CD154 CRISPR Activation Plasmid (h): sc-401682-ACT

CD40LG encodes CD154 (CD40 ligand), a type II transmembrane protein transiently displayed on activated CD4+ T cells and other immune subsets that provides essential costimulatory signals to CD40-expressing B cells, dendritic cells, and macrophages. CD154–CD40 engagement promotes germinal center formation, immunoglobulin class-switch recombination, and dendritic cell licensing, integrating antigen presentation with adaptive immune polarization. Downstream signaling involves TRAF-dependent activation of NF-κB, MAPK, and PI3K pathways that regulate cytokine production, survival, and differentiation. Dysregulated CD40LG/CD154 activity is linked to altered humoral immunity and inflammatory immune phenotypes, supporting its relevance in studies of immune regulation and autoimmunity-associated mechanisms.

CD154 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CD40LG expression without altering the underlying DNA sequence.

CD154 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CD40LG 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 CD40LG transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CD154 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CD40LG locus and enabling the study of CD154-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CD154 pathway restoration in tumor cells with silenced or reduced CD40LG expression.

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