VSTM3 CRISPR Activation Plasmid (h): sc-404147-ACT

TIGIT, also known as VSTM3, encodes an inhibitory immunoreceptor expressed on activated T cells, regulatory T cells, and NK cells that helps maintain immune homeostasis. By binding poliovirus receptor family ligands such as PVR (CD155) and PVRL2 (CD112), TIGIT modulates ITIM-like signaling and counterbalances costimulatory pathways, shaping cytotoxicity, cytokine production, and immune synapse function. Altered TIGIT signaling is implicated in immune evasion and dysfunctional antitumor immunity as well as chronic inflammatory and autoimmune contexts, making it a key node in immune checkpoint biology. As a surface receptor integrating adhesion and inhibitory cues, TIGIT provides a mechanistic entry point to study receptor–ligand interactions, transcriptional programs of exhausted lymphocytes, and cross-talk with CD226/DNAM-1 signaling.

VSTM3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TIGIT expression without altering the underlying DNA sequence.

VSTM3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TIGIT 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 TIGIT transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous VSTM3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TIGIT locus and enabling the study of VSTM3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of VSTM3 pathway restoration in tumor cells with silenced or reduced TIGIT expression.

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