TRIM5α CRISPR Activation Plasmid (h): sc-409348-ACT

TRIM5α (tripartite motif-containing 5 alpha) is an interferon-inducible restriction factor that recognizes retroviral capsids and blocks infection through premature uncoating and downstream inhibition of reverse transcription. As an E3 ubiquitin ligase with a RING-B-box-coiled-coil architecture and a capsid-binding PRY/SPRY domain, TRIM5α couples pattern recognition to ubiquitin-dependent signaling and proteostasis, intersecting with innate immune pathways such as NF-κB and inflammatory cytokine responses. Variation in TRIM5 influences host range and susceptibility to retroviruses, making it a widely studied node in antiviral defense, immune regulation, and host–pathogen coevolution. Dysregulated innate sensing and ubiquitin signaling involving TRIM family proteins is also relevant to immune-mediated pathology and cellular stress responses.

TRIM5α CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TRIM5 expression without altering the underlying DNA sequence.

TRIM5α CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TRIM5 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 TRIM5 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous TRIM5α expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TRIM5 locus and enabling the study of TRIM5α-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of TRIM5α pathway restoration in tumor cells with silenced or reduced TRIM5 expression.

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