PIRT CRISPR Activation Plasmid (h): sc-416125-ACT

Human PIRT (phosphoinositide-interacting regulator of TRP channels) encodes a small membrane-associated protein that binds phosphatidylinositol 4,5-bisphosphate (PIP2) and modulates the activity of select transient receptor potential (TRP) ion channels. Through regulation of TRP-dependent calcium and cation flux, PIRT contributes to sensory signaling processes including nociception, thermosensation, and itch, linking membrane phosphoinositide dynamics to neuronal excitability. PIRT-associated signaling intersects with pathways governing GPCR- and lipid-mediated channel gating, impacting downstream second messenger responses and activity-dependent transcriptional programs. Altered TRP channel regulation and phosphoinositide signaling have been implicated in pain and neuroinflammatory phenotypes, making PIRT a useful node for mechanistic studies in sensory neuron biology and related disease models.

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

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

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