GSK-3α CRISPR Activation Plasmid (h): sc-400518-ACT

GSK3A encodes glycogen synthase kinase-3 alpha (GSK-3α), a serine/threonine kinase that integrates multiple signaling inputs to regulate phosphorylation-dependent control of metabolism, proliferation, and differentiation. GSK-3α functions in canonical Wnt/β-catenin signaling through regulation of β-catenin turnover and also intersects with insulin/PI3K–AKT signaling, where inhibitory AKT phosphorylation modulates its activity. Through these pathways, GSK-3α influences glycogen synthesis, cell cycle progression, and transcriptional programs linked to developmental patterning and cellular stress responses. Dysregulated GSK3A activity and downstream signaling have been associated with mechanisms relevant to cancer biology, neurodegeneration, and metabolic disease models.

GSK-3α CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous GSK3A expression without altering the underlying DNA sequence.

GSK-3α CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the GSK3A 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 GSK3A transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous GSK-3α expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native GSK3A locus and enabling the study of GSK-3α-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of GSK-3α pathway restoration in tumor cells with silenced or reduced GSK3A expression.

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