DAP10 CRISPR Activation Plasmid (h): sc-402426-ACT

HCST encodes DNAX activation protein 10 (DAP10), a transmembrane adaptor that couples NKG2D and related receptors to intracellular signaling in human cytotoxic lymphocytes. Through its YINM motif, DAP10 recruits PI3K and Grb2/Vav1 to promote Akt and MAPK pathway activation, supporting immune synapse formation, cytokine production, and cell-mediated cytotoxicity. This signaling axis regulates NK cell and CD8⁺ T cell surveillance of stressed, infected, or transformed cells and influences inflammatory crosstalk within the tumor microenvironment. Dysregulated HCST/DAP10 activity has been implicated in altered innate and adaptive immune responses relevant to cancer immunobiology, viral infection, and autoimmune inflammation.

DAP10 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous HCST expression without altering the underlying DNA sequence.

DAP10 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the HCST 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 HCST transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous DAP10 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native HCST locus and enabling the study of DAP10-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of DAP10 pathway restoration in tumor cells with silenced or reduced HCST expression.

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