The DNAJB5 knockout MES-OV polyclonal cells represent a CRISPR/Cas9-edited polyclonal knockout cell population derived from the MES-OV murine embryonic fibroblast cell line, designed for targeted disruption of the DNAJB5 gene. This loss-of-function model abolishes DNAJB5 protein expression, providing a heterogeneous pool of edited cells that captures a range of CRISPR-mediated gene-disruption events. The polyclonal format enables robust functional genomics and drug-screening applications without the clonal limitations of single-cell-derived lines, offering a physiologically relevant system for investigating co-chaperone-dependent proteostasis.
MES-OV cells are a well-established mouse embryonic fibroblast line frequently employed as feeder layers to support stem cell growth through the secretion of growth factors and extracellular matrix components. Their fibroblast identity offers a pertinent context for modeling cellular stress responses, including proteotoxic stress, due to their robust proliferation and ease of genetic manipulation. As a host for CRISPR/Cas9 editing, MES-OV provides a reproducible background that facilitates high-throughput assays and detailed mechanistic studies of protein quality control networks.
DNAJB5, a J-domain co-chaperone, specifically stimulates the ATPase activity of Hsp70 chaperones such as HSPA1A and HSPA8, driving the chaperone cycle that mediates client binding and release. Functioning downstream of the transcription factor HSF1, which is activated by heat shock and proteotoxic stress, DNAJB5 recruits misfolded protein substrates to Hsp70 for refolding or, in collaboration with the E3 ubiquitin ligase STUB1 and co-chaperone BAG1, targets terminally misfolded clients for ubiquitin-dependent proteasomal degradation. This functional coupling links the Hsp70 system to the ubiquitin-proteasome pathway, positioning DNAJB5 as a critical node in the cellular protein quality control apparatus.
In the MES-OV feeder-layer context, DNAJB5 knockout disrupts proteostasis, impairing the cell??s ability to manage proteotoxic stress. Loss of DNAJB5 function abrogates efficient Hsp70-mediated refolding and promotes the accumulation of aggregation-prone proteins, particularly under HSF1-activating conditions. Given the supportive role of MES-OV cells in stem cell niches, compromised protein homeostasis may also alter the secretome, potentially impacting stem cell maintenance. This model thus enables investigation of both cell-autonomous and non-cell-autonomous consequences of proteostasis failure, making it valuable for studying feeder-layer dysfunction in regenerative biology.
These polyclonal DNAJB5 knockout cells support diverse research applications, including mechanistic dissection of the Hsp70 co-chaperone network via co-immunoprecipitation and proteasome activity assays, investigation of protein aggregation dynamics using immunofluorescence and biochemical aggregation assays, and screening for small-molecule modulators of proteostasis through flow cytometry-based phenotypic readouts. They are also suited for modeling neurodegenerative disease-relevant proteotoxicity, assessing heat shock response pathways by RT-qPCR and western blotting, and evaluating chaperone dysfunction in stem cell coculture systems. For additional information, custom modifications, or bulk orders, please contact Ascent Research.
