Pontin 52 CRISPR Activation Plasmid (h): sc-402304-ACT

Human RUVBL1 encodes Pontin 52, an AAA+ ATPase that functions in multi-protein assemblies regulating chromatin architecture, transcriptional control, and macromolecular complex biogenesis. Pontin 52 is a core component of the R2TP co-chaperone complex and supports HSP90-dependent maturation of several client complexes, linking it to processes such as DNA damage responses, chromatin remodeling, and ribonucleoprotein assembly. Through interactions with transcriptional regulators and remodeling factors, RUVBL1 influences cell-cycle progression and stress-adaptive gene expression programs. Dysregulated RUVBL1/Pontin 52 activity and altered expression have been associated with proliferative signaling and genome maintenance defects, making it a useful node for mechanistic studies in cancer-relevant pathways and proteostasis networks.

Pontin 52 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RUVBL1 expression without altering the underlying DNA sequence.

Pontin 52 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RUVBL1 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 RUVBL1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Pontin 52 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RUVBL1 locus and enabling the study of Pontin 52-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Pontin 52 pathway restoration in tumor cells with silenced or reduced RUVBL1 expression.

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