The KLHL36 Knockout NCI-H1975 Polyclonal Cells represent a CRISPR/Cas9-edited polyclonal knockout cell population derived from the NCI-H1975 human lung adenocarcinoma cell line. This product enables loss-of-function studies of the KLHL36 gene, which encodes a substrate-specific adaptor protein for the Cullin-3-RING E3 ubiquitin ligase complex. Through CRISPR/Cas9-mediated gene disruption, the polyclonal pool encompasses a heterogeneous mix of edited alleles, providing a robust model for investigating KLHL36-mediated ubiquitination pathways without the clonal artifacts that can arise from single-cell derived lines.
The host cell line, NCI-H1975, is a widely used non-small cell lung cancer (NSCLC) model established from a female patient with lung adenocarcinoma. These cells harbor activating EGFR L858R and T790M mutations, the latter conferring resistance to first-generation tyrosine kinase inhibitors such as gefitinib and erlotinib. The NCI-H1975 line retains epithelial characteristics and is commonly employed to study mechanisms of acquired drug resistance and to evaluate next-generation EGFR inhibitors. Its well-characterized oncogenic background makes it an ideal platform for dissecting the contributions of ubiquitin ligase adaptors to NSCLC pathophysiology.
KLHL36 functions as a substrate recognition subunit of the CUL3-RBX1 E3 ubiquitin ligase, forming complexes with CUL3, RBX1, and E2 ubiquitin-conjugating enzymes to catalyze the polyubiquitination of target proteins, thereby marking them for proteasomal degradation by the 26S proteasome. The kelch repeat domains of KLHL36 are predicted to mediate interactions with specific substrates, though its downstream targets remain largely uncharacterized. Disruption of KLHL36 perturbs CUL3-mediated ubiquitination dynamics, potentially altering the stability of proteins involved in cell cycle control, apoptosis, and signaling, and thus provides a tool to uncover novel substrates and regulatory mechanisms within the ubiquitin-proteasome system.
In the context of NCI-H1975 cells, KLHL36 knockout offers a unique opportunity to explore how Cullin-RING E3 ligase adaptors influence EGFR-mutant NSCLC phenotypes, including proliferation, drug resistance, and metastatic potential. By eliminating KLHL36, researchers can assess its role in modulating oncogenic signaling downstream of mutant EGFR, and determine whether it contributes to the maintenance of the resistant state. This model is particularly valuable for interrogating the crosstalk between ubiquitin-mediated proteolysis and kinase-driven oncogenesis, and may reveal vulnerabilities that can be exploited therapeutically in T790M-positive lung adenocarcinomas.
These polyclonal knockout cells are suited for a variety of applications, including functional characterization of KLHL36 through western blotting for ubiquitin conjugates and co-immunoprecipitation with CUL3, identification of ubiquitination substrates, and assessment of cellular phenotypes using viability assays (MTS/MTT), colony formation, flow cytometry for cell cycle and apoptosis, and drug sensitivity testing with osimertinib or gefitinib. Additional studies may involve migration and invasion assays or RT-qPCR analysis of downstream gene expression changes. This knockout model serves as a powerful resource for advancing understanding of ubiquitin ligase biology in lung cancer. For further information, please contact Ascent Research.
