TRAIL CRISPR Activation Plasmid (h): sc-418124-ACT

Human TNFSF10 encodes TRAIL (TNF-related apoptosis-inducing ligand), a type II membrane protein that can be released as a soluble cytokine and engage death receptors to initiate extrinsic apoptosis. TRAIL signaling activates caspase-8 and downstream effector caspases, while intersecting with NF-κB, JNK, and mitochondrial amplification pathways to shape cell fate decisions. TNFSF10/TRAIL activity influences immune surveillance, inflammatory crosstalk, and the regulation of tissue homeostasis, with dysregulation linked to altered apoptosis sensitivity and tumor–immune interactions. As a pathway node, TRAIL is frequently studied in contexts of cancer cell death programs, resistance mechanisms, and immune-mediated cytotoxicity.

TRAIL CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TNFSF10 expression without altering the underlying DNA sequence.

TRAIL CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TNFSF10 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 TNFSF10 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous TRAIL expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TNFSF10 locus and enabling the study of TRAIL-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of TRAIL pathway restoration in tumor cells with silenced or reduced TNFSF10 expression.

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