HLA-DRβ5 CRISPR Activation Plasmid (h): sc-402985-ACT

HLA-DRB5 encodes the HLA-DRβ5 chain, which pairs with HLA-DRA to form MHC class II HLA-DR heterodimers that present extracellularly derived peptides to CD4+ T cells. This antigen presentation program is central to adaptive immune activation, thymic selection, and peripheral immune tolerance, and is regulated by the class II transactivator (CIITA) and interferon-γ–responsive transcriptional networks. HLA-DRB5 expression is most prominent in professional antigen-presenting cells and can be induced in other cell types during inflammation, influencing T cell priming and cytokine signaling outcomes. Variation in HLA class II haplotypes and dysregulated MHC II expression are widely studied in the context of autoimmunity, transplant immunobiology, and tumor immune microenvironments.

HLA-DRβ5 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous HLA-DRB5 expression without altering the underlying DNA sequence.

HLA-DRβ5 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the HLA-DRB5 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 HLA-DRB5 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous HLA-DRβ5 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native HLA-DRB5 locus and enabling the study of HLA-DRβ5-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of HLA-DRβ5 pathway restoration in tumor cells with silenced or reduced HLA-DRB5 expression.

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