NBR1 CRISPR Activation Plasmid (h): sc-402592-ACT

Human NBR1 (neighbor of BRCA1 gene 1) encodes a selective autophagy receptor that recognizes ubiquitinated cargo and links it to LC3-positive autophagosomes through LIR-dependent interactions. NBR1 cooperates with SQSTM1/p62 in aggrephagy, mitophagy, and proteostasis control, supporting turnover of protein aggregates and damaged organelles during cellular stress. Through these functions, NBR1 contributes to regulation of innate immune signaling, oxidative stress responses, and lysosome-dependent quality control. Dysregulated NBR1-associated autophagy has been implicated in pathways relevant to neurodegeneration, cancer biology, and inflammatory phenotypes, making it a useful node for mechanistic studies of autophagic flux and ubiquitin signaling.

NBR1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous NBR1 expression without altering the underlying DNA sequence.

NBR1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the NBR1 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 NBR1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous NBR1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native NBR1 locus and enabling the study of NBR1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of NBR1 pathway restoration in tumor cells with silenced or reduced NBR1 expression.

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