PNUTS CRISPR Activation Plasmid (h): sc-403688-ACT

PPP1R10 encodes PNUTS (protein phosphatase 1 regulatory subunit 10), a nuclear targeting subunit that binds PP1 and modulates phosphatase activity on chromatin-associated substrates. PNUTS participates in transcriptional control, DNA damage response signaling, and cell-cycle progression by coordinating protein dephosphorylation with genome maintenance pathways. It has been linked to regulation of RNA polymerase II–dependent transcription and to processes affecting telomere stability and replication stress. Altered PPP1R10/PNUTS activity has been associated with dysregulated proliferation and genome instability phenotypes relevant to cancer biology and other disorders involving impaired DNA repair and nuclear signaling.

PNUTS CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous PPP1R10 expression without altering the underlying DNA sequence.

PNUTS CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the PPP1R10 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 PPP1R10 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous PNUTS expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native PPP1R10 locus and enabling the study of PNUTS-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of PNUTS pathway restoration in tumor cells with silenced or reduced PPP1R10 expression.

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