DNAM-1 CRISPR Activation Plasmid (h): sc-402189-ACT

CD226 encodes DNAM-1, an immunoglobulin superfamily adhesion and co-stimulatory receptor expressed on NK cells, CD8+ T cells, and subsets of CD4+ T cells and monocytes. DNAM-1 binds the ligands CD155 (PVR) and CD112 (NECTIN2) to promote immune synapse formation, cytotoxic granule polarization, and cytokine release, integrating with signaling networks that shape lymphocyte activation and target cell recognition. Through its role in balancing activating and inhibitory inputs, CD226 influences antitumor immune surveillance and responses to viral infection. Genetic variation and dysregulated expression of CD226 have been associated with immune-mediated disease susceptibility, supporting its use as a research node for studying inflammatory pathways and immune cell functional heterogeneity.

DNAM-1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CD226 expression without altering the underlying DNA sequence.

DNAM-1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CD226 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 CD226 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous DNAM-1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CD226 locus and enabling the study of DNAM-1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of DNAM-1 pathway restoration in tumor cells with silenced or reduced CD226 expression.

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