DUSP18 Knockout HEK293T Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population designed to disrupt the DUSP18 gene in the widely used HEK293T host cell line. This product provides a heterogeneous pool of cells carrying targeted gene disruptions, enabling loss-of-function studies without the constraints of clonal selection. The polyclonal format preserves population-level diversity while ensuring robust knockout representation across the cell pool, making it suitable for experiments where bulk cellular responses are assessed. Researchers can employ this model to investigate DUSP18-mediated regulation of stress-activated signaling cascades under physiologically relevant conditions.
HEK293T cells are human embryonic kidney epithelial cells that stably express the SV40 large T antigen, which promotes episomal replication of plasmids containing the SV40 origin of replication. This feature dramatically enhances protein expression and is a cornerstone of their utility in transient transfection, lentivirus/retrovirus production, and large-scale biochemical studies. The cell line is extensively characterized for its ease of manipulation, rapid growth, and responsiveness to a broad range of stimuli, including toll-like receptor ligands, inflammatory cytokines, and cellular stresses. Consequently, HEK293T is a preferred platform for dissecting signaling pathways, particularly those involving innate immunity and mitogen-activated protein kinase (MAPK) cascades.
DUSP18 encodes a dual-specificity phosphatase that specifically dephosphorylates phosphotyrosine and phosphoserine/threonine residues on stress-activated MAP kinases JNK and p38. In the canonical signaling hierarchy, upstream stimuli such as TLR activation by LPS or CpG, inflammatory cytokines (e.g., TNF, IL-1??), and reactive oxygen species trigger MAP3Ks like ASK1 and MEKK1. These kinases phosphorylate MAP2Ks including MKK4/MKK7 (for JNK) and MKK3/MKK6 (for p38), which in turn activate JNK and p38. Active JNK and p38 translocate to the nucleus to phosphorylate transcription factors such as c-Jun and ATF2, driving AP-1-dependent gene expression. DUSP18 directly binds to and dephosphorylates JNK and p38, serving as a critical negative feedback regulator that terminates signal propagation. Loss of DUSP18 thus disrupts this feedback, leading to sustained or amplified downstream signaling.
In the HEK293T cellular context, knockout of DUSP18 results in prolonged activation of JNK and p38 upon challenge with TLR agonists or stress inducers. This heightened MAPK activity enhances transcriptional programs governed by AP-1 and other stress-responsive factors, including those controlling inflammatory cytokines and apoptotic regulators. The HEK293T background allows researchers to exploit its efficient transfection capabilities to introduce reporters, mutant constructs, or additional pathway components, facilitating detailed mechanistic dissection. Because DUSP18 is itself implicated in inflammatory diseases, cancer, and autoimmune disorders, this model provides a tractable system for evaluating its role in these pathophysiological processes.
Typical research applications include western blotting for phospho-JNK and phospho-p38 to monitor kinase activation dynamics, RT-qPCR of AP-1 target genes (e.g., c-JUN, FOS, ATF3) to assess transcriptional outcomes, and dual-luciferase reporter assays for AP-1 activity. Flow cytometry using Annexin V/PI staining allows quantification of apoptosis under pro-inflammatory conditions, while co-immunoprecipitation experiments can probe DUSP18?CMAPK interactions in a clean knockout background. Additionally, the cells are suitable for high-throughput screening of small-molecule phosphatase inhibitors and for functional rescue experiments with wild-type or mutant DUSP18 constructs. For further technical details or ordering information, please contact Ascent Research.
