USP45 CRISPR Activation Plasmid (m2): sc-429548-ACT-2

Mouse Usp45 encodes USP45, a ubiquitin-specific protease that removes ubiquitin modifications from target proteins to regulate their stability and signaling output. USP45 is implicated in the DNA damage response, supporting genome maintenance through deubiquitination events that influence repair pathway choice and replication stress tolerance, including coordination with post-translational modification networks such as ubiquitin and SUMO signaling. Disruption of USP45-mediated deubiquitination can perturb chromatin-associated repair processes, elevate genomic instability, and has been linked to phenotypes relevant to cancer biology and developmental defects. Usp45-targeted gene editing and functional genomics studies in mouse cells enable mechanistic dissection of ubiquitin-dependent DNA repair circuitry, identification of USP45 substrates and interaction partners, and modeling of genotype–phenotype relationships in genome integrity pathways.

USP45 CRISPR Activation Plasmid (m2) provides a targeted, non-destructive approach to upregulating endogenous Usp45 expression without altering the underlying DNA sequence.

USP45 CRISPR Activation Plasmid (m2) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Usp45 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 Usp45 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous USP45 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Usp45 locus and enabling the study of USP45-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of USP45 pathway restoration in tumor cells with silenced or reduced Usp45 expression.

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