USP21 CRISPR Activation Plasmid (h): sc-404690-ACT

Human USP21 encodes a ubiquitin-specific protease that removes ubiquitin from protein substrates to regulate protein stability, signaling amplitude, and subcellular trafficking. USP21 has been implicated in controlling inflammatory and innate immune signaling by modulating ubiquitin-dependent steps within NF-κB and interferon-associated pathways, and it can influence transcriptional programs through deubiquitination of chromatin-associated factors. By shaping ubiquitin homeostasis, USP21 impacts cell-cycle progression, stress responses, and differentiation-related processes. Dysregulated USP21 activity has been associated with aberrant immune signaling and oncogenic phenotypes, making it a useful target for mechanistic studies of ubiquitin-driven regulation in human cells.

USP21 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous USP21 expression without altering the underlying DNA sequence.

USP21 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the USP21 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 USP21 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous USP21 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native USP21 locus and enabling the study of USP21-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of USP21 pathway restoration in tumor cells with silenced or reduced USP21 expression.

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