AMPKγ3 CRISPR Activation Plasmid (h): sc-403280-ACT

PRKAG3 encodes the AMPKγ3 regulatory subunit of AMP-activated protein kinase (AMPK), a central energy sensor that integrates AMP/ADP:ATP status to coordinate metabolic adaptation. By modulating AMPK complex activation and substrate targeting, AMPKγ3 influences pathways controlling glucose uptake, glycogen metabolism, mitochondrial function, and lipid homeostasis through downstream nodes such as mTORC1 and autophagy regulators. Although best characterized in striated muscle, altered AMPK signaling involving PRKAG3 is relevant to cellular stress responses that intersect with insulin sensitivity, metabolic remodeling, and inflammation-associated signaling. Dysregulated AMPK pathway activity is frequently examined in contexts including metabolic disease mechanisms and tumor cell nutrient stress, where PRKAG3-dependent regulation can shape bioenergetic phenotypes.

AMPKγ3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous PRKAG3 expression without altering the underlying DNA sequence.

AMPKγ3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the PRKAG3 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 PRKAG3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous AMPKγ3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native PRKAG3 locus and enabling the study of AMPKγ3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of AMPKγ3 pathway restoration in tumor cells with silenced or reduced PRKAG3 expression.

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