Adenosine A2B-R CRISPR Activation Plasmid (h): sc-401402-ACT

ADORA2B encodes the human adenosine A2B receptor (A2B-R), a low-affinity, G protein–coupled receptor induced under hypoxia and inflammatory stress where extracellular adenosine accumulates. A2B-R signals primarily through Gs to elevate cAMP and can also couple to Gq to mobilize intracellular calcium, engaging downstream programs such as PKA/CREB, MAPK/ERK, and PI3K signaling. Through these pathways, ADORA2B modulates endothelial barrier function, angiogenic and fibrotic responses, and cytokine production in immune and stromal compartments. Dysregulated ADORA2B activity has been associated with chronic inflammatory states, hypoxia-driven tissue remodeling, and tumor microenvironment signaling, supporting its use as a mechanistic node for receptor signaling and microenvironment biology studies.

Adenosine A2B-R CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ADORA2B expression without altering the underlying DNA sequence.

Adenosine A2B-R CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ADORA2B 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 ADORA2B transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Adenosine A2B-R expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ADORA2B locus and enabling the study of Adenosine A2B-R-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Adenosine A2B-R pathway restoration in tumor cells with silenced or reduced ADORA2B expression.

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