DNAJC3 Knockout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HT29 human colorectal adenocarcinoma cell line. This product provides a targeted loss-of-function model in which the DNAJC3 gene is disrupted via CRISPR/Cas9-mediated gene editing, generating a heterogeneous pool of edited cells suitable for functional studies. The polyclonal format preserves population-level effects of DNAJC3 ablation without single-cell cloning, enabling researchers to assess phenotypic changes in a context that more closely mimics the complexity of tumor cell populations. It serves as a versatile tool for investigating DNAJC3-dependent mechanisms in colorectal cancer biology, particularly under conditions of endoplasmic reticulum (ER) stress.
The host cell line, HT29, is an established human colorectal adenocarcinoma epithelial line originally derived from a 44-year-old Caucasian female. These cells exhibit an epithelial morphology and retain characteristics of intestinal epithelial cells, making them a widely utilized model for studying intestinal cell physiology, colorectal tumorigenesis, and drug responses. HT29 cells display basal activation of some signaling pathways common in colorectal cancer, including those responsive to ER stress and inflammatory cues, providing a relevant backdrop for dissecting DNAJC3 function in a malignancy-associated context.
DNAJC3, also known as P58IPK, acts as a molecular co-chaperone for the ER-resident heat shock protein BiP (HSPA5), facilitating protein folding and attenuating the unfolded protein response (UPR) under proteotoxic stress. In parallel, DNAJC3 functions as a negative regulator of the double-stranded RNA-dependent protein kinase PKR (EIF2AK2), thereby modulating translational control and inflammatory signaling. Upstream, its expression is transcriptionally induced by UPR sensors such as XBP1 and ATF6 upon ER stress, while downstream it interacts with BiP, HSP70, IRE1??, and PERK to dampen stress signaling. Through these interactions, DNAJC3 suppresses PERK-mediated eIF2?? phosphorylation and downstream CHOP (DDIT3) induction, while also limiting PKR-driven NF-??B activation. Thus, DNAJC3 operates at the interface of the UPR and the PKR pathway, coordinating adaptive responses to protein-folding perturbations.
In the HT29 colorectal cancer model, knockout of DNAJC3 removes a critical brake on both the UPR and PKR pathways, potentially rendering cells hypersensitive to ER stress and altering downstream inflammatory responses. Because colorectal tumors often encounter nutrient deprivation, hypoxia, and chemotherapeutic insults that trigger ER stress, DNAJC3 loss may influence tumor cell survival, proliferation, and drug sensitivity. This knockout model therefore enables dissection of how DNAJC3 contributes to UPR-dependent cytoprotection and PKR-mediated signaling in intestinal epithelial cells, shedding light on mechanisms of tumorigenesis and therapeutic resistance.
Key research applications include investigating ER stress response dynamics, screening for ER stress modulators, and exploring UPR-mediated drug resistance in colorectal cancer. Typical assays suited to these polyclonal cells are western blotting for UPR markers (e.g., BiP, CHOP, phospho-eIF2??), RT-qPCR for XBP1 mRNA splicing, co-immunoprecipitation of DNAJC3?CBiP complexes, and cell viability or apoptosis assays under ER stress inducers such as tunicamycin. The pooled knockout population is also ideal for RNA-sequencing studies to capture transcriptomic changes linked to DNAJC3 loss, and for immunofluorescence-based assessment of BiP subcellular distribution. For further technical details, please contact Ascent Research.
