and intersects with NF-??B signaling, linking genotoxic stress to inflammatory and survival outputs.
In the HT29 colorectal adenocarcinoma context, ANKRD44 knockout is predicted to compromise the cell’s ability to correctly regulate DNA damage checkpoints, thereby promoting genomic instability??a hallmark of colorectal cancer progression. Since HT29 cells are commonly used to model therapeutic response, this polyclonal knockout population offers a valuable platform to dissect how PP6-mediated dephosphorylation influences sensitivity to platinum-based drugs and topoisomerase inhibitors. The model may reveal synthetic lethal interactions and adaptive mechanisms that cancer cells exploit to survive genotoxic stress, providing insight into resistance pathways.
Researchers can employ these polyclonal knockout cells in various assays. Applications include western blotting for phospho-ATM, phospho-CHK1, and phospho-CHK2; ??H2AX immunofluorescence to quantify DNA breaks; cell viability and colony formation assays with cisplatin or doxorubicin; cell cycle flow cytometry to assess checkpoint integrity; and co-immunoprecipitation of the PP6 complex. The model is also suitable for synthetic lethality screens and phosphatase signaling studies. For further details, please contact Ascent Research.
