ATP5B CRISPR Activation Plasmid (h): sc-401009-ACT

ATP5B encodes the beta subunit of mitochondrial ATP synthase (Complex V), a core catalytic component of the F1 sector that couples proton-motive force to ATP production during oxidative phosphorylation. By supporting cellular bioenergetics and mitochondrial membrane potential maintenance, ATP5B influences processes such as reactive oxygen species homeostasis, apoptosis susceptibility, and metabolic adaptation. Altered expression or function of ATP synthase subunits, including ATP5B, has been associated with mitochondrial dysfunction phenotypes observed across neurodegeneration, cardiometabolic stress, and cancer-related metabolic reprogramming, making it a common readout in studies of mitochondrial biology. ATP5B is therefore frequently interrogated in pathways linking electron transport chain activity to ATP demand, mitochondrial quality control, and stress response signaling.

ATP5B CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ATP5B expression without altering the underlying DNA sequence.

ATP5B CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ATP5B 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 ATP5B transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous ATP5B expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ATP5B locus and enabling the study of ATP5B-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of ATP5B pathway restoration in tumor cells with silenced or reduced ATP5B expression.

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