NIMP CRISPR Activation Plasmid (h): sc-413705-ACT

RTN4IP1 encodes NIMP, a mitochondrial matrix protein that interacts with the NOGO receptor pathway component RTN4 and supports oxidative phosphorylation by maintaining respiratory chain function and redox balance. NIMP contributes to mitochondrial bioenergetics, cofactor homeostasis, and protection from oxidative stress, linking its activity to ATP production and reactive oxygen species management. Altered RTN4IP1 function has been associated with mitochondrial dysfunction phenotypes, including neurodegenerative and optic neuropathy presentations, making it relevant for studies of mitochondrial quality control and stress-response signaling. In human cell models, modulation of RTN4IP1 expression is used to interrogate pathways connecting mitochondrial metabolism to neuronal maintenance and disease-relevant cellular vulnerability.

NIMP CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RTN4IP1 expression without altering the underlying DNA sequence.

NIMP CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RTN4IP1 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 RTN4IP1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous NIMP expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RTN4IP1 locus and enabling the study of NIMP-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of NIMP pathway restoration in tumor cells with silenced or reduced RTN4IP1 expression.

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