FOXP3 CRISPR Activation Plasmid (h): sc-400542-ACT

FOXP3 encodes a forkhead family transcription factor that serves as a lineage-defining regulator of CD4⁺ regulatory T cells, coordinating immune tolerance and limiting excessive inflammatory responses. By shaping transcriptional programs downstream of T cell receptor signaling and cytokine pathways such as IL-2/STAT5 and TGF-β/SMAD, FOXP3 modulates effector cytokine production, metabolic remodeling, and suppressive function. Dysregulated FOXP3 expression or activity is linked to breakdown of peripheral tolerance and broad immune pathology, and FOXP3-positive tumor-infiltrating T cells are studied for roles in immunosuppressive microenvironments. These features make FOXP3 a key node for dissecting gene regulatory networks controlling immune homeostasis.

FOXP3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous FOXP3 expression without altering the underlying DNA sequence.

FOXP3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the FOXP3 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 FOXP3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous FOXP3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native FOXP3 locus and enabling the study of FOXP3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of FOXP3 pathway restoration in tumor cells with silenced or reduced FOXP3 expression.

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