EP3 CRISPR Activation Plasmid (h): sc-402485-ACT

PTGER3 encodes the human prostaglandin E2 receptor EP3, a GPCR that couples predominantly to Gi to suppress adenylyl cyclase and reduce cAMP, with context-dependent coupling to other G proteins that can modulate MAPK signaling and intracellular Ca2+. EP3 participates in prostaglandin-mediated control of inflammation, vascular tone, platelet function, smooth muscle contractility, and neuronal signaling, integrating lipid mediator cues into cell-type-specific transcriptional and metabolic programs. Through these pathways, EP3 signaling is relevant to models of inflammatory regulation, cardiovascular and thrombotic biology, and neurophysiology, and it is frequently investigated in settings where prostanoid receptor balance influences tissue responses.

EP3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous PTGER3 expression without altering the underlying DNA sequence.

EP3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the PTGER3 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 PTGER3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous EP3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native PTGER3 locus and enabling the study of EP3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of EP3 pathway restoration in tumor cells with silenced or reduced PTGER3 expression.

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