P2Y1 CRISPR Activation Plasmid (m): sc-422095-ACT

Mouse P2ry1 encodes the purinergic receptor P2Y1, a G protein–coupled receptor activated by extracellular ADP that couples primarily to Gq/11 to stimulate phospholipase C signaling, inositol phosphate production, and intracellular Ca2+ mobilization. P2Y1 contributes to purinergic neurotransmission, platelet-like aggregation pathways, vascular tone regulation, and immune cell activation by integrating nucleotide release with downstream second-messenger networks. Through cross-talk with MAPK and other calcium-dependent pathways, P2Y1 modulates secretion, migration, and excitability in diverse tissues. Dysregulated purinergic signaling involving P2Y1 has been implicated in thrombosis and hemostasis-related phenotypes, inflammatory responses, and neurovascular dysfunction, supporting its relevance for mechanistic disease modeling.

P2Y1 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous P2ry1 expression without altering the underlying DNA sequence.

P2Y1 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the P2ry1 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 P2ry1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous P2Y1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native P2ry1 locus and enabling the study of P2Y1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of P2Y1 pathway restoration in tumor cells with silenced or reduced P2ry1 expression.

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