This product consists of a CRISPR/Cas9-edited polyclonal knockout population of HCT 116 cells carrying a disrupted KLHL18 gene. The polyclonal format provides a heterogeneous mixture of edited alleles, offering a robust loss-of-function model free from clonal selection artifacts. It is intended for researchers exploring ubiquitin-dependent regulation of cell migration and cancer metastasis.
The parental HCT 116 cell line is an epithelial colorectal carcinoma model derived from a human tumor. It displays microsatellite instability (MSI) and contains a KRAS G13D oncogenic driver mutation, reflecting molecular features of aggressive colorectal cancers. HCT 116 cells maintain key oncogenic signaling and are extensively used to study cytoskeletal dynamics and metastatic progression.
KLHL18 is a substrate adaptor of the CUL3?CRBX1 E3 ubiquitin ligase that selectively targets DDA3 (PSRC1) for proteasomal degradation. DDA3 associates with microtubules and actin filaments, and its ubiquitination by KLHL18?CCUL3?CRBX1 promotes microtubule destabilization and actin remodeling, thereby facilitating cell migration. KLHL18 activity is regulated by growth factor receptor signaling and Rho GTPase pathways; specifically, RhoA activation via ROCK can stimulate KLHL18-mediated DDA3 degradation. In this circuit, KLHL18 acts as a central node converting extracellular cues into cytoskeletal rearrangements. Knockout of KLHL18 leads to DDA3 accumulation, which in turn stabilizes microtubules and modulates actin organization, typically impairing cell motility. The interplay between KLHL18, CUL3, RBX1, DDA3, and RhoA thus integrates pathways controlling migration, with potential importance in cancer cell invasion.
Within HCT 116 colorectal cancer cells, KLHL18 disruption enables dissection of ubiquitin-dependent control of metastasis-associated behaviors. Because these cells harbor MSI and an activated KRAS allele, the knockout model permits examination of how KLHL18 loss impacts DDA3-driven cytoskeletal alterations and migratory competence in a clinically relevant genetic background. The polyclonal nature avoids clonal bias and yields data more representative of heterogeneous tumor populations, enhancing the translational relevance of findings. Researchers can assess changes in wound closure, transwell invasion, and the interaction between the ubiquitin-proteasome system and Rho GTPase signaling.
Typical experimental applications include Western blotting and co-immunoprecipitation to monitor DDA3 protein levels and CUL3 complex integrity, alongside RT-qPCR for mRNA quantification. Migration and invasion are assessed by wound healing and Transwell assays, while immunofluorescence microscopy reveals alterations in microtubule networks and focal adhesions. These approaches facilitate screening for small-molecule inhibitors that disrupt the KLHL18?CDDA3 interface, potentially identifying anti-metastatic leads. For further information, technical assistance, or to place an order, please contact Ascent Research.
