TRIM32 CRISPR Activation Plasmid (h): sc-402731-ACT

TRIM32 encodes an E3 ubiquitin ligase of the TRIM family that regulates protein homeostasis by catalyzing substrate ubiquitination and coordinating proteasomal turnover. Through its RING, B-box, coiled-coil, and NHL repeat domains, TRIM32 influences myogenic differentiation, cytoskeletal organization, and stress-responsive signaling, with reported links to pathways including NF-κB and autophagy-related quality control. In human biology, altered TRIM32 function is associated with neuromuscular phenotypes such as limb-girdle muscular dystrophy and congenital myopathy, and dysregulated expression has been studied in tumor-relevant contexts where proteostasis and transcriptional programs are remodeled. These features make TRIM32 a useful node for dissecting ubiquitin-dependent regulation, muscle cell state transitions, and genotype–phenotype relationships in cellular models.

TRIM32 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TRIM32 expression without altering the underlying DNA sequence.

TRIM32 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TRIM32 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 TRIM32 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous TRIM32 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TRIM32 locus and enabling the study of TRIM32-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of TRIM32 pathway restoration in tumor cells with silenced or reduced TRIM32 expression.

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