USP32 CRISPR Activation Plasmid (m): sc-433591-ACT

Usp32 encodes the deubiquitinating enzyme USP32, a ubiquitin-specific protease that reverses ubiquitin conjugation to regulate protein stability, trafficking, and signal transduction. As part of ubiquitin-dependent quality control, USP32 is implicated in controlling proteostasis and turnover of pathway components that influence cell-cycle progression, stress responses, and membrane-associated signaling. Dysregulated deubiquitination has broad relevance to oncology and neurobiology by altering degradation of regulatory proteins and shifting pathway outputs. In mouse model systems, modulating USP32 activity supports mechanistic studies of ubiquitin signaling networks and their roles in disease-associated cellular phenotypes.

USP32 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Usp32 expression without altering the underlying DNA sequence.

USP32 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Usp32 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 Usp32 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous USP32 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Usp32 locus and enabling the study of USP32-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of USP32 pathway restoration in tumor cells with silenced or reduced Usp32 expression.

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