PINK1 CRISPR Activation Plasmid (h): sc-400739-ACT

PINK1 (PTEN-induced putative kinase 1) encodes a mitochondria-targeted serine/threonine kinase that functions as a key sensor of mitochondrial damage and initiator of mitophagy. Upon loss of mitochondrial membrane potential, PINK1 accumulates on the outer mitochondrial membrane and promotes recruitment and activation of PRKN/Parkin, coordinating ubiquitin-dependent clearance of impaired organelles and remodeling of mitochondrial dynamics. This pathway interfaces with cellular stress responses, including oxidative stress signaling, proteostasis, and innate immune activation linked to mitochondrial dysfunction. Altered PINK1 activity has been implicated in neurodegeneration and other disorders characterized by defective mitochondrial quality control, making it a widely used target in studies of mitochondrial homeostasis.

PINK1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous PINK1 expression without altering the underlying DNA sequence.

PINK1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the PINK1 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 PINK1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous PINK1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native PINK1 locus and enabling the study of PINK1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of PINK1 pathway restoration in tumor cells with silenced or reduced PINK1 expression.

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