GalNAc-T3 CRISPR Activation Plasmid (h): sc-407682-ACT

GALNT3 encodes polypeptide N-acetylgalactosaminyltransferase 3 (GalNAc-T3), a Golgi-resident initiating enzyme for mucin-type O-linked glycosylation that transfers GalNAc to serine/threonine residues on secretory and membrane proteins. By shaping O-glycan initiation sites, GalNAc-T3 influences protein folding, trafficking, protease susceptibility, and receptor/adhesion molecule function across epithelial and secretory pathways. Altered GALNT3 activity has been linked to dysregulated glycoprotein processing and extracellular signaling, with relevance to disorders of phosphate homeostasis and broader glycosylation-associated phenotypes. Modulation of GalNAc-T3 is therefore useful for dissecting how O-glycosylation controls cell–cell interactions, secretory programs, and ligand–receptor communication.

GalNAc-T3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous GALNT3 expression without altering the underlying DNA sequence.

GalNAc-T3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the GALNT3 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 GALNT3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous GalNAc-T3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native GALNT3 locus and enabling the study of GalNAc-T3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of GalNAc-T3 pathway restoration in tumor cells with silenced or reduced GALNT3 expression.

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