The KLHL25 Knockout HCT 116 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population targeting the KLHL25 gene in the human HCT 116 colorectal carcinoma cell line. This heterogeneous cell pool provides a loss-of-function model for investigating KLHL25-dependent cellular processes without clonal isolation, capturing a range of genetic disruptions. The polyclonal format is particularly suited for studying complex phenotypes where population-level effects are informative.
HCT 116 is a well-established human colorectal carcinoma epithelial cell line characterized by a homozygous KRAS G13D activating mutation, MLH1 deficiency causing high microsatellite instability (MSI-H), and wild-type p53 status. These features make it a relevant model for microsatellite-unstable colorectal cancers, commonly used to explore oncogenic signaling, metastatic mechanisms, and responses to chemotherapeutic agents. Its genetic background provides a clinically pertinent context for studying gene function in cancer biology.
KLHL25 (Kelch-like protein 25) functions as a substrate adaptor for Cullin-3 (CUL3)-based E3 ubiquitin ligase complexes, binding directly to actin and interacting with KEAP1. It modulates the KEAP1-Nrf2 oxidative stress pathway: under homeostatic conditions, KEAP1 promotes Nrf2 ubiquitination and proteasomal degradation; upon oxidative stress or PI3K/AKT pathway activation, KEAP1-mediated repression is relieved, and Nrf2 translocates to the nucleus to transactivate antioxidant response element (ARE)-containing genes. KLHL25??s interaction with actin suggests a role in coupling cytoskeletal organization to redox sensing. Downstream Nrf2 targets include genes encoding antioxidant proteins such as heme oxygenase-1 and NAD(P)H quinone oxidoreductase 1. Additionally, KLHL25 has been implicated in neuronal development, though its primary cancer-relevant functions involve cytoskeletal dynamics and oxidative stress adaptation.
Disruption of KLHL25 in the HCT 116 background is predicted to impair actin cytoskeleton integrity and perturb KEAP1-Nrf2 signaling. This dual disruption may reduce cell migration and invasion capacity while increasing susceptibility to oxidative damage and chemotherapeutic stress. Since HCT 116 cells harbor oncogenic KRAS, which elevates intrinsic oxidative stress, the loss of KLHL25 could unmask synthetic lethal interactions or alter stress-adaptive responses. Consequently, this polyclonal knockout model enables detailed dissection of the interplay between oncogenic signaling, cytoskeletal remodeling, and antioxidant defense mechanisms in colorectal cancer.
Researchers can utilize this model in diverse applications: investigating KLHL25??s role in colorectal cancer progression and metastasis, elucidation of oxidative stress response pathways, and screening for chemotherapeutic sensitivity or resistance. Key assays include Western blotting for KLHL25 and Nrf2, immunofluorescence imaging of the actin cytoskeleton, transwell migration and invasion assays, cell viability under oxidative challenge (e.g., H?O?), quantitative RT-PCR for Nrf2 target genes (HO-1, NQO1), and co-immunoprecipitation to study KLHL25-KEAP1 interactions. For technical inquiries and support, please contact Ascent Research.
