TRB-3 CRISPR Activation Plasmid (m): sc-432803-ACT

Mouse Trib3 encodes TRB-3, a catalytically inactive pseudokinase that functions as an adaptor regulating signal transduction, transcriptional programs, and protein turnover. TRB-3 is best known for modulating PI3K–AKT signaling via interaction with AKT and related pathway components, thereby influencing glucose and lipid metabolism, cell survival decisions, and stress-responsive gene expression. Trib3 is induced by endoplasmic reticulum stress and integrated stress response pathways, linking nutrient availability and proteostasis to cellular adaptation. Dysregulated TRB-3 activity has been associated with insulin resistance, metabolic inflammation, and context-dependent effects on proliferation and apoptosis, supporting its use as a mechanistic node in metabolic and stress biology studies.

TRB-3 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Trib3 expression without altering the underlying DNA sequence.

TRB-3 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Trib3 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 Trib3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous TRB-3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Trib3 locus and enabling the study of TRB-3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of TRB-3 pathway restoration in tumor cells with silenced or reduced Trib3 expression.

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