PYPAF7 CRISPR Activation Plasmid (m): sc-436175-ACT

Mouse Nlrp12 (PYPAF7) encodes a NOD-like receptor that functions as an innate immune sensor and regulator of inflammatory signaling. It contributes to the control of NF-κB and MAPK pathway outputs downstream of pattern-recognition and cytokine receptors, shaping myeloid activation, chemokine expression, and leukocyte trafficking. Through its role in inflammasome-associated processes and negative regulation of excessive inflammation, altered Nlrp12 activity has been linked to dysregulated immune homeostasis and inflammatory phenotypes in experimental disease models. These properties make Nlrp12 a useful node for studying innate immune checkpoints, microbiota-responsive inflammation, and signaling crosstalk that influences tissue inflammation.

PYPAF7 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Nlrp12 expression without altering the underlying DNA sequence.

PYPAF7 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Nlrp12 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 Nlrp12 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous PYPAF7 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Nlrp12 locus and enabling the study of PYPAF7-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of PYPAF7 pathway restoration in tumor cells with silenced or reduced Nlrp12 expression.

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