ASK 1 CRISPR Activation Plasmid (h): sc-400658-ACT

MAP3K5 encodes apoptosis signal-regulating kinase 1 (ASK1), a stress-responsive MAP3K that integrates oxidative stress, endoplasmic reticulum stress, calcium flux, and inflammatory cues to propagate signaling through the JNK and p38 MAPK cascades. ASK1 activity is regulated by redox-dependent interactions with thioredoxin and other scaffolding/adaptor proteins, coupling upstream stress sensing to phosphorylation of downstream MAP2Ks and transcriptional programs that shape apoptosis, cytokine production, and innate immune responses. Dysregulated ASK1 signaling has been implicated in cellular injury contexts relevant to neurodegeneration, cardiometabolic stress, and chronic inflammation, where altered MAPK output can influence survival, remodeling, and inflammatory gene expression. Because ASK1 sits at a key node connecting stress inputs to MAPK effector pathways, it is frequently studied for its role in signaling network rewiring and context-dependent cell fate decisions.

ASK 1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous MAP3K5 expression without altering the underlying DNA sequence.

ASK 1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the MAP3K5 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 MAP3K5 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous ASK 1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native MAP3K5 locus and enabling the study of ASK 1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of ASK 1 pathway restoration in tumor cells with silenced or reduced MAP3K5 expression.

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