TRAF6 CRISPR Activation Plasmid (h): sc-400117-ACT

TRAF6 (TNF receptor–associated factor 6) is a RING-type E3 ubiquitin ligase and adaptor protein that transduces signals from members of the TNF receptor superfamily and Toll-like/IL-1 receptors. Through K63-linked polyubiquitination of itself and partner proteins, TRAF6 promotes activation of TAK1 and downstream NF-κB and MAPK pathways, shaping inflammatory gene expression, innate immune responses, and cell survival programs. TRAF6 also interfaces with PI3K–AKT signaling and contributes to osteoclast differentiation via RANK–RANKL signaling, linking immune regulation to bone remodeling. Dysregulated TRAF6 signaling has been associated with chronic inflammation, immune dysfunction, and oncogenic pathway activation in multiple tumor contexts, supporting its use as a mechanistic node in pathway-focused studies.

TRAF6 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TRAF6 expression without altering the underlying DNA sequence.

TRAF6 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TRAF6 locus in human cell lines. The system is built around a catalytically inactive Cas9 (dCas9) carrying two inactivating mutations (D10A and N863A) that eliminate nuclease activity while preserving DNA binding. This dCas9 is fused to VP64, a potent transcriptional activator, and is co-expressed with a blasticidin resistance gene for selection. The second plasmid encodes the MS2-p65-HSF1 fusion protein, a secondary activator complex that works in concert with dCas9-VP64, alongside a hygromycin resistance gene. The third plasmid encodes a target-specific 20 nt sgRNA fused to two MS2 RNA aptamers that recruit the MS2-p65-HSF1 complex to the activation site, accompanied by a puromycin resistance gene. The three plasmids are delivered at a 1:1:1 mass ratio for balanced expression of all system components.

Once assembled at the target locus, the SAM complex binds within approximately 200 bp upstream of the TRAF6 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous TRAF6 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TRAF6 locus and enabling the study of TRAF6-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of TRAF6 pathway restoration in tumor cells with silenced or reduced TRAF6 expression.

For Research Use Only. Not Intended for Diagnostic or Therapeutic Use.