PORCN CRISPR Activation Plasmid (h): sc-413044-ACT

Human PORCN encodes a membrane-bound O-acyltransferase in the endoplasmic reticulum that catalyzes palmitoleoylation of WNT ligands, a modification required for WNT secretion, extracellular gradient formation, and receptor engagement. Through this essential processing step, PORCN regulates canonical and non-canonical WNT signaling, influencing β-catenin–dependent transcription, cell fate specification, proliferation, and tissue patterning. Altered PORCN-dependent WNT output is implicated in developmental abnormalities and diseases characterized by dysregulated WNT pathway activity, supporting its use in mechanistic studies of differentiation programs and signaling network rewiring. PORCN function is also relevant to studies of epithelial homeostasis, stem cell biology, and pathway crosstalk affecting morphogenesis and oncogenic signaling states.

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

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

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