CHIP/STUB1 CRISPR Activation Plasmid (h): sc-400815-ACT

STUB1 encodes the E3 ubiquitin ligase CHIP, a co-chaperone that couples HSP70/HSP90 client recognition to ubiquitin-dependent protein turnover via its tetratricopeptide repeat (TPR) and U-box domains. CHIP/STUB1 regulates proteostasis by coordinating protein quality control, chaperone-mediated refolding decisions, and degradation through the ubiquitin–proteasome system, with downstream effects on cellular stress responses and mitochondrial and ER homeostasis. Through these processes, STUB1 influences pathways linked to protein aggregation and neuronal maintenance, and altered CHIP/STUB1 activity has been associated with neurodegeneration phenotypes and ataxia-related disease mechanisms. Its central role in controlling misfolded protein burden makes STUB1 a useful node for studying stress signaling and proteome stability in human cell models.

CHIP/STUB1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous STUB1 expression without altering the underlying DNA sequence.

CHIP/STUB1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the STUB1 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 STUB1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CHIP/STUB1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native STUB1 locus and enabling the study of CHIP/STUB1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CHIP/STUB1 pathway restoration in tumor cells with silenced or reduced STUB1 expression.

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