CD40 CRISPR Activation Plasmid (h): sc-400524-ACT

Human CD40 encodes a TNF receptor superfamily co-stimulatory receptor expressed primarily on antigen-presenting cells, including B cells, dendritic cells, and macrophages, where it integrates signals from CD40L to shape adaptive immunity. CD40 engagement activates canonical and non-canonical NF-κB signaling, MAPK pathways, and PI3K/AKT-dependent programs that regulate antigen presentation, cytokine production, survival, and B cell proliferation and class-switch recombination. Through these pathways, CD40 influences germinal center dynamics and T cell–dependent humoral responses, and altered CD40 activity has been associated with immune dysregulation and inflammatory pathobiology. Dysregulated CD40 signaling is also relevant to tumor immunology and B cell malignancy research due to its role in cell activation states and microenvironmental communication.

CD40 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CD40 expression without altering the underlying DNA sequence.

CD40 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CD40 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 CD40 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CD40 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CD40 locus and enabling the study of CD40-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CD40 pathway restoration in tumor cells with silenced or reduced CD40 expression.

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