CTLA-4 CRISPR Activation Plasmid (h): sc-400948-ACT

Human CTLA4 encodes CTLA-4 (CD152), an inhibitory immune checkpoint receptor predominantly expressed on activated T cells and regulatory T cells that restrains T cell activation by competing with CD28 for CD80/CD86 binding on antigen-presenting cells. Through attenuation of co-stimulatory signaling, CTLA-4 modulates downstream pathways including PI3K–AKT, NF-κB, and MAPK, helping maintain peripheral tolerance and limiting excessive cytokine production. Dysregulated CTLA-4 expression or signaling is linked to breakdown of immune homeostasis and altered lymphocyte responses implicated in autoimmune and inflammatory phenotypes. As a key node in immunoregulatory circuits, CTLA4 is widely studied in T cell functional states, antigen-driven activation dynamics, and mechanisms of immune suppression.

CTLA-4 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CTLA4 expression without altering the underlying DNA sequence.

CTLA-4 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CTLA4 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 CTLA4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CTLA-4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CTLA4 locus and enabling the study of CTLA-4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CTLA-4 pathway restoration in tumor cells with silenced or reduced CTLA4 expression.

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