CD274/PD-L1 CRISPR Activation Plasmid (h): sc-401140-ACT

CD274 encodes the immune checkpoint ligand PD-L1, a cell-surface glycoprotein that binds PD-1 (PDCD1) on activated T cells to attenuate TCR signaling, cytokine production, and cytotoxic function. PD-L1 expression is regulated by inflammatory and oncogenic cues, including IFN-γ–JAK/STAT signaling and pathways such as PI3K/AKT and NF-κB, shaping adaptive immune resistance in tissue microenvironments. Dysregulated CD274/PD-L1 contributes to immune evasion across multiple cancers and is also implicated in chronic infection and autoimmune-associated immune regulation through modulation of antigen presentation and T cell exhaustion programs. As a measurable node in immune signaling, PD-L1 is frequently studied in tumor–immune co-culture systems, interferon response assays, and pathway dissection of checkpoint regulation.

CD274/PD-L1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CD274 expression without altering the underlying DNA sequence.

CD274/PD-L1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CD274 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 CD274 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CD274/PD-L1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CD274 locus and enabling the study of CD274/PD-L1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CD274/PD-L1 pathway restoration in tumor cells with silenced or reduced CD274 expression.

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