VCP CRISPR Activation Plasmid (h): sc-401052-ACT

Human valosin-containing protein (VCP, p97) is an AAA+ ATPase that drives ubiquitin-dependent protein quality control by extracting and remodeling ubiquitinated substrates for proteasomal degradation and organelle maintenance. VCP functions centrally in ER-associated degradation, autophagy and lysosomal homeostasis, cell cycle regulation, and the DNA damage response through coordinated action with cofactors such as UFD1–NPL4. By controlling proteostasis and stress-adaptive pathways, VCP helps maintain mitochondrial and ER integrity and regulates signaling linked to inflammatory and apoptotic programs. Dysregulated VCP activity is associated with multisystem proteinopathy phenotypes, including neurodegeneration and myopathy, and is frequently investigated in models of proteotoxic stress and cancer cell fitness.

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

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

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