The ANAPC16 Knockout HT29 Polyclonal Cells represent a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HT29 human colon adenocarcinoma cell line. This product enables loss-of-function studies of ANAPC16, a core subunit of the anaphase-promoting complex/cyclosome (APC/C). The polyclonal nature of the knockout population captures a heterogeneous disruption of the target gene, providing a robust model for investigating APC/C-dependent processes without single-cell clonal selection. Researchers can utilize this tool to dissect mitotic regulation and ubiquitin-proteasome system dynamics in a colon cancer background.
HT29 cells are a well-established epithelial model isolated from a female patient with colorectal adenocarcinoma. They harbor a TP53 mutation, which contributes to impaired DNA damage checkpoints and chromosomal instability, making them particularly suitable for studying tumorigenic mechanisms. As a colorectal cancer line, HT29 is extensively used in cancer biology, intestinal barrier function research, and anti-cancer drug testing. The knockout of ANAPC16 in this TP53-mutant context offers a unique platform to examine synthetic interactions between cell cycle regulation and tumor suppressor pathways.
ANAPC16 is an integral subunit of the APC/C E3 ubiquitin ligase, which orchestrates ordered progression through mitosis and G1 phase by targeting key regulators for proteasomal degradation. Within the APC/C holoenzyme, ANAPC16 stably associates with structural components including ANAPC2, ANAPC10, and ANAPC11. APC/C activity is temporally controlled by co-activators CDC20 and CDH1, which direct substrate recognition. Critical substrates include Cyclin B1 and securin (PTTG1), whose degradation inactivates CDK1 and triggers anaphase onset. Upstream kinases such as CDK1 and PLK1 phosphorylate APC/C subunits to modulate activity, while EMI1 (FBXO5) and MAD2L2 provide checkpoint-mediated inhibition. Representative pathway components??CDC20, CDH1, Cyclin B1, securin, CDK1-cyclin B, and E2 enzymes UBE2C and UBE2S??converge on APC/C-dependent ubiquitination cascades.
Knocking out ANAPC16 in HT29 cells impairs APC/C assembly or function, leading to accumulation of mitotic substrates and consequent cell cycle dysregulation. This disruption is expected to exacerbate the inherent chromosomal instability of the TP53-mutant background, creating a heightened state of genomic instability. The model enables dissection of how compromised APC/C activity contributes to colorectal cancer progression, resistance to anti-mitotic agents, and aberrant mitotic exit. It also permits exploration of the interplay between APC/C dysfunction and pathways affected by TP53 loss, such as apoptosis and G1/S checkpoint control.
This polyclonal knockout cell population is a versatile resource for multiple research applications. It can be employed to study APC/C-dependent degradation mechanisms using ubiquitination assays, to profile cell cycle distribution by flow cytometry, and to evaluate mitotic arrest via immunofluorescence of mitotic markers. Western blotting for Cyclin B1 and securin provides direct readouts of substrate accumulation. Functional assays such as colony formation and viability testing with nocodazole or other spindle poisons facilitate screening of anti-mitotic therapeutics. Additionally, the cells serve as a tool to investigate genomic instability mechanisms in colorectal cancer. For further information or to discuss customized applications, please contact Ascent Research.
