The DNAJB14 Knockout HEK293T Polyclonal Cells represent a CRISPR/Cas9-mediated polyclonal knockout cell population engineered to disrupt the DNAJB14 gene, which encodes a J-domain co-chaperone critical for Hsp70 chaperone function. This loss-of-function model provides an essential tool for examining Hsp70-mediated protein folding and quality control pathways in a human epithelial cell context. The polyclonal format enables robust population-level studies without imposing clonal selection constraints, making it suitable for mechanistic and phenotypic analyses in proteostasis research.
The host cell line, HEK293T, is an immortalized human embryonic kidney epithelial cell line stably expressing the SV40 large T antigen, which enhances plasmid replication and protein expression. Widely employed across biomedical research, HEK293T cells offer ease of genetic manipulation, rapid growth, and high transfection efficiency, providing a consistent and reliable cellular background for knockout studies. Their epithelial origin further permits investigation of chaperone function and stress responses in a physiologically relevant adherent cell model.
DNAJB14 functions as an Hsp40-type co-chaperone that recruits unfolded or misfolded substrates to Hsp70, facilitating ATP hydrolysis and subsequent protein folding, assembly, or degradation. The mechanistic cycle involves DNAJB14 binding exposed hydrophobic patches on client proteins, interacting with Hsp70, and stimulating its ATPase activity, a process tightly regulated by the heat shock transcription factor HSF1 under conditions of heat shock and unfolded protein stress. Representative pathway components thus include DNAJB14, Hsp70, ATP, and unfolded substrates, with upstream activation through HSF1-mediated transcriptional upregulation. DNAJB14 is also known to interact with other Hsp40 co-chaperones, positioning it within a network that modulates the fate of Hsp70 client proteins.
Disruption of DNAJB14 in HEK293T cells creates a defined cellular context for dissecting the contribution of this specific co-chaperone to Hsp70 biology. Without endogenous DNAJB14 activity, researchers can delineate substrate specificity, assess compensatory mechanisms among Hsp40 family members, and probe the impact on downstream Hsp70 client protein handling. This model is particularly valuable for investigating endoplasmic reticulum protein quality control, given the epithelial secretory phenotype of HEK293T cells and the proposed involvement of DNAJB14 in ER-associated processes.
This polyclonal knockout population is ideally suited for a broad range of assays, including Western blotting and RT-qPCR for knockout confirmation, co-immunoprecipitation with Hsp70 to examine protein?Cprotein interactions, and cell viability measurements under proteotoxic stress induced by agents that cause protein misfolding. Fluorescence-based aggregation assays can further quantify the model??s impact on the partitioning of misfolded proteins. Applications span protein quality control studies, chaperone function analysis, cellular stress response modulation, and proteostasis research, with relevance to cancer, neurodegeneration, and protein conformational disorders. For further details and technical support, please contact Ascent Research.
