P2P-R CRISPR Activation Plasmid (h): sc-404165-ACT

Human RBBP6 encodes the P2P-R protein, a multi-domain factor that associates with p53 and retinoblastoma pathway components to influence cell-cycle progression, apoptosis, and transcriptional control. RBBP6 contains motifs consistent with roles in ubiquitin-dependent protein regulation and RNA processing, linking it to proteostasis and mRNA maturation programs that coordinate proliferation. Altered RBBP6 activity has been reported across tumor biology contexts, where perturbations in p53/RB signaling and checkpoint control are common, making it relevant for mechanistic studies of oncogenic stress responses. In cellular models, modulation of P2P-R levels can be used to interrogate pathway crosstalk between DNA damage signaling, transcriptional regulation, and post-transcriptional RNA metabolism.

P2P-R CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RBBP6 expression without altering the underlying DNA sequence.

P2P-R CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RBBP6 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 RBBP6 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous P2P-R expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RBBP6 locus and enabling the study of P2P-R-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of P2P-R pathway restoration in tumor cells with silenced or reduced RBBP6 expression.

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