PBEF CRISPR Activation Plasmid (h): sc-416248-ACT

NAMPT, also known as PBEF, encodes nicotinamide phosphoribosyltransferase, the rate-limiting enzyme in the NAD salvage pathway that converts nicotinamide to nicotinamide mononucleotide. By controlling intracellular NAD availability, PBEF influences redox homeostasis and supports NAD-dependent enzymes including sirtuins and PARPs, linking metabolic state to chromatin regulation, DNA repair, and stress responses. NAMPT activity intersects with mitochondrial function, inflammatory signaling, and cell-cycle control, and dysregulation has been associated with altered immunometabolism and tumor biology. As a multifunctional node in NAD metabolism, NAMPT is frequently studied in contexts of metabolic adaptation, oxidative stress, and inflammation-related cellular phenotypes.

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

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

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