The KIFAP3 Knockout HT29 Polyclonal Cells constitute a targeted loss-of-function model generated by CRISPR/Cas9-mediated disruption of the human KIFAP3 gene in the HT29 colorectal adenocarcinoma cell line. This product is provided as a polyclonal knockout cell population, meaning it consists of a heterogeneous pool of edited cells without single-cell clonal isolation. The polyclonal format preserves population-level diversity while ensuring uniform gene disruption across the culture, offering a robust tool for experiments where monoclonal selection is not required. This knockout model is designed to interrogate KIFAP3-dependent molecular mechanisms in an intestinal epithelial context relevant to cancer biology and ciliary signaling research.
The HT29 cell line is a well-characterized human colorectal adenocarcinoma model of epithelial origin, widely employed for studies of intestinal differentiation, mucin production, and polarized monolayer formation. These cells form tight junctions and exhibit characteristics of absorptive enterocytes, making them valuable for investigating barrier function, cell polarity, and tumorigenic processes. The epithelial morphology and ability to polarize provide a physiologically relevant substrate for analyzing the role of primary cilia and associated signaling pathways in colorectal cancer progression.
KIFAP3 encodes kinesin-associated protein 3, a non-motor adaptor subunit of the heterotrimeric kinesin-2 motor complex, which also contains KIF3A and KIF3B motor subunits. This complex is essential for anterograde intraflagellar transport and primary cilia biogenesis. KIFAP3 functions as a scaffold that facilitates cargo binding and regulates motor activity. In HT29 cells, KIFAP3 disruption impairs kinesin-2-dependent trafficking of ciliary components, such as those loaded by the BBSome and IFT complex B proteins. Consequently, primary cilia assembly is compromised, altering downstream Hedgehog and Wnt signaling cascades. Key molecular players include upstream RFX transcription factors and FoxJ1, which transcriptionally regulate ciliary genes; downstream effectors like Smoothened and Gli transcription factors; and representative pathway components such as IFT88, IFT140, BBS4, and ARL13B.
In the context of the HT29 colorectal cancer model, KIFAP3 knockout disrupts primary cilia formation, which is increasingly recognized as a modulator of oncogenic signaling. Loss of cilia may dysregulate Hedgehog and Wnt pathways, potentially influencing tumor cell proliferation, differentiation, and migration. This model therefore enables dissection of cilia-dependent versus cilia-independent contributions to colorectal cancer pathogenesis. Additionally, it provides a platform for studying ciliopathy-associated mechanisms, as KIFAP3 dysfunction is implicated in polycystic kidney disease and other ciliopathies. The HT29 background adds epithelial malignancy relevance, bridging basic cilia biology with translational oncology research.
Researchers can employ this polyclonal knockout model for a variety of advanced applications, including functional analyses of primary cilia in colorectal cancer, mechanistic studies of intraflagellar transport, and screening of small molecules that modulate ciliogenesis. Typical readouts include immunofluorescence detection of acetylated-tubulin and ARL13B to visualize cilia, western blotting for KIFAP3 and downstream targets, RT-qPCR profiling of cilia-related genes, cell cycle analysis, colony formation assays, and migration assays. The model is also suitable for drug response profiling in cilia-deficient cancer cells. For further technical specifications or custom projects, please contact Ascent Research.
