PAK4 CRISPR Activation Plasmid (h): sc-401895-ACT

PAK4 (p21-activated kinase 4) is a serine/threonine kinase that functions downstream of Cdc42/Rac GTPases to coordinate cytoskeletal remodeling, cell polarity, and adhesion dynamics. It modulates actin organization and focal adhesion turnover through phosphorylation of substrates that influence Rho family signaling, and it interfaces with MAPK and PI3K/AKT-associated networks affecting survival and motility. In human cells, altered PAK4 activity or expression has been linked to dysregulated proliferation, invasive phenotypes, and resistance to stress signals in multiple disease contexts. These properties make PAK4 a useful node for studying kinase signaling, migration/invasion programs, and transcriptional responses to cytoskeletal cues.

PAK4 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous PAK4 expression without altering the underlying DNA sequence.

PAK4 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the PAK4 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 PAK4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous PAK4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native PAK4 locus and enabling the study of PAK4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of PAK4 pathway restoration in tumor cells with silenced or reduced PAK4 expression.

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