Pyrin CRISPR Activation Plasmid (h): sc-403193-ACT

MEFV encodes pyrin, a cytosolic pattern-recognition protein that regulates inflammasome signaling and innate immune homeostasis in human myeloid cells. Pyrin senses pathogen- or danger-associated perturbations that impact Rho GTPase signaling and cytoskeletal dynamics, coordinating assembly of inflammasome complexes that control caspase-1 activity and downstream processing of IL-1 family cytokines. Through these pathways, pyrin helps tune inflammatory thresholding and programmed inflammatory responses, linking cytoskeletal surveillance to NF-κB and cytokine signaling networks. Dysregulated MEFV/pyrin activity is genetically associated with autoinflammatory phenotypes, making it a frequently studied node in inflammatory signaling and host defense research.

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

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

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