The Kdm4a Knockout RAW 264.7 Cell Line is a stable CRISPR/Cas9-edited murine macrophage model that carries a targeted disruption of the Kdm4a gene, generating a loss-of-function system for the KDM4A histone demethylase. This product is supplied as a ready-to-use cell line derived from the well-characterized RAW 264.7 monocyte/macrophage background. It enables precise investigation of KDM4A-dependent epigenetic regulation, chromatin modification, and downstream signaling pathways in innate immune cells.
The parental RAW 264.7 cell line originates from an Abelson murine leukemia virus-induced tumor and exhibits adherent, macrophage-like morphology. These cells are phagocytic and respond to inflammatory stimuli by secreting key cytokines such as tumor necrosis factor (TNF) and interleukin-6 (IL-6). As a widely adopted model for studying macrophage biology, RAW 264.7 cells are extensively used in research on innate immunity, host defense, and cytokine signaling. Their genetic tractability makes them an ideal host for interrogating gene function through CRISPR/Cas9-mediated editing.
KDM4A is a histone demethylase that removes methyl groups from H3K9me2/3 and H3K36me2/3, thereby modulating chromatin accessibility and gene expression. It operates downstream of HIF-1?? stabilization, MAPK cascades, and JAK-STAT pathways. KDM4A physically interacts with NF-??B, HDACs, and nuclear receptors such as the androgen receptor, forming complexes that link extracellular signals to chromatin. It demethylates histones at the promoters of NF-??B-regulated genes (e.g., TNF, IL6) and HIF-1?? target genes, facilitating their transcriptional activation. Consequently, KDM4A serves as a critical integrator of inflammatory, hypoxic, and growth factor signals to control cytokine output and stress responses in macrophages.
Disruption of Kdm4a in RAW 264.7 cells is anticipated to alter the histone methylation landscape, particularly at H3K9 and H3K36 residues, impacting the expression of key inflammatory mediators and stress-response genes. This model provides a critical tool for deciphering how KDM4A-driven chromatin dynamics govern macrophage polarization between pro-inflammatory and anti-inflammatory states. It is particularly valuable for studying the epigenetic basis of chronic inflammation, tumor-associated macrophage reprogramming, and metabolic disorders where macrophage plasticity is decisive. The knockout enables dissection of KDM4A??s specific contributions amid the complex regulatory networks active in activated macrophages.
This knockout cell line supports a diverse array of experimental approaches. Standard methods such as western blotting and RT-qPCR can confirm Kdm4a disruption and quantify changes in downstream targets. Chromatin immunoprecipitation with qPCR (ChIP-qPCR) for H3K9me3 and H3K36me3 reveals locus-specific alterations in histone methylation. Transcriptome-wide analysis via RNA-seq uncovers global gene expression shifts, while ELISA assays measure secretion of cytokines like TNF-?? and IL-6. Functional studies employing phagocytosis assays and flow cytometry for macrophage surface markers further characterize the knockout phenotype. The cell line is also amenable to high-throughput screening of small-molecule KDM4A inhibitors. For further technical information or custom applications, please contact Ascent Research.
