This product consists of a CRISPR/Cas9-edited polyclonal knockout cell population derived from the human NCI-H1975 lung adenocarcinoma epithelial cell line, in which the DNAJB14 gene has been disrupted. The polyclonal population comprises a heterogeneous pool of cells carrying diverse loss-of-function edits, providing a robust model to study gene function without clonal bias. DNAJB14 encodes an endoplasmic reticulum (ER) co-chaperone that collaborates with Hsp70 family proteins to maintain proteostasis, and its ablation allows direct interrogation of ER quality control mechanisms in a cancer model.
The NCI-H1975 cell line was established from the pleural effusion of a non-smoking female with lung adenocarcinoma and harbors the activating EGFR L858R point mutation together with the T790M gatekeeper mutation, which confers resistance to first-generation EGFR tyrosine kinase inhibitors. These well-characterized cells are widely used as a preclinical model to investigate EGFR-mutant non-small cell lung cancer (NSCLC) biology, drug resistance mechanisms, and the tumor cell response to endoplasmic reticulum stress. The epithelial origin and defined genetic background make this line particularly suitable for dissecting oncogenic signaling and its intersection with cellular stress pathways.
DNAJB14 is a J-domain-containing co-chaperone localized to the ER lumen, where it recruits Hsp70 chaperones such as HSPA5 (BiP) and HSPA8 to facilitate de novo protein folding, translocation, and assembly. Upon accumulation of misfolded proteins, the unfolded protein response (UPR) sensors IRE1, PERK, and ATF6 transcriptionally upregulate DNAJB14 as part of an adaptive program to restore ER homeostasis. DNAJB14 coordinates with the HSPA5-HSPA8 chaperone cycle to direct terminally misfolded clients toward the ER-associated degradation (ERAD) machinery, thereby preventing proteotoxic stress. Disruption of DNAJB14 functionally uncouples Hsp70 activity from ER quality control, leading to accumulation of unfolded polypeptides and potentiation of UPR signaling through the IRE1-XBP1 and PERK-ATF4-CHOP arms. Downstream consequences may involve alterations in ERAD substrate handling and chaperone network dynamics, impacting cell fate decisions under proteotoxic burden.
In the context of NCI-H1975 cells expressing oncogenic EGFR mutants, which drive heightened protein synthesis and constitutively elevated ER stress, loss of DNAJB14 provides a unique opportunity to dissect the reliance of lung adenocarcinoma on ER proteostatic adaptation. This knockout model enables systematic analysis of how co-chaperone-dependent quality control influences tumor cell proliferation, survival, and sensitivity to EGFR-targeted agents or ER stress?Cinducing chemotherapies. It is particularly valuable for exploring the functional link between the UPR and drug resistance in NSCLC, given the established role of EGFR signaling in modulating stress responses. By interrogating DNAJB14 in a clinically relevant EGFR-mutant background, researchers can uncover vulnerabilities that may be therapeutically exploited.
Researchers can employ these polyclonal knockout cells in a variety of experimental settings, including western blotting to monitor UPR activation markers such as BiP and CHOP, RT-qPCR to quantify XBP1 mRNA splicing, and cell viability assays under treatment with ER stress inducers (e.g., tunicamycin, thapsigargin) or EGFR inhibitors. The model is also suitable for co-immunoprecipitation studies to assess disrupted Hsp70 interactions, immunofluorescence to visualize ER morphology and chaperone localization, and high-content screening for modulators of ER stress responses. Additionally, it serves as a platform for investigating mechanisms of acquired drug resistance in EGFR-mutant adenocarcinoma. For further information or to discuss custom applications, please contact Ascent Research.
