The DNAJC16 Knockout HT29 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal population derived from HT29 human colorectal adenocarcinoma cells, offering a model to study DNAJC16 loss-of-function. This heterogeneous pool, generated by CRISPR/Cas9-mediated disruption of the DNAJC16 gene, contains diverse knockout alleles, avoiding clonal selection bias. It is suited for population-level phenotypic analyses, such as bulk proteomics, pooled functional screens, and drug response profiling under ER stress conditions.
HT29 is a well-characterized colorectal adenocarcinoma cell line with epithelial morphology, widely used in cancer and epithelial biology. These cells retain intestinal epithelial features, including polarized monolayer formation, mucin secretion, and tight junction expression, relevant for barrier function and absorption studies. HT29 cells are highly responsive to unfolded protein response (UPR) triggers, making them an ideal host for investigating ER proteostasis in colorectal cancer.
DNAJC16 encodes a J domain co-chaperone that regulates Hsp70 ATPase activity, primarily interacting with HSPA5 (BiP) and HSPA8, and is involved in protein folding and ER-associated degradation (ERAD). As part of the UPR, DNAJC16 is transcriptionally activated by ATF6 and XBP1 upon ER stress induced by agents like tunicamycin or thapsigargin. It forms complexes with BiP, HSPA8, DNAJB11, and DNAJC3 to coordinate chaperone cycles. Downstream, DNAJC16 modulates UPR target expression, including BiP, HSPA8, DDIT3 (CHOP), and spliced XBP1s, integrating stress signals to regulate apoptosis, survival, and secretory function.
In HT29 cells, DNAJC16 knockout impairs ER proteostasis, sensitizing cells to ER stress-induced apoptosis. Colorectal cancers often exploit the UPR for survival under hypoxia or nutrient stress; loss of this co-chaperone disrupts folding of secretory/membrane proteins, leading to misfolded protein accumulation and a shift from pro-survival to pro-death signaling. This model enables dissection of DNAJC16’s role in ER stress adaptation and protein secretion, with the polyclonal design ensuring phenotypic averaging across genetic backgrounds for robust functional assays.
Applications include mechanistic studies of UPR/ERAD in colorectal cancer, protein folding diseases, and drug screening for ER stress modulators. Key assays include western blotting for BiP, CHOP, and XBP1s; RT-qPCR for XBP1 splicing; apoptosis and viability assays under ER stress; immunofluorescence for ER morphology; and colony formation assays to assess proliferative consequences. This loss-of-function model supports discovery in Hsp70 co-chaperone biology. For further assistance, contact Ascent Research.
