GRHL2 CRISPR Activation Plasmid (m): sc-434250-ACT

Grhl2 encodes the mouse transcription factor GRHL2, a Grainyhead-like regulator of epithelial differentiation, apical–basal polarity, and barrier formation. GRHL2 coordinates gene programs involved in adherens junction assembly, tight junction integrity, and stratified epithelial development, linking cell–cell adhesion with proliferation and tissue morphogenesis. In diverse model systems, altered GRHL2 activity has been connected to dysregulated epithelial state transitions, including processes related to epithelial–mesenchymal plasticity, wound repair, and organogenesis. These functions make Grhl2 a useful node for studying epithelial lineage specification and pathways that influence epithelial homeostasis and disease-associated remodeling.

GRHL2 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Grhl2 expression without altering the underlying DNA sequence.

GRHL2 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Grhl2 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 Grhl2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous GRHL2 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Grhl2 locus and enabling the study of GRHL2-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of GRHL2 pathway restoration in tumor cells with silenced or reduced Grhl2 expression.

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