The KIFC3 knockout HT29 polyclonal cells are a CRISPR/Cas9-edited polyclonal knockout cell population for studying KIFC3 function in colorectal cancer. This genetically disrupted model provides a heterogeneous HT29 pool lacking KIFC3 protein activity, suitable for population-based assays of impaired microtubule-based transport and signaling.
HT29 is an adherent human colorectal adenocarcinoma line derived from a primary tumor of a 44-year-old female. It contains mutations in APC and TP53, resulting in constitutive Wnt/??-catenin signaling and widely used for colon cancer biology and drug response studies.
KIFC3 encodes a minus-end-directed kinesin-14 motor protein that transports cargo along microtubules, with roles in mitotic spindle organization, vesicle trafficking, and cellular polarization. In colorectal cancer, KIFC3 promotes proliferation and migration. It is transcriptionally upregulated by the ??-catenin/TCF complex and modulated by MYC, CDK1, Cyclin B, and Rho GTPases. KIFC3 interacts with cytoplasmic dynein (DYNC1H1), dynactin (DCTN1), Rab11, NuMA, and the centralspindlin complex to coordinate retrograde transport. Downstream, KIFC3 influences mitotic spindle integrity, cytokinesis machinery, vesicle delivery to the cell periphery, E-cadherin-mediated adhesion, and ??-catenin nuclear translocation. Disruption of KIFC3 impairs minus-end-directed microtubule transport, leading to defective spindle assembly, reduced proliferation, and altered Wnt/??-catenin pathway output.
In HT29 cells, APC truncation leads to ligand-independent ??-catenin accumulation and TCF/LEF-driven transcription, including KIFC3 expression. Knockout of KIFC3 in this context uncouples Wnt-dependent gene expression from microtubule-based processes essential for mitosis and migration. This model therefore enables investigation of how loss of kinesin-14 function compromises colon cancer cell fitness, particularly under oncogenic stress, and allows assessment of KIFC3-dependent mechanisms in spindle assembly, cytokinesis, and polarized cell movement.
Applications include KIFC3 knockout confirmation by Western blotting and RT-qPCR, immunofluorescence analysis of microtubule and spindle structures, MTT/CCK-8 viability assays, Transwell migration and invasion assays, and flow cytometry for cell cycle profiling. Transcriptome-wide RNA-seq, TOP/FOP Flash Wnt reporter assays, co-immunoprecipitation of interaction partners, and apoptosis assays are also enabled. Researchers are invited to contact Ascent Research for further details.
